1.1.1.1 alcohol dehydrogenase synthesis enzyme can be used in preparative scale enantioselective oxidation of sec-alcohol in asymmetric reduction of ketones, using acetone and 2-propanol, respectively, as cosubstrates for cofactor-regeneration via a coupled-substrate approach 1.1.1.1 alcohol dehydrogenase synthesis production of (3R,5S)-6-benzyloxy-3,5-dihydroxy-hexanoic acid ethyl ester, which is a key chiral intermediate for anticholesterol drugs that act by inhibition of hydroxy methyl glutaryl coenzyme A reductase 1.1.1.1 alcohol dehydrogenase synthesis production of (4S,6S)-5,6-dihydro-4-hydroxy-6-methyl-4H-thieno[2,3b]thiopyran-7,7dioxide, which is an intermediate in the synthesis of the carbonic anhydrase inhibitor trusopt. Trusopt is a novel, topically active treatment for glaucoma 1.1.1.1 alcohol dehydrogenase synthesis production of (S)-1-Phenyl-2-propanol, which is used as an intermediate for the synthesis of amphetamines and as a precursor for anti-hypertensive agents and spasmolytics or anti-epileptics 1.1.1.1 alcohol dehydrogenase synthesis production of (S)-4-(3,4-methylenedioxyphenyl)-2-propanol, which is converted to LY300164, an orally active benzodiazepine 1.1.1.1 alcohol dehydrogenase synthesis LSADH catalyzed the enantioselective reduction of some ketones with high enantiomeric excesses: phenyl trifluoromethyl ketone to (S)-1-phenyltrifluoroethanol (>99% e.e.), acetophenone to (R)-1-phenylethanol (99% e.e.), and 2-heptanone to (R)-2-heptanol (>99% e.e.) in the presence of 2-propanol without an additional NADH regeneration system. Therefore, it would be a useful biocatalyst 1.1.1.1 alcohol dehydrogenase synthesis the photochemical and enzymatic synthesis of methanol from formaldehyde with alcohol dehydrogenase and NAD+ photoreduction by the visible-light photosensitization of zinc tetraphenylporphyrin tetrasulfonate in the presence of methylviologen, diaphorase, and triethanolamine is developed 1.1.1.1 alcohol dehydrogenase synthesis alcohol dehydrogenases represent an important group of biocatalysts due to their ability to stereospecifically reduce prochiral carbonyl compounds 1.1.1.1 alcohol dehydrogenase synthesis alpha-ketoisovalerate decarboxylase Kivd from Lactococcus lactis combined with alcohol dehydrogenase Adh3 from Zymomonas mobilis are the optimum candidates for 3-methyl-1-butanol production in Corynebacterium glutamicum. The recombinant strain produces 0.182 g/l of 3-methyl-1-butanol and 0.144 g/l of isobutanol after 12 h of incubation. Further inactivation of the E1 subunit of pyruvate dehydrogenase complex gene (aceE) and lactic dehydrogenase gene (ldh) improves the 3-methyl-1-butanol titer to 0.497 g/l after 12 h of incubation 1.1.1.1 alcohol dehydrogenase synthesis construction of a synthetic pathway for bioalcohol production at 70°C by insertion of the gene for alcohol dehydrogenase AdhA into the archaeon Pyrococcus furiosus. The engineered strain converts glucose to ethanol via acetate and acetaldehyde, catalyzed by the host-encoded aldehyde ferredoxin oxidoreductase AOR and heterologously expressed AdhA, in an energy-conserving, redox-balanced pathway. The AOR/AdhA pathway also converts exogenously added aliphatic and aromatic carboxylic acids to the corresponding alcohol using glucose, pyruvate, and/or hydrogen as the source of reductant. By heterologous coexpression of a membrane-bound carbon monoxide dehydrogenase, CO is used as a reductant for converting carboxylic acids to alcohols 1.1.1.1 alcohol dehydrogenase synthesis construction of an enzyme-immobilized bioanode that can operate at high temperatures. The catalytic current for ethanol oxidation at Ru complex-modified electrodes increases at 80°C up to 12fold compared with room temperature 1.1.1.1 alcohol dehydrogenase synthesis deletion of the hypoxanthine phosphoribosyltransferase gene in ethanol tolerant strain adhE*(EA), carrrying mutation P704L/H734R in the alcohol dehydrogenase gene, and deletion of lactate dehydrogenase (ldh) to redirect carbon flux towards ethanol reults in a strain producing 30% more ethanol than wild type on minimal medium. The engineered strain retains tolerance to 5% v/v ethanol, resulting in an ethanol tolerant platform strain 1.1.1.1 alcohol dehydrogenase synthesis engineering of a strain of Corynebacterium glutamicum, based on inactivation of the pyruvate dehydrogenase complex, pyruvate:quinone oxidoreductase, transaminase B, and additional overexpression of the IlvBNCD genes, encoding acetohydroxyacid synthase, acetohydroxyacid isomeroreductase, and dihydroxyacid dehydratase, for the production of isobutanol from glucose under oxygen deprivation conditions by inactivation of L-lactate and malate dehydrogenases, implementation of ketoacid decarboxylase from Lactococcus lactis, alcohol dehydrogenase 2 (ADH2) from Saccharomyces cerevisiae, and expression of the pntAB transhydrogenase genes from Escherichia coli. The resulting strain produces isobutanol with a substrate-specific yield (YP/S) of 0.60 mol per mol of glucose. Chromosomally encoded alcohol dehydrogenase AdhA rather than the plasmid-encoded ADH2 from Saccharomyces cerevisiae is involved in isobutanol formation, and overexpression of the corresponding AdhA gene increases the YP/S to 0.77 mol of isobutanol per mol of glucose. Inactivation of the malic enzyme significantly reduces the YP/S, indicating that the metabolic cycle consisting of pyruvate and/or phosphoenolpyruvate carboxylase, malate dehydrogenase, and malic enzyme is responsible for the conversion of NADH + H+ to NADPH + H+. In fed-batch fermentations with an aerobic growth phase and an oxygen-depleted production phase, the most promising strain produces about 175 mM isobutanol, with a volumetric productivity of 4.4 mM per h, and shows an overall YP/S of about 0.48 mol per mol of glucose in the production phase 1.1.1.1 alcohol dehydrogenase synthesis engineering of Klebsiella pneumoniae to produce 2-butanol from crude glycerol as a sole carbon source by expressing acetolactate synthase (IlvH), keto-acid reducto-isomerase (IlvC) and dihydroxyacid dehydratase (IlvD) from Klebsiella pneumoniae, and alpha-oxoisovalerate decarboxylase (Kivd) and alcohol dehydrogenase (AdhA) from Lactococcus lactis. The engineered strain produce 2-butanol (160 mg/l) from crude glycerol. Elimination of the 2,3-butanediol pathway by inactivating alpha-acetolactate decarboxylase (Adc) further improves the yield of 2-butanol from 160 to 320 mg/l 1.1.1.1 alcohol dehydrogenase synthesis enhancement of ethanol production capacity of Clostridium thermocellum by transferring pyruvate decarboxylase and alcohol dehydrogenase genes of the homoethanol pathway from Zymomonas mobilis. Both transferring pyruvate decarboxylase and alcohol dehydrogenase are functional in Clostridium thermocellum, but the presence of and alcohol dehydrogenase severely limits the growth of the recombinant strains, irrespective of the presence or absence of the pyruvate decarboxylase gene. The recombinant strain shows two-fold increase in pyruvate carboxylase activity and ethanol production when compared with the wild type strain 1.1.1.1 alcohol dehydrogenase synthesis enzyme catalyses the reduction of alpha-methyl and alpha-ethyl benzoylformate, and methyl o-chlorobenzoylformate with 100% conversion to methyl (S)-mandelate [17% enantiomeric excess (ee)], ethyl (R)-mandelate (50% ee), and methyl (R)-o-chloromandelate (72% ee), respectively, with an efficient in situ NADH-recycling system which involves glucose and a thermophilic glucose dehydrogenase 1.1.1.1 alcohol dehydrogenase synthesis enzyme catalyzes the following reactions with Prelog specificity: the reduction of acetophenone, 2,2,2-trifluoroacetophenone, alpha-tetralone, and alpha-methyl and alpha-ethyl benzoylformates to (S)-1-phenylethanol (>99% enantiomeric excess), (R)-alpha-(trifluoromethyl)benzyl alcohol (93% enantiomeric excess), (S)-alpha-tetralol (>99% enantiomeric excess), methyl (R)-mandelate (92% enantiomeric excess), and ethyl (R)-mandelate (95% enantiomeric excess), respectively, by way of an efficient in situ NADH-recycling system involving 2-propanol and a second thermophilic ADH 1.1.1.1 alcohol dehydrogenase synthesis expression of enzyme in auxotrophic Arxula adeninivorans, Hansenula polymorpha, and Saccharomyces cerevisiae strains using yeast ribosomal DNA integrative expression cassettes. Recombinant ADH accumulates intracellularly in all strains tested. The best yields of active enzyme are obtained from A. adeninivorans, with Saccharomyces cerevisiae producing intermediate amounts. Although Hansenula polymorpha is the least efficient producer overall, the product obtained is most similar to the enzyme synthesized by Rhodococcus ruber 219 with respect to its thermostability 1.1.1.1 alcohol dehydrogenase synthesis expression of pyruvate decarboxylase and alcohol dehydrogenase in Clostridium thermocellum DSM 1313. Though both enzymes are functional in Clostridium thermocellum, the presence of alcohol dehydrogenase severely limits the growth of the recombinant strains, irrespective of the presence or absence of the pyruvate decarboxylase gene 1.1.1.1 alcohol dehydrogenase synthesis in order to increase production of isobutanol, 2-oxoacid decarboxylase (KDC) and alcohol dehydrogenase (ADH) are expressed in Saccharomyces cerevisiae to enhance the endogenous activity of the Ehrlich pathway. Overexpression Ilv2, which catalyzes the first step in the valine synthetic pathway, and deletion of the PDC1 gene encoding a major pyruvate decarboxylase alters the abundant ethanol flux via pyruvate. Along with modification of culture conditions, the isobutanol titer is elevated 13fold, from 11 mg/l to 143 mg/l, and the yield is 6.6 mg/g glucose 1.1.1.1 alcohol dehydrogenase synthesis overexpression of the adhB gene results in a significant increase in the ethanol level 1.1.1.1 alcohol dehydrogenase synthesis protocol for the synthesis of [4R-(2)H]NADH with high yield by enzymatic oxidation of 2-propanol-d(8) 1.1.1.1 alcohol dehydrogenase synthesis recombinant enzyme activity can be improved by coexpression of archaeal chaperones (i.e., gamma-prefoldin and thermosome). Ricinoleic acid biotransformation activity of recombinant Escherichia coli expressing Micrococcus luteus alcohol dehydrogenase and the Pseudomonas putida KT2440 Baeyer-Villiger monooxygenase improves significantly with coexpression of gamma-prefoldin or recombinant themosome originating from the deep-sea hyperthermophile archaea Methanocaldococcus jannaschii. The degree of enhanced activity is dependent on the expression levels of the chaperones 1.1.1.1 alcohol dehydrogenase synthesis semi-preparative biocatalysis at 60°C using the stabilized mutant C257L, employing butyraldehyde for in situ cofactor regeneration with only catalytic amounts of NAD+, yields up to 23% conversion of omega-hydroxy lauric acid methyl ester to omega-oxo lauric acid methyl ester after 30 min 1.1.1.1 alcohol dehydrogenase synthesis simplified production scheme for isobutanol based on a cell-free immobilized enzyme system. Immobilized enzymes keto-acid decarboxylase (KdcA) and alcohol dehydrogenase (ADH) plus formate dehydrogenase (FDH) for NADH recycle in solution produce isobutanol titers 8 to 20 times higher than the highest reported titers with Saccharomyces cerevisiae on a mol/mol basis. Conversion rates and low protein leaching are achieved by covalent immobilization on methacrylate resin. The enzyme system without in situ removal of isobutanol achieves a 55% conversion of ketoisovaleric acid to isobutanol at a concentration of 0.135 mol isobutanol produced for each mol ketoisovaleric acid consumed 1.1.1.1 alcohol dehydrogenase synthesis synthesis of the cinnamyl alcohol by means of enzymatic reduction of cinnamaldehyde using alcohol both as an isolated enzyme, and in recombinant Escherichia coli whole cells in an efficient and sustainable one-phase system. The reduction of cinnamaldehyde (0.5 g/l, 3.8 mmol/l) by the isolated enzyme occurrs in 3 h at 50°C with 97% conversion, and yields high purity cinnamyl alcohol (98%) with a yield of 88% and a productivity of 50 g/g enzyme. The reduction of 12.5 g/l (94 mmol/l) cinnamaldehyde by whole cells in 6 h, at 37°C and no requirement of external cofactor occurrs with 97% conversion, 82% yield of 98% pure alcohol and a productivity of 34 mg/g wet cell weight 1.1.1.1 alcohol dehydrogenase synthesis synthetic pathway for n-butanol production from acetyl coenzyme at 70°C, using beta-ketothiolase Thl, 3-hydroxybutyryl-CoA dehydrogenase Hbd, and 3-hydroxybutyryl-CoA dehydratase Crt from Caldanaerobacter subterraneus subsp. tengcongensis, trans-2-enoyl-CoA reductase Ter from Spirochaeta thermophila and bifunctional aldehyde dehydrogenase AdhE and and butanol dehydrogenase in vitro. n-Butanol is produced at 70°C, but with different amounts of ethanol as a coproduct, because of the broad substrate specificities of AdhE, Bad, and Bdh. A reaction kinetics model, validated via comparison to in vitro experiments, is used to determine relative enzyme ratios needed to maximize n-butanol production. By using large relative amounts of Thl and Hbd and small amounts of Bad and Bdh, >70% conversion to n-butanol is observed in vitro, but with a 60% decrease in the predicted pathway flux 1.1.1.1 alcohol dehydrogenase synthesis yeast alcohol dehydrogenase with its cofactor NAD+ can be stably encapsulated in liposomes composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine. The liposomes are 100 nm in mean diameter, the liposomal ADH and NAD+ concentrations are 2.3 mg/ml and 3.9 mM, respectively. Free ADH is increasingly deactivated during its incubation at 45°C for 2 h with decrease of the enzyme concentration from 3.3 to 0.01 mg/ml because of the dissociation of tetrameric ADH into its subunits. Both liposomal enzyme systems, in presence and absence of NAD+, show stabilities at both 45 and 50°C much higher than those of the free enzyme systems, implying that the liposome membranes stabilize the enzyme tertiary and quaternary structures. The enzyme activity of the liposomes in presence of NAD+ show a stability higher than that in absence of NAD+ with a more remarkable effect of NAD+ at 50°C than at 45°C 1.1.1.1 alcohol dehydrogenase synthesis immobilization of enzyme on metal-derivatized epoxy Sepabeads. The highest immobilization efficiency (100%) and retention activity (60%) are achieved after 48 h of incubation of the enzyme with Niepoxy Sepabeads support in 100 mM Tris-HCl buffer, pH 8, containing 3 M KCl at 5°C. A significant increase in the stability of the immobilized enzyme is achieved by blocking the unreacted epoxy groups with ethylamine. The immobilization process increases the enzyme stability, thermal activity, and organic solvents. One step purification-immobilization can be carried out on metal chelate-epoxy Sepabeads 1.1.1.1 alcohol dehydrogenase synthesis under optimized conditions, the enzyme produces 600 mg all-trans-retinol per l after 3 h, with a conversion yield of 27.3% (w/w) and a productivity of 200 mg per l and h 1.1.1.1 alcohol dehydrogenase synthesis the alcohol dehydrogenase from Pyrococcus furiosus is a very robust enzyme in some organic solvents. From a synthetic point of view, this property is particularly important and useful for the reduction of ketones with a low solubility in aqueous buffers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenase. The enzyme from Acetobacter aceti catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenase. The enzyme from Aminobacter aminovorans slightly catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenase. The enzyme from Gluconobacter diazotrophicus slightly catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenase. The enzyme from Komagataeibacter medellinensis catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenase. The enzyme from Komagataeibacter xylinus catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis development of biotransformation process for asymmetric reduction with anti-Prelog NADH-dependent alcohol dehydrogenases. The enzyme from Acetobacter senegalensis catalyzes the formation of (S)-ethyl-4-chloro-3-hydroxybutanoate ((S)-CHBE), a key chiral intermediate in the synthesis of HMG-CoA reductase inhibitors (cholesterol lowering drugs like lipitor), slagenins B, slagenins C, and 1,4-dihydropyridine type beta-blockers 1.1.1.1 alcohol dehydrogenase synthesis horse liver alcohol dehydrogenase (HLADH) together with the NADH oxidase from Streptococcus mutans (SmNOX) are applied for the oxidative lactamization of various amino alcohols, direct synthesis of lactams (5-, 6-, and 7-membered) starting from amino-alcohols in a bienzymatic cascade. A direct approach for biocatalytic oxidative lactamization reaction. In situ regeneration of NAD+ with SmNOX in the HLADH-catalyzed oxidative lactamization of 4-amino-1-butanol to gamma-butyrolactam. The bienzymatic reaction cascade exhibits an optimum between pH 8 and pH 10, which can be attributed to the rather narrow pH range of SmNOX compared to that of HLADH. The fast reoxidation of NADH eliminated inhibitory effects of NADH on the HLADH-catalyzed oxidation 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis - 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis develpoment of conversion processes for petrochemicals and oil-contaminated environments, cinnamyl aldehyde and cinnamyl alcohol used in flavor and perfume industry, anisaldehyde is used for perfume and toilet soaps, decylalcohol is used in the manufacture of plasticizers, a production system for this enzyme may be useful for industrial application as a biocatalyst in the future 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis potential use for industrial production of ethanol by fermentation, thermophilic fermentations offer the potential to separate ethanol from continous cultures at process temperature and reduced pressure during growth 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis reduction of industrially important compounds cinnamyl aldehyde and anisaldehyde, industrial bioconversion of useful alcohols and aldehydes 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis useful for asymmetric production of L-carnitine 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis industrial ethanol production 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis conversion of prochiral ketones to chiral alcohols by Escherichia coli coexpressing enzyme with NAD+-dependent formate dehydrogenase and pyridine nucleotide transhydrogenase genes pnta and pntb, conversion of 66% acetophenone to (R)-phenylethanol over 12 h 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis the NADP(H)-dependent enzyme is useful in the selective chemoenzymatic synthesis of the tert-butyl (S)-6-chloro-5-hydroxy-3-ketohexanoate, a highly regio- and enantioselective reduction of a beta,delta-diketohexanoate ester, scale up of the continous fed-batch method, overview 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis 50 microg of alcohol dehydrogenase AdhA, EC 1.1.1.2, and 50 microg actaldehyde dehydrogenase AldH, EC 1.2.1.10,in buffer solution (pH 8.0) containing NADPH, NADH and acetyl-CoA at 60°C, produce 1.6 mM ethanol from 3 mM acetyl-CoA after 90 min 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis expression of BdhA enzyme in Caldicellulosiruptor bescii confers increased resistance of the engineered strain to both furfural and 5-hydroxymethylfurfural. In presence of 15 mM of either furan aldehyde, the ability to eliminate furfural or 5-hydroxymethylfurfural from the culture medium is significantly improved in the engineered strain 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis synthetic pathway for bioalcohol production at 70°C by insertion of the gene for bacterial alcohol dehydrogenase AdhA into the archaeon Pyrococcus furiosus. The engineered strain converts glucose to ethanol via acetate and acetaldehyde, catalyzed by the host-encoded aldehyde ferredoxin oxidoreductase AOR and heterologously expressed AdhA, in an energy-conserving, redox-balanced pathway. The AOR/AdhA pathway also converts exogenously added aliphatic and aromatic carboxylic acids to the corresponding alcohol using glucose, pyruvate, and/or hydrogen as the source of reductant. By heterologous coexpression of a membrane-bound carbon monoxide dehydrogenase, CO is used as a reductant for converting carboxylic acids to alcoholsThe AOR/AdhA pathway is a potentially game-changing strategy for syngas fermentation, especially in combination with carbon chain elongation pathways 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis overexpression of the endogenous zwf gene, which encodes glucose-6-phosphate dehydrogenase of the pentose phosphate pathway, in Synechocystis sp. PCC 6803 results in increased NADPH production, and promoted biomass production. Ethanol production by alcohol dehydrogenase YqhD is increased in autotrophic conditions by zwf overexpression 1.1.1.2 alcohol dehydrogenase (NADP+) synthesis engineering ADHs for regenerating NADPH by oxidation of diols 1.1.1.3 homoserine dehydrogenase synthesis contrary to wild-type MGA3 cells that secrete 0.4 g/l L-lysine and 59 g/l L-glutamate under optimised fed batch methanol fermentation, the hom-1 mutant M168-20 secretes 11 g/l L-lysine and 69 g/l of L-glutamate. Overproduction of pyruvate carboxylase and its mutant enzyme P455S in M168-20 has no positive effect on the volumetric L-lysine yield and the L-lysine yield on methanol, and causes significantly reduced volumetric L-glutamate yield and L-glutamate yield on methanol 1.1.1.3 homoserine dehydrogenase synthesis enzyme HSD is utilized in the large scale production of L-lysine 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis production of ethyl (R)-4-chloro-3-hydroxybutanoate using whole recombinant cells of Escherichia coli and 2-propanol as an energy source to regenerate NADH. Yield reaches 36.6 g/l with purity of more than 99% enantiomeric excess and 95.2% conversion 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis synthesis of (R)-1,3-butanediol from its racemate by stereoselective oxidation of the (S)-isomer using (S)-specific secondary alcohol dehydrogenase in whole recombinant Escherichia coli cells. Yield of the (R)-product reaches 72.6 g/l, with a molar recovery yield of 48.4% and an optical purity of 95% enantiomeric excess 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis the enzyme is useful in production of chiral compounds for organic synthesis 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis the immobilized enzyme is utilized in the asymmetric reduction of acetophenone to produce (S)-1-phenylethanol, with an enantiomeric excess of more than 99% 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis synthesis of ethyl (S)-4-chloro-3-hydroxybutanoate in Escherichia coli. Coexpression of carbonyl reductase CRII and a glucose dehydrogenase gives an activity of 15 U/mg protein using ethyl 4-chloro-3-oxobutanoate as a substrate in a water/butyl acetate system. The transformants give a molar yield of 91%, and an optical purity of the (S)-isomer of more than 99% enantiomeric excess 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis the enzyme can be used for stereospecific interconversion of (R)-1-phenylethanol and (S)-1-phenylethanol via the oxoform together with the (R)-specific secondary alcohol dehydrogenase using whole cells as biocatalysts that include the required cofactor regenration system, method, overview. Optically pure secondary alcohols are widely used in pharmaceuticals, flavors, agricultural chemicals and specialty materials 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis the enzyme catalyzes the asymmetric reduction of ethyl 4-chloro-3-oxobutanoate, the activity is 6.2 U/mg. Using two coexisting recombinant Escherichia coli strains, in which a strain expressing glucose dehydrogenase is used as an NADPH regenerator. An optical purity of 99% (e.e.) and a maximum yield of 1240 mM (S)-4-chloro-3-hydroxybutanoate are obtained, and highest turnover number of 53900 can be achieved without adding extra NADP+/NADPH 1.1.1.B3 (S)-specific secondary alcohol dehydrogenase synthesis using recombinant Scr2 in an aqueous-organic solvent system with a substrate fed-batch strategy and a final substrate concentration of 1 M, a yield of 95.3% and e.e. of 99% is obtained after 6-h reaction 1.1.1.4 (R,R)-butanediol dehydrogenase synthesis increase in production of (R,R)-butanediol from xylose in batch and continuous cultures by increase of temperature from 30 to 39°C, analysis of byproducts 1.1.1.4 (R,R)-butanediol dehydrogenase synthesis the enzyme is useful in production of 2,3-butanediol, an important starting material for the manufacture of bulk chemicals such as methyl ethyl ketone and 1,3-butadiene 1.1.1.4 (R,R)-butanediol dehydrogenase synthesis Paenibacillus brasilensis produces 2,3-butanediol (2,3-BDO) and can be utilized for large scale production 1.1.1.4 (R,R)-butanediol dehydrogenase synthesis two coexpressed enantiocomplementary carbonyl reductases, BDHA (2, 3-butanediol dehydrogenase from Bacillus subtilis) and GoSCR (polyol dehydrogenase from Gluconobacter oxydans) are used for asymmetric reduction of 2-hydroxyacetophenone (2-HAP) to (R)-1-phenyl-1,2-ethanediol ((R)-PED) or (S)-1-phenyl-1,2-ethanediol ((S)-PED) with excellent stereochemical selectivity and coupled with cofactor regeneration by GDH. Enantiomerically pure (R)-1-phenyl-1,2-ethanediol ((R)-PED) can be used as a building block for the preparation of (R)-norfluoxetine, (R)-fluoxetine, and beta-lactam antibiotics 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis the enzym is useful in production of chiral compounds for organic synthesis 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis the enzyme can be used for stereospecific interconversion of (R)-1-phenylethanol and (S)-1-phenylethanol via the oxoform together with the (S)-specific secondary alcohol dehydrogenase using whole cells as biocatalysts that include the required cofactor regenration system, method, overview. Optically pure secondary alcohols are widely used in pharmaceuticals, flavors, agricultural chemicals and specialty materials 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis ethyl benzoylformate is asymmetrically reduced by the purified enzyme, using an additional coupled NADH regeneration system, with 95% conversion and in an enantiomeric excess of 99.9% 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis using recombinant Escherichia coli cells expressing Sdr, a yield of 82.5% for (R)-[3,5-bis(trifluoromethyl)phenyl]ethanol can be achieved within 12 h at a substrate concentration of up to 1000 M 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis the enzyme is coupled with formate dehydrogenase and co-immobilized on SiO2 particles, the system is used for continuous catalytic conversion of beta-hydroxyacetophenone to optically pure (r)-phenylethanediol with in situ NADH regeneration and recycling. Reusable system, method overview 1.1.1.B4 (R)-specific secondary alcohol dehydrogenase (NADH) synthesis the enzyme might be useful in application as a replacement of chemical synthesis of aromatic chiral beta-amino alcohols 1.1.1.6 glycerol dehydrogenase synthesis high specificity of enzyme for secondary alcohols in R-configuration, use of enzyme for production of chiral compounds 1.1.1.6 glycerol dehydrogenase synthesis biotransformation of glycerol to dihydroxyacetone by recombinant Gluconobacter oxydans DSM 2343. Overproduction of the glycerol dehydrogenase to improve production of dihydroxyacetone 1.1.1.6 glycerol dehydrogenase synthesis Thermoanaerobacter mathranii can produce ethanol from lignocellulosic biomass at high temperatures. Deletion of the Ldh gene coding for lactate dehydrogenase eliminates an NADH oxidation pathway. To further facilitate NADH regeneration used for ethanol formation, heterologous gene GldA is expressed leading to increased ethanol yield in the presence of glycerol using xylose as a substrate. The metabolism of the cells is shifted toward the production of ethanol over acetate, hence restoring the redox balance. The recombinant acquired the capability to utilize glycerol as an extra carbon source in the presence of xylose resulting in a higher ethanol yield 1.1.1.6 glycerol dehydrogenase synthesis efficiency of a cofactor regeneration enzyme co-expressed with a glycerol dehydrogenase for the production of 1,3-dihydroxyacetone. In vitro biotransformation of glycerol is achieved with the cell-free extracts containing recombinant glycerol dehydrogenase from Escherichia coli, lactate dehydrogenase form Bacillus subtilis, or NADH oxidase LpNox1 from Lactobacillus pentosus, giving1,3-dihydroxyacetone (DHA), no expensive consumption of NAD+ for the production of DHA, overview. DHA is a valuable chemical with a wide range of applications in the cosmetics 1.1.1.6 glycerol dehydrogenase synthesis GlyDH is active with immobilized N6-CM-NAD+, suggesting that N6-CM-NAD+ can be immobilized on an electrode to allow TmGlyDH activity in a system that reoxidizes the cofactor electrocatalytically, development of a bioelectrocatalytic reactor 1.1.1.6 glycerol dehydrogenase synthesis engineering and immobilizing of glycerol dehydrogenase to accept alkyl/aryl glyceryl monoethers and catalyze their enantioselective oxidation to yield the corresponding 3-alkoxy/aryloxy-1-hydroxyacetones. The enzyme is highly enantioselective towards S-isomers (ee > 99%). Application of mutant L252A in a one-pot chemoenzymatic process to convert glycidol and ethanol into 3-ethoxy-1-hydroxyacetone and (R)-3-ethoxypropan-1,2-diol, without affecting the oxidation activity 1.1.1.6 glycerol dehydrogenase synthesis engineering of a hypertransformable variant of Clostridium pasteurianum for bioconversion of glycerol into hydrogen via increasing product yield by overexpression of enzyme catalyzing H2 production, and increasing substrate uptake by overexpression of enzymes involved in glycerol utilization. Overexpression of the HydA gene encoding hydrogenase, and overexpression of DhaD1 and DhaK genes encoding glycerol dehydrogenase and dihydroxyacetone kinase result in two recombinant strains (HydA++/HhaD1K++) capable of producing 97% H2 (v/v), with yields of 1.1 mol H2/mol glycerol in HydA overexpressed strain, and 0.93 mol H2/mol glycerol in DhaD1K overexpressing strain 1.1.1.6 glycerol dehydrogenase synthesis immobilization of Escherichia coli cells harboring the recombinant glycerol dehydrogenase gene on mannose-functionalized magnetic nanoparticles for conversion of glycerol to 1,3-dihydroxyacetone. Immobilization uses specific binding between mannose on the nanoparticles and the FimH lectin on the Escherichia coli cell surface via hydrogen bonds and hydrophobic interactions. Compared with the free cells, the thermostability of the immobilized cells is improved 2.56fold at 37°C. More than 50% of the initial activity of the immobilized cells remains after 10 cycles 1.1.1.6 glycerol dehydrogenase synthesis regeneration of NAD+ in enzyme-catalyzed reactions using aggreagets of glycerol dehydrogenase and NADH oxidase. After optimization, the activities of combi-aggregates and separate aggregates mixtures are 950 and 580 U/g, respectively. After ten cycles of reuse, the catalytic efficiency may still retain 63.3% of its initial activity. The conversion of glycerol to 1,3-dihydroxyacetone is 4.6%, which is 2.5 times of the free enzyme system 1.1.1.8 glycerol-3-phosphate dehydrogenase (NAD+) synthesis fermentative production of L-glycerol 3-phosphate utilizing a Saccharomyces cerevisiae strain with an engineered glycerol biosynthetic pathway (strain with deletions in both genes encoding specific L-G3Pases (GPP1 and GPP2) and multicopy overexpression of L-glycerol 3-phosphate dehydrogenase). Up-scaling the process employs fed-batch fermentation with repeated glucose feeding, plus an aerobic growth phase followed by an anaerobic product accumulation phase. This produces a final product titer of about 325 mg total L-glycerol 3-phosphate per liter of fermentation broth 1.1.1.8 glycerol-3-phosphate dehydrogenase (NAD+) synthesis successful introduction of a glycerol production pathway into Klebseiella pneumoniae by coexpression of genes encoding glycerol-3-phosphate dehydrogenase and glycerol-3-phosphatase (EC 3.1.3.21) organized into the plasmid pUC18K under control of the respective lac promoter. An engineered Klebsiella pneumoniae that can produce glycerol from glucose is achieved. It is still difficult to efficiently produce 1,3-propanediol from glucose. Only 0.58 g/l 1,3-propanediol is produced 1.1.1.8 glycerol-3-phosphate dehydrogenase (NAD+) synthesis deletion of the NAD+-dependent glycerol-3-phosphate dehydrogenase gene in an industrial ethanol-producing strain and expression of either the non-phosphorylating NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase from Bacillus cereus, strain AG2A, or the NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase GAPDH from Kluyveromyces lactis, strain AG2B, in the deletion strain. Recombinant strain AG2A exhibits a 48.70% decrease in glycerol production and a 7.60% increase in ethanol yield relative to the amount of substrate consumed, while recombinant strain AG2B exhibits a 52.90% decrease in glycerol production and a 7.34% increase in ethanol yield relative to the amount of substrate consumed, compared with the wild-type strain. The maximum specific growth rates of the recombinant AG2A and AG2B are higher than that of the gpd2 deletion strain and are indistinguishable compared with the wild-type strain in anaerobic batch fermentations 1.1.1.8 glycerol-3-phosphate dehydrogenase (NAD+) synthesis Camelina sativa coexpressing Arabidopsis thaliana diacylglycerol acyltransferase1 (DGAT1) and yeast cytosolic glycerol-3-phosphate dehydrogenase (GPD1) genes exhibit up to 13% higher seed oil content and up to 52% increase in seed mass compared to wild-type plants. DGAT1- and GDP1-coexpressing lines show significantly higher seed and oil yields on a dry weight basis than the wild-type controls or plants expressing DGAT1 and GPD1 alone. The oil harvest index (g oil per g total dry matter) for DGTA1- and GPD1-coexpressing lines is almost twofold higher as compared to wild type and the lines expressing DGAT1 and GPD1 alone 1.1.1.8 glycerol-3-phosphate dehydrogenase (NAD+) synthesis upon heterologous expression of diacylglycerol acyltransferase DGAT1, glycerol-3-phosphate dehydrogenase GPD1 and DGAT1 + GPD1 in Camelina sativa, increase in triacylglycerol production is limited by utilization of fixed carbon from the source tissues supported by the increase in glycolysis pathway metabolites and decreased transcripts levels of transcription factors controlling fatty acids synthesis, and triacylglycerol accumulation is limited by the activity of lipases/hydrolases that hydrolyze triacylglycerol pool supported by the increase in free fatty acids and monoacylglycerols 1.1.1.9 D-xylulose reductase synthesis optimization of xylitol production, using fed-batch process and controlled pH 6.0 gives maximum enzyme activity 1.1.1.9 D-xylulose reductase synthesis use of enzyme in production of xylitol from bagasse hydrolysate, enzyme activity is higher in medium containing acetic acid than in control medium 1.1.1.9 D-xylulose reductase synthesis use of enzyme in xylose fermentation, metabolic flux partitioning from xylitol to xylulose depends on aeration and enzyme activity, increased aeration results in less xylitol accumulation and more xylulose accumulation, increase in enzyme activity can reduce xylitol formation 1.1.1.9 D-xylulose reductase synthesis the enzyme is useful for xylitol bioproduction, profiles, overview 1.1.1.9 D-xylulose reductase synthesis Gluconobacter oxydans strain NH-10 is useful for production of xylitol from D-arabitol via D-xylulose 1.1.1.9 D-xylulose reductase synthesis enzyme IoXyl2p from Issatchenkia orientalis is considered to be an attractive candidate for the construction of genetically engineered Saccharomyces cerevisiae for efficient fermentation of carbohydrate in lignocellulosic hydrolysate 1.1.1.9 D-xylulose reductase synthesis enzyme TdXyl2p from Torulaspora delbrueckii is considered to be an attractive candidate for the construction of genetically engineered Saccharomyces cerevisiae for efficient fermentation of carbohydrate in lignocellulosic hydrolysate 1.1.1.10 L-xylulose reductase synthesis the microalga Chlorella sorokiniana and provide a target for genetic engineering to improve D-xylose utilization for microalgal lipid production 1.1.1.10 L-xylulose reductase synthesis potential approach for industrial-scale production of xylitol from hemicellulosic hydrolysate involving the enzyme 1.1.1.12 L-arabinitol 4-dehydrogenase synthesis immobilization of HjLAD onto silicon oxide nanoparticles has the potential for use in the industrial production of rare sugars, e.g. L-xylulose, due to the thermostability and reusability of the immobilized enzyme 1.1.1.12 L-arabinitol 4-dehydrogenase synthesis rare L-sugar L-xylulose is produced by the enzymatic oxidation of arabinitol to give a yield of approximately 86% 1.1.1.14 L-iditol 2-dehydrogenase synthesis L-sorbose is an important intermediate in the industrial vitamin C production process 1.1.1.14 L-iditol 2-dehydrogenase synthesis two co-expressed enantiocomplementary carbonyl reductases, BDHA (2,3-butanediol dehydrogenase from Bacillus subtilis) and GoSCR (polyol dehydrogenase from Gluconobacter oxydans) are used for asymmetric reduction of 2-hydroxyacetophenone (2-HAP) to (R)-1-phenyl-1,2-ethanediol ((R)-PED) or (S)-1-phenyl-1,2-ethanediol ((S)-PED) with excellent stereochemical selectivity and coupled with cofactor regeneration by GDH. Enantiomerically pure (R)-1-phenyl-1,2-ethanediol ((R)-PED) can be used as a building block for the preparation of (R)-norfluoxetine, (R)-fluoxetine, and beta-lactam antibiotics 1.1.1.16 galactitol 2-dehydrogenase synthesis the enzyme can be used to produce optically pure building blocks and for the bioconversion of bioactive compounds 1.1.1.16 galactitol 2-dehydrogenase synthesis a yeast strain capable of consuming lactose intracellularly is engineered to produce tagatose from lactose. GAL1 coding for galactose kinase is deleted to eliminate galactose utilization. Heterologous xylose reductase (XR) and galactitol dehydrogenase (GDH) are introduced into the Gal1 deletion strain. The expression levels of XR and GDH are adjusted to maximize tagatose production. The resulting engineered yeast produces 37.69 g/l of tagatose from lactose with a tagatose and galactose ratio of 9:1 in the reaction broth 1.1.1.17 mannitol-1-phosphate 5-dehydrogenase synthesis strategy for mannitol production in Lactococcus, most promising is overexpression of enzyme in a lactate-dehydrogenase deficient strain 1.1.1.17 mannitol-1-phosphate 5-dehydrogenase synthesis hydrogen transfer from formate to D-fructose 6-phosphate, mediated by NAD(H) and catalyzed by a coupled enzyme system of purified Candida boidinii formate dehydrogenase and AfM1PDH, is used for the preparative synthesis of D-mannitol 1-phosphate or, by applying an analogous procedure using deuterio formate, the 5-[2H] derivative thereof, overview 1.1.1.B18 L-1-amino-2-propanol dehydrogenase synthesis coversion of 1-(3-hydroxyphenyl)-2-(methylamino) ethanone to (S)-phenylephrine with with more than 99% enantiomeric excess, 78% yield and a productivity of 3.9 mmol(S)-phenylepinephrine/l h in 12 h at 30°C and pH 7. The (S)-phenylepinephrine, recovered from reaction mixture by precipitation at pH 11.3, can be converted to (R)-phenylepinephrine by Walden inversion reaction 1.1.1.B19 xylitol dehydrogenase (NAD+) synthesis production of L-xylulose from xylitol using a resting cell reaction leads to 35% L-xylulose within 24 h, starting from 5% xylitol as initial concentration 1.1.1.B19 xylitol dehydrogenase (NAD+) synthesis enzyme XDH from Erwinia aphidicola can be useful for production of L-xylulose, a rare ketose, for apllication in pharmaceutical and food industries 1.1.1.B20 meso-2,3-butandiol dehydrogenase synthesis discovery of the (S)-selective alcohol dehydrogenase enables a novel production process of (R)-acetoin from meso-2,3-butanediol 1.1.1.B20 meso-2,3-butandiol dehydrogenase synthesis Bacillus licheniformis strain MW3 (DELTAbudCDELTAgdh) can be useful for the production of acetoin on a commercial scale 1.1.1.B20 meso-2,3-butandiol dehydrogenase synthesis Bacillus subtilis enzyme BS-BDH is a potential candidate for L-(+)-acetoin production 1.1.1.B20 meso-2,3-butandiol dehydrogenase synthesis Paenibacillus brasilensis produces 2,3-butanediol (2,3-BDO) and can be utilized for large scale production 1.1.1.21 aldose reductase synthesis homochiral 3-hydroxy-4-substituted beta-lactams serve as precursors to the corresponding alpah-hydroxy-beta-amino acids, the enzyme might be useful insynthesis of these key components of many biologically and therapeutically important compounds 1.1.1.22 UDP-glucose 6-dehydrogenase synthesis the high activity combined with the simple purification procedure used make GbUGD a valuable alternative biocatalyst for the synthesis of UDP-glucuronic acid or the development of NAD+ regeneration systems 1.1.1.22 UDP-glucose 6-dehydrogenase synthesis use of recombinant Triton-permeabilized cells of Schizosaccharomyces pombe to synthesize UDP-glucuronic acid with 100 % yield and selectivity. 5 mM UDP-glucose are converted into 5 mM UDP-glucuronic acid within 3 h 1.1.1.27 L-lactate dehydrogenase synthesis development of yeast-based bioprocesses to produce lactate from lignocellulosic raw material 1.1.1.27 L-lactate dehydrogenase synthesis the enzyme has a commercial significance, as it can be used to produce chiral building blocks for the synthesis of key pharmaceuticals and agrochemicals, optimization of enzyme reaction by engineering to eliminate the substrate inhibition 1.1.1.27 L-lactate dehydrogenase synthesis the enzyme might be useful in the production of phenyllactate 1.1.1.27 L-lactate dehydrogenase synthesis a lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta) deletion strain is evolved for 2,000 h, resulting in a stable strain with 40:1 ethanol selectivity and a 4.2-fold increase in ethanol yield over the wild-type strain. In a coculture of organic acid-deficient engineered strains of both Clostridium thermocellum and Thermoanaerobacterium saccharolyticum, fermentation of 92 g/liter Avicel results in 38 g/liter ethanol, with acetic and lactic acids below detection limits, in 146 h. engineering is based on a phosphoribosyl transferase (Hpt) deletion strain, which produces acetate, lactate, and ethanol in a ratio of 1.7:1.5:1.0, similar to the 2.1:1.9:1.0 ratio produced by the wild type. The Hpt/Ldh double mutant strain does not produce significant levels of lactate and has a 1.4:1.0 ratio of acetate to ethanol. Similarly, the Hpt/Pta double mutant strain does not produce acetate and has a 1.9:1.0 ratio of lactate to ethanol. The Hpt/Ldh/Pta triple mutant strain achieves ethanol selectivity of 40:1 relative to organic acids 1.1.1.27 L-lactate dehydrogenase synthesis construction of a markerless strain lacking phosphotransacetylase Pta, acetate kinase Ack and lactate dehydrogenase Ldh genes. The gene deletion strain ferments 50 g/liter of cellobiose, with a yield of 0.44 g ethanol per g glucose equivalent substrate and a maximum volumetric productivity of 1.13 g ethanol per liter and h. A system for genetic marker removal allows for enactment of further modifications and creation of strains for industrial applications 1.1.1.27 L-lactate dehydrogenase synthesis metabolic engineering of Geobacillus thermoglucosidasius to divert the fermentative carbon flux from a mixed acid pathway, to one in which ethanol becomes the major product, involving elimination of the lactate dehydrogenase and pyruvate formate lyase pathways by disruption of the ldh and pflB genes, respectively, and upregulation of expression of pyruvate dehydrogenase. Strains with all three modifications form ethanol efficiently and rapidly at temperatures in excess of 60°C in yields in excess of 90% of theoretical. The strains also efficiently ferment cellobiose and a mixed hexose and pentose feed 1.1.1.27 L-lactate dehydrogenase synthesis Thermoanaerobacter mathranii can produce ethanol from lignocellulosic biomass at high temperatures. Deletion of the Ldh gene coding for lactate dehydrogenase eliminates an NADH oxidation pathway. To further facilitate NADH regeneration used for ethanol formation, a heterologous gene GldA encoding an NAD+-dependent glycerol dehydrogenase is expressed leading to increased ethanol yield in the presence of glycerol using xylose as a substrate. The metabolism of the cells is shifted toward the production of ethanol over acetate, hence restoring the redox balance. The recombinant enzyme acquired the capability to utilize glycerol as an extra carbon source in the presence of xylose resulting in a higher ethanol yield 1.1.1.27 L-lactate dehydrogenase synthesis L-nLDH is an efficient catalyst that can be used in the enantioselective reduction of alpha-keto acids to alpha-hydroxy acids 1.1.1.27 L-lactate dehydrogenase synthesis coexpression of enzyme and glucose dehydrogenase gene in Escherichia coli efficiently reduces 3,4-dihydroxyphenylpyruvate to L-3,4-dihydroxyphenyllactate with 95.45% isolation yield 1.1.1.27 L-lactate dehydrogenase synthesis engineering of Kluyveromyces marxianus to express and coexpress various heterologous LDH enzymes for L-lactic acid production. LDH enzymes originating from Staphylococcus epidermidis (SeLDH, optimal at pH 5.6), Lactobacillus acidophilus (LaLDH, optimal at pH 5.3), and Bos taurus (BtLDH, optimal at pH 9.8) are functionally expressed individually and in combination. A strain co-expressing SeLDH and LaLDH produces 16.0 g/l L-lactic acid, whereas the strains expressing those enzymes individually produces only 8.4 and 6.8 g/l, respectively. This coexpressing strain produces 24.0 g/l L-lactic acid with a yield of 0.48 g/g glucose in the presence of CaCO3 1.1.1.27 L-lactate dehydrogenase synthesis engineering of Kluyveromyces marxianus to express and coexpress various heterologous LDH enzymes for L-lactic acid production. LDH enzymes originating from Staphylococcus epidermidis (SeLDH, optimal at pH 5.6), Lactobacillus acidophilus (LaLDH, optimal at pH 5.3), and Bos taurus (BtLDH, optimal at pH 9.8) are functionally expressed individually and in combination. A strain coexpressing SeLDH and LaLDH produces 16.0 g/l L-lactic acid, whereas the strains expressing those enzymes individually produces only 8.4 and 6.8 g/l, respectively. This coexpressing strain produces 24.0 g/l L-lactic acid with a yield of 0.48 g/g glucose in the presence of CaCO3 1.1.1.27 L-lactate dehydrogenase synthesis production of phenyllactic acid from L-Phe by recombinant Escherichia coli coexpressing L-phenylalanine oxidase and L-lactate dehydrogenase. At optimal conditions (L-Phe 6 g/l, pH 7.5, 35°C, CDW 24.5 g/l and 200 rpm), the recombinant strain produces 1.62 g L-phenylalanine/l with a conversion of 28% from L-Phe 1.1.1.28 D-lactate dehydrogenase synthesis production of (R)-2-hydroxy-4-phenyl-butyric acid, which is a precursor for different ACE-inhibitors 1.1.1.28 D-lactate dehydrogenase synthesis enzymatic synthesis of (R)-3,4-dihydrixyphenyllactic acid, a pharmacological compound that is used for the treatment of menstrual disorders, menostasis, menorrhalgia, insomnia, blood circulation diseases and Angina pectoris. Regeneration of NADH by formate dehydrogenase system. Use of genetic algorithm as a stochastic optimization method seems to be the best choice for the optimization 1.1.1.28 D-lactate dehydrogenase synthesis D-LDH is a candidate for thermophilic D-lactic acid production 1.1.1.28 D-lactate dehydrogenase synthesis the polymers of lactic acid are used as biodegradable bioplastics. Polylactic acid, biodegradable polyester polymer of lactic acid, is mainly produced through a bacterial fermentation process 1.1.1.28 D-lactate dehydrogenase synthesis the recombinant enzyme from Pediococcus pentosaceus can be used for production of 3-phenyllactic acid (2-hydroxy-3-phenylpropanoic acid, PLA), an antimicrobial compound with broad spectrum activity against both bacteria and fungi, optimization by coexpression of Ogataea parapolymorpha formate dehydrogenase, EC 1.2.1.2, for NADH regeneration 1.1.1.28 D-lactate dehydrogenase synthesis asymmetric synthesis of (R)-2-hydroxy-4-phenylbutyric acid using recombinant Pichia pastoris expressing the Tyr52Leu variant of D-lactate dehydrogenase from Lactobacillus plantarum. The recombinant yeast cells show catalytic activity at a high concentration of 2-oxo-4-phenylbutyric acid (380 mM, 76 g/l). Under optimized reaction conditions (pH 7.5, 37°C, and 2% glucose), a full conversion with over 95% reaction yield and about 100% product enantiomeric excess is achieved 1.1.1.28 D-lactate dehydrogenase synthesis recombinant Escherichia coli expressing D-lactate dehydrogenase, without coexpression of a cofactor regeneration system, can produce 20.5 g/l D-phenyllactic acid with enantiomeric excess above 99% from phenylpyruvic acid in a fed-batch biotransformation process, with a productivity of 49.2 g/l per day 1.1.1.28 D-lactate dehydrogenase synthesis recombinant Escherichia coli expressing Ldb0101 achieves a D-lactate concentration of 949.6 mg/l under aerobic and 1.94 g/l under anaerobic conditions, respectively 1.1.1.28 D-lactate dehydrogenase synthesis recombinant Escherichia coli expressing Ldb1010 achieves a D-lactate concentration of 850 mg/l 1.1.1.28 D-lactate dehydrogenase synthesis synthesis of D-phenyllactic acid by Escherichia coli expressing D-lactate dehdrogenase plus Exiguobacterium sibiricum glucose dehydrogenase. The total enzyme activity in the fermentation broth reaches 2359.0 U/l when induced by 10 g/l lactose at 28°C and 150 rpm for 14 h. Under the optimized biocatalysis conditions, 50 g/l sodium phenylpyruvate is completely converted to D-phenyllactic acid with a space-time yield and enantiomeric excess of 262.8 g/l day and over 99.5%, respectively 1.1.1.30 3-hydroxybutyrate dehydrogenase synthesis the engineered enzyme mutant H144L/W187F is used for production of 4-hydroxyvaleric acid, a monomer of bio-polyester and a precursor of bio-fuels, from levulinic acid 1.1.1.35 3-hydroxyacyl-CoA dehydrogenase synthesis the enzyme is used in the biosynthesis of n-butanol from acetyl-CoA by the reduction of acetoacetyl-CoA to (S)-3-hydroxybutyryl-CoA 1.1.1.35 3-hydroxyacyl-CoA dehydrogenase synthesis the highly efficient mutant enzyme K50A/K54A/L232Y can be useful for increasing the production rate of n-butanol 1.1.1.36 acetoacetyl-CoA reductase synthesis PHB-synthesis for thermoplastics 1.1.1.36 acetoacetyl-CoA reductase synthesis establishing of an enzyme-catalyzed synthesis system for production of poly(3-hydroxybutyrate) in vitro on the basis of the poly(3-hydroxybutyrate) biosynthesis pathway of Ralstonia eutropha, recycling CoA for synthesis of acetyl-CoA and deriving NADPH from regeneration by GDH, overview 1.1.1.36 acetoacetyl-CoA reductase synthesis the engineered enzyme mutant T173S can be used for increased production of poly(3-hydroxybutyrate) in a Corynebacterium glutamicum expression system 1.1.1.39 malate dehydrogenase (decarboxylating) synthesis the enzyme is useful for production of L-malic acid with NADH generation including the reverse reaction of malic enzyme and the activity of glucose-6-phosphate dehydrogenase, EC1.1.1.49, from Leuconostoc mesenteroides, overview 1.1.1.44 phosphogluconate dehydrogenase (NADP+-dependent, decarboxylating) synthesis thermostability may lead to some practical applications 1.1.1.44 phosphogluconate dehydrogenase (NADP+-dependent, decarboxylating) synthesis NADPH regeneration. When coupled with glucose-6-phosphate dehydrogenase the enzyme generates two moles of NADPH per mole of glucose-6-phosphate 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis enzyme can be used for gluconic acid production in low water systems 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis usage as NADP+ cofactor regenerator for enzymatic synthesis of chiral compounds such as ethyl-(S)-4-chloro-3-hydroxybutanoate and ethyl 4-chloro-3-oxobutanoate 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis production of recombinant glucose 1-dehydrogenase in Escherichia coli, optimization of culture and induction conditions. Glucose 1-dehydrogenase is used to regenerate NADPH in vivo and in vitro and coupled with a NADPH-dependent bioreduction for efficient synthesis of ethyl (R)-4-chloro-3-hydroxybutanoate from ethyl-4-chloro-3-oxobutanoate 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis (±)-ethyl mandelate are important intermediates in the synthesis of numerous pharmaceuticals. Efficient routes for the production of these derivatives are highly desirable. A co-immobilization strategy is developed to overcome the issue of NADPH demand in the short-chain dehydrogenase/reductase (SDR) catalytic process. The SDR from Thermus thermophilus HB8 and the NAD(P)-dependent glucose dehydrogenase (GDH) from Thermoplasma acidophilum DSM 1728 are co-immobilized on silica gel. This dual-system offers an efficient route for the biosynthesis of (+/-)-ethyl mandelate 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis co-immobilization of ketoreductase (KRED) and glucose dehydrogenase (GDH) on highly cross-linked agarose (sepharose) via affnity interaction between His-tagged enzymes (six histidine residues on the N-terminus of the protein) and agarose matrix charged with nickel (Ni2+ ions). Immobilized enzymes are applied in a set of biotransformation reactions in repeated batch flow-reactor mode. Immobilization reduces the requirement for cofactor (NADP+) and allows the use of higher substrate concentration in comparison with free enzymes 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis glucose dehydrogenase is a general tool for driving nicotinamide (NAD(P)H) regeneration in synthetic biochemistry. Coupled with a Candida glabrata carbonyl reductase, the mutant glucose dehydrogenase Q252L/E170K/S100P/K166R/V72I/K137R is successfully used for the asymmetric reduction of deactivating ethyl 2-oxo-4-phenylbutyrate with total turnover number of 1800 for the nicotinamide cofactor, thus making it attractive for commercial application 1.1.1.47 glucose 1-dehydrogenase [NAD(P)+] synthesis production of tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate, an important chiral intermediate for the synthesis of rosuvastatin, using carbonyl reductase coupled with glucose dehydrogenase. A recombinant Escherichia coli strain harboring carbonyl reductase R9M and glucose dehydrogenase is constructed with high carbonyl reduction activity and cofactor regeneration efficiency. The recombinant Escherichia coli cells are applied for the efficient production of tert-butyl (3R,5S)-6-chloro-3,5-dihydroxyhexanoate with a substrate conversion of 98.8%, a yield of 95.6% and an enantiomeric excess of more than 99.0% under 350 g/l of tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate after 12 h reaction. A substrate fed-batch strategy is further employed to increase the substrate concentration to 400 g/l resulting in an enhanced product yield to 98.5% after 12 h reaction in a 1 l bioreactor. Meanwhile, the space-time yield is 1182.3 g/l*day 1.1.1.49 glucose-6-phosphate dehydrogenase (NADP+) synthesis bacitracin, a kind of cyclic peptide antibiotic mainly produced by Bacillus, has wide ranges of applications. The bacitracin production is enhanced by by NADPH generation via overexpressing glucose-6-phosphate dehydrogenase Zwf in Bacillus licheniformis 1.1.1.50 3alpha-hydroxysteroid 3-dehydrogenase (Si-specific) synthesis the enzyme is useful foe androsterone production in a coupled system with formate dehydrogenase in enhancing Tris-HCl/co-solvent 1-butyl-3-methylimidazolium L-lactate, at pH 7.6 and 25°C, method optimization, overview 1.1.1.50 3alpha-hydroxysteroid 3-dehydrogenase (Si-specific) synthesis the enzyme is useful in reductive production of steroids. In a coupled-enzyme system comprising HSDH and formate dehydrogenase, a twofold increase in production rate of androsterone is obtained when utilizing 1-butyl-3-methylimidazolium L-lactate with NADH regeneration 1.1.1.51 3(or 17)beta-hydroxysteroid dehydrogenase synthesis engineering Mycobacterium smegmatis for testosterone production. Mycobacterium smegmatis is an excellent chassis to develop biotechnological processes for the biotransformation of sterols and their derivatives into valuable pharmaceutical compounds. Overexpression of the gene encoding microbial 17beta-hydroxysteroid: NADP 17-oxidoreductase, from the bacterium Comamonas testosteroni. The host strains are Mycobacterium smegmatis wild type and a genetic engineered androst-4-ene-3,17-dione producing mutant. The recombinant strains are able to produce testosterone from androst-4-ene-3,17-dione and/or from sterols with high yields 1.1.1.51 3(or 17)beta-hydroxysteroid dehydrogenase synthesis engineering Mycobacterium smegmatis for testosterone production. Mycobacterium smegmatis is an excellent chassis to develop biotechnological processes for the biotransformation of sterols and their derivatives into valuable pharmaceutical compounds. Overexpression of the gene encoding microbial 17beta-hydroxysteroid: NADP 17-oxidoreductase, from the fungus Cochliobolus lunatus. The host strains are Mycobacterium smegmatis wild type and a genetic engineered androst-4-ene-3,17-dione producing mutant. The recombinant strains are able to produce testosterone from androst-4-ene-3,17-dione and/or from sterols with high yields 1.1.1.B51 3-quinuclidinone reductase (NADPH) synthesis stereospecific production of (R)-3-quinuclidinol, an important chiral building block for the synthesis of various pharmaceuticals 1.1.1.B51 3-quinuclidinone reductase (NADPH) synthesis the enzyme can be used for synthesis of optically pure 3-quinuclidinol. Optically pure 3-quinuclidinol is an important intermediate for the synthesis of various anticholinergic drugs. (R)-3-Quinuclidinol is used to synthesize muscarinic M1 or M3 receptor antagonists such as talsaclidine, revatropate, and solifenacin 1.1.1.B52 3-quinuclidinone reductase (NADH) synthesis stereospecific production of (R)-3-quinuclidinol, an important chiral building block for the synthesis of various pharmaceuticals 1.1.1.B52 3-quinuclidinone reductase (NADH) synthesis stereospecific production of (R)-3-quinuclidinol, an important chiral building block for the synthesis of various pharmaceuticals, high yield of (R)-3-quinuclidinol up to 916 g/L * d using a bioreduction approach 1.1.1.B52 3-quinuclidinone reductase (NADH) synthesis stereospecific production of (R)-3-quinuclidinol, an important chiral building block for the synthesis of various pharmaceuticals. The 3-quinuclidinone reductase and Leifsonia sp. alcohol dehydrogenase genes are efficiently expressed in Escherichia coli cells. A number of constructed Echerichia coli biocatalysts (intact or immobilized) are applied to the resting cell reaction and optimized. Under the optimized conditions, (R)-(-)-3-quinuclidinolis synthesized from 3-quinuclidinone (15% w/v, 939 mM) giving a conversion yield of 100% for the immobilized enzyme. The optical purity of the (R)-(-)-3-quinuclidinol produced by the enzymatic reactions is above 99.9% 1.1.1.B52 3-quinuclidinone reductase (NADH) synthesis (R)-3-quinuclidinol is a valuable intermediate for pharmaceuticals. The enzyme can be used for the synthesis of the enantiopure compound 1.1.1.56 ribitol 2-dehydrogenase synthesis the recombinant Escherichia coli expressing D-psicose-3-epimerase (DPE), ribitol dehydrogenase (RDH) and formate dehydrogenase (FDH) is constructed and used together with immobilized GI for allitol bioproduction from D-glucose. The conditions of allitol biotransformation, the cell catalytic activity resistance, the cell cultivation medium, and fed-batch culture conditions are optimized 1.1.1.58 tagaturonate reductase synthesis expression of Lactococcus lactis uxaB and uxaC genes encoding D-tagaturonate reductase and D-galacturonate isomerase, in Saccharomyces cerevisiae to investigate in vivo activity of the first steps of the D-galacturonate pathway. Although D-tagaturonate reductase could, in principle, provide an alternative means for re-oxidizing cytosolic NADH, addition of D-galacturonate does not restore anaerobic growth, possibly due to absence of a functional D-altronate exporter in Saccharomyces cerevisiae 1.1.1.B60 D-sorbitol dehydrogenase (NADP+) synthesis synthesis of L-sorbose is largely used as a starting material for L-ascorbic acid biosynthesis. It can been also used to synthesize the potent glycosidase inhibitor 1-deoxygalactonojirim and rare sugars such as L-tagatose and L-iditol. Escherichia coli(gosldh-lrenox) producing both GoSLDH for D-sorbitol oxidation and LreNOX (NAD(P)H oxidase from Lactobacillus reuteri) for NADP+ regeneration is generated and used for L-sorbose production. L-Sorbose production by Escherichi coli(gosldh-lrenox) reaches 4.1 g/l after 40 min, which was 20.5fold higher than that of Escherichia coli(gosldh). This system reduces the NADPH inhibition effect in the GoSLDH reaction and enables high production of L-sorbose from D-sorbitol 1.1.1.B63 3-quinuclidinone reductase (NADP+) synthesis the enzyme can be used for synthesis of optically pure 3-quinuclidinol. Optically pure 3-quinuclidinol is an important intermediate for the synthesis of various anticholinergic drugs. (S)-3-Quinuclidinol is a very promising chiral building blocks for synthesis of serotonin receptor antagonist drugs and anticholinergic drugs 1.1.1.67 mannitol 2-dehydrogenase synthesis the recombinant enzyme expressed in Bacillus megaterium is useful in production of D-mannitol using a resting cell biotransformation approach 1.1.1.67 mannitol 2-dehydrogenase synthesis an effective strategy for producing high yields of mannitol is developed. The combined strategies of aeration induction and redox modulation significantly increases the glucose consumption rate, intracellular NADH level and the specific activity of mannitol dehydrogenase (MDH), resulting in an increase in mannitol production from 64.6 to 88.1 g/l with the yield increased from 0.69 to 0.94 g/g 1.1.1.67 mannitol 2-dehydrogenase synthesis mannitol is a natural hexitol with important applications in medicine and food industry. Development of a production method on an industrial scale, optimization and evaluation of production in a batch reactor (BR, or BRP operation) for a complex bi-enzymatic system with suspended enzymes and cofactor regeneration, method modeling, molecular calculations and simulations, detailed overview 1.1.1.67 mannitol 2-dehydrogenase synthesis the enzyme might be useful for enzymatic D-mannitol production in an industrial scale. The purified mannitol dehydrogenase have been reported to produce D-mannitol with no sorbitol formation at temperatures of 90-120°C. The pathway for D-mannitol production using MtDH isolated from Thermotoga maritima involves production from glucose via Thermotoga neapolitana xylose isomerase (gene xylA, UniProt ID P45687) followed by the conversion of the formed D-fructose using Thermotoga maritima MtDH of enzymatic to chemical synthesis process, overview 1.1.1.67 mannitol 2-dehydrogenase synthesis the enzyme might be useful for enzymatic D-mannitol production in an industrial scale. The purified mannitol dehydrogenase have been reported to produce D-mannitol with no sorbitol formation at temperatures of 90-120°C. The pathway for D-mannitol production using MtDH isolated from Thermotoga neapolitana is via D-fructose in a single step procedure. Comparison of enzymatic to chemical synthesis process, overview 1.1.1.69 gluconate 5-dehydrogenase synthesis strain overexpressing enzyme plus Escherichia coli transhydrogenase sthA, enhanced accumulation of 5-ketoD-gluconate, precursor of L-(+)-tartaric acid 1.1.1.76 (S,S)-butanediol dehydrogenase synthesis preparation of chiral acetoinic compounds, enzymic identification for chiral acetoinic compounds or as model enzyme for studying the interrelation between enzymic stereospecificity and structure 1.1.1.76 (S,S)-butanediol dehydrogenase synthesis the key enzymes in the microbial production of 2,3-butanediol 1.1.1.77 lactaldehyde reductase synthesis fermentation of L-rhamnose, L-fucose and D-fucose to a mixture of 1,2-propanediol, acetone, H2, CO2 and ethanol 1.1.1.81 hydroxypyruvate reductase synthesis potential application in the enzymatic synthesis of glyoxylate 1.1.1.87 homoisocitrate dehydrogenase synthesis novel glutarate biosynthetic pathway by incorporation of a +1 carbon chain extension pathway from 2-oxoglutarate in combination with 2-oxo acid decarboxylation pathway in Escherichia coli. Introduction of homocitrate synthase, homoaconitase and homoisocitrate dehydrogenase from Saccharomyces cerevisiae into Escherichia coli enables +1 carbon extension from 2-oxoglutarate to 2-oxoadipate, which is subsequently converted into glutarate by a promiscuous 2-oxo acid decarboxylase (KivD) and a succinate semialdehyde dehydrogenase (GabD). The recombinant Escherichia coli coexpressing all five genes produces 0.3 g/l glutarate from glucose. To further improve the titers, 2-oxoglutarate is rechanneled into carbon chain extension pathway via the clustered regularly interspersed palindromic repeats system mediated interference (CRISPRi) of essential genes sucA and sucB in tricarboxylic acid cycle. The final strain can produce 0.42 g/l glutarate, which is increased by 40% compared with the parental strain. Glutarate is one of the most potential building blocks for bioplastics 1.1.1.90 aryl-alcohol dehydrogenase synthesis biotechnological production of vanillin 1.1.1.94 glycerol-3-phosphate dehydrogenase [NAD(P)+] synthesis development of a whole-cell biocatalyst for NAD(P)H cofactor regeneration that employs permeabilized Escherichia coli cells in which the glpD and gldA genes are deleted and the gpsA gene is overexpressed. The biocatalyst involves an economical substrate, bifunctional regeneration of NAD(P)H, and simple reaction conditions as well as a stable environment for enzymes, and is applicable to a variety of oxidoreductase reactions requiring NAD(P)H regeneration 1.1.1.95 phosphoglycerate dehydrogenase synthesis metabolic engineering of Corynebacterium glutamicum for L-serine production by enzyme overexpression 1.1.1.100 3-oxoacyl-[acyl-carrier-protein] reductase synthesis coexpression with fabH mutant F87T and polyhydroxyalkanoate synthase genes enhances the production of short chain length-medium chain length polyhydroxyalkanoate copolymer from both related and unrelated carbon sources. Analysis of polyhydroxyalkanoate accumulation and physical characterization of copolymer 1.1.1.103 L-threonine 3-dehydrogenase synthesis over-expression of a feedback-resistant threonine operon thrA*BC, with deletion of the genes that encode threonine dehydrogenase tdh and threonine transporters tdcC and sstT, and introduction of a mutant threonine exporter rhtA23 in Escherichia coliMDS42. The resulting strain shows about 83% increase in L-threonine production when cells are grown by flask fermentation, compared to a wild-type Escherichia coli strain MG1655 engineered with the same threonine-specific modifications described above 1.1.1.105 all-trans-retinol dehydrogenase (NAD+) synthesis under optimized conditions, the enzyme produces 600 mg all-trans-retinol per l after 3 h, with a conversion yield of 27.3% (w/w) and a productivity of 200 mg per l and h 1.1.1.108 carnitine 3-dehydrogenase synthesis immobilized in nanofiltration membrane bioreactor for the continuous production of L-carnitine 1.1.1.116 D-arabinose 1-dehydrogenase (NAD+) synthesis production of L-ascorbic acid and secretion into culture medium by overexpression of enzyme and arabinone-1,4-lactone oxidase in Saccharomyces cerevisiae and Zygosaccharomyces bailii 1.1.1.118 glucose 1-dehydrogenase (NAD+) synthesis glucose dehydrogenase and L-carnitine dehydrogenase are coimmobilized in a nanofiltration membrane bioreactor for the continuous production of 1-carnitine from 3-dehydrocarnitine with NADH regeneration 1.1.1.119 glucose 1-dehydrogenase (NADP+) synthesis use of enzyme in enzyme-catalyzed synthesis system for poly(3-hydroxybutyrate), enzyme catalyzes regeneration of NADPH, system yields 5.6 mg of poly(3-hydroxybutyrate), in a 5 ml-reaction mixture 1.1.1.119 glucose 1-dehydrogenase (NADP+) synthesis Escherichia coli strain expressing both recombinant glucose 1-dehydrogenase and a glucose facilitator for uptake of unphosphorylated glucose shows a nine times higher initial alpha-pinene oxide formation rate corresponding to a sixfold higher yield of 20 mg per g cell dry weight after 1.5 h and to a sevenfold increased alpha-pinene oxide yield in the presence of glucose compared to glucose-free conditions 1.1.1.119 glucose 1-dehydrogenase (NADP+) synthesis glucose dehydrogenase is generally used to regenerate the expensive cofactor NADPH by oxidation of D-glucose to gluconolactone 1.1.1.133 dTDP-4-dehydrorhamnose reductase synthesis development and evaluation of a modular system for large scale production of important dTDP-activated deoxyhexoses from dTMP and sucrose, overview 1.1.1.133 dTDP-4-dehydrorhamnose reductase synthesis six enzymes, including the dTDP-4-keto-rhamnose reductase, are involved in the pathway and are prepared by recombinant expression in Escherichia coli for large scale production of O-antigen precursor sTDP-L-rhamnose in a one-pot reaction, overview 1.1.1.135 GDP-6-deoxy-D-talose 4-dehydrogenase synthesis use of enzyme for synthesis of GDP-deoxyhexoses 1.1.1.138 mannitol 2-dehydrogenase (NADP+) synthesis commercial mannitol production as alternatives to less efficient chemical reduction of fructose 1.1.1.138 mannitol 2-dehydrogenase (NADP+) synthesis the enzyme of strain strain HH-01, KCCM-10252, is useful in production of D-mannitol 1.1.1.140 sorbitol-6-phosphate 2-dehydrogenase synthesis constitutive expression of the two sorbitol-6-phosphate dehydrogenase genes srlD1 and srlD2 in a mutant strain deficient for both L- and D-lactate dehydrogenase activities. Both Stl6PDH enzymes are active, and high specific activity can be detected in the overexpressing strains. Using resting cells under pH control with glucose as a substrate, both Stl6PDHs are capable of rerouting the glycolytic flux from fructose-6-phosphate toward sorbitol production with a remarkably high efficiency of 61 to 65% glucose conversion 1.1.1.144 perillyl-alcohol dehydrogenase synthesis expression of the genes for (-)-limonene synthase (SdLS), a limonene 7-hydroxylase (SdL7H, CYP71A76), a perillyl alcohol dehydrogenase (SdPOHDH) and perillic acid O-methyltransferase (SdPAOMT) in Nicotiana benthamiana in combination with a geranyl diphosphate synthase to boost precursor formation, results in production of methylperillate 1.1.1.159 7alpha-hydroxysteroid dehydrogenase synthesis the enzyme is useful as biocatalyst in the reduction of 7-keto bile acids, co-operation with cholylglycine hydrolase, EC 3.5.1.24, overview 1.1.1.159 7alpha-hydroxysteroid dehydrogenase synthesis the enzyme is useful in production of ursodeoxycholic acid, a secondary bile acid, which is used as a drug for the treatment of various liver diseases 1.1.1.175 D-xylose 1-dehydrogenase synthesis expression of gene xylB in Saccharomyces cerevisiae results in production of 17 g D-xylonate/l at 0.23g/l/h from 23 g D-xylose/l. D-Xylonate accumulates intracellularly to 70 mg/g, xylitol to 18 mg/g. Cells expressing D-xylonolactone lactonase xylC from Caulobacter crescentus with xylB initially produce more extracellular D-xylonate than cells lacking xylC at both pH5.5 and pH3, and sustain higher production at pH3. Cell vitality and viability decreases during D-xylonate production at pH 3.0 1.1.1.176 12alpha-hydroxysteroid dehydrogenase synthesis the enzyme is useful in production of ursodeoxycholic acid, a secondary bile acid, which is used as a drug for the treatment of various liver diseases 1.1.1.179 D-xylose 1-dehydrogenase (NADP+, D-xylono-1,5-lactone-forming) synthesis production of up to19 g D-xylonate per litre in Kluyveromyces lactis expressing gene xyd1 upon growth on D-galactosel and D-xylose. D-Xylose uptake is not affected by deletion of either the D-xylose reductase XYL1 or a putative xylitol dehydrogenase encoding gene XYL2 in xyd1 expressing strains 1.1.1.184 carbonyl reductase (NADPH) synthesis enzyme is useful for production of ethyl (S)-4-chloro-3-hydroxybutanoate, which is used in the synthesis of pharmacologically and biologically important compouds 1.1.1.184 carbonyl reductase (NADPH) synthesis enzyme might be useful for production of ethyl (S)-4-chloro-3-hydroxybutanoate, which is used in the synthesis of pharmacologically and biologically important compounds 1.1.1.184 carbonyl reductase (NADPH) synthesis semienzymatic production of (R)-3-styrene oxide 1.1.1.184 carbonyl reductase (NADPH) synthesis the enzyme is useful for stereoselective enzymatic synthesis of chiral alcohols and chiral alcohol intermediates of pharmaceutical importance 1.1.1.184 carbonyl reductase (NADPH) synthesis production of carbonyl reductase by Candida viswanathii in 6.6 l fermentor, using controlled pH value at 8.0, aeration rate 1 vvm and an agitation speed of 250 rpm at 25°C. Use of enzyme for preparative scale reduction of N,N-dimethyl-3-keto-2-thienyl-propanamine to (S)-N,N-dimethyl-3-keto-2-thienyl-propanamine 1.1.1.184 carbonyl reductase (NADPH) synthesis production of Geotrichum candidum carbonyl reductase in a laboratory scale stirred tank bioreactor, optimization of conditions. At controlled pH value of 5.5 the specific enzyme activity is highest with 306 U/mg. Optimization of glucose concentration yields 21 g/l cell mass with 9770 U enzyme activity/g glucose 1.1.1.184 carbonyl reductase (NADPH) synthesis synthesis of (R)-beta-hydroxynitriles with good optical purity by use of recombinant carbonyl reductase and further conversion to (R)-beta-hydroxy carboxylic acids via a nitrilase-catalyzed hydrolysis 1.1.1.184 carbonyl reductase (NADPH) synthesis enzyme product ethyl (S)-4-chloro-3-hydroxybutanoate is a chiral compound valuable as a building block for pharmaceuticals 1.1.1.184 carbonyl reductase (NADPH) synthesis synthesis of (3S)-acetoin in a coupled system consisting of glucose dehydrogenase from Bacillus subtilis 168 and the NADPH-dependent carbonyl reductase. Under the optimal conditions, 12.2 g/l (3S)-acetoin is produced from 14.3 g/l diacetyl in 75 min 1.1.1.184 carbonyl reductase (NADPH) synthesis optimum substrate, 2,2,2-trifluoroacetophenone, is asymmetrically reduced in a coupled NADPH-regeneration system with an enantioselectivity of 99.8% and a conversion of 98% 1.1.1.184 carbonyl reductase (NADPH) synthesis coexpression with glucose dehydrogenase from Bacillus subtilis for NADPH regeneration in Escherichia coli and optimization of linker peptides used for the fusion expression of carbonyl reductase and glucose dehydrogenase. Up to 297.3 g/L (R)-[3,5-bis(trifluoromethyl)phenyl] ethanol with enantiopurity >99.9% ee is produced via reduction of 1.2 M of substrate with a 96.7% yield and productivity of 29.7 g/(L h) 1.1.1.184 carbonyl reductase (NADPH) synthesis in situ expression in Candida leads to over fourfold higher activity for conversion of racemic (R,S)-1-phenyl-1,2-ethanediol to 2-hydroxyacetophenone, while maintaining the activity for catalyzing 2-hydroxyacetophenone to (S)-1-phenyl-1,2-ethanediol. A recombinant Candida parapsilosis converts racemic (R,S)-1-phenyl-1,2-ethanediol to its (S)-isomer with an optical purity of 98.8% and a yield of 48.4%. The biotransformation duration is reduced from 48 to 13 h 1.1.1.188 prostaglandin-F synthase synthesis development of a coupled assay method for enzymatic formation of prostamide F2alpha from anandamide by the cyclooxygenase-II and the prostaglandin synthase F involving the intermediate metabolite, prostamide H2 1.1.1.194 coniferyl-alcohol dehydrogenase synthesis the recombinant Rhodococcus opacus strain PD630, expressing the coniferyl alcohol dehydrogenase from Rhodococcus sp. strain HR199, together with the coniferyl aldehyde dehydrogenase, and the vanillyl alcohol oxidase, the latter from Penicillium simplicissimus strain CBS, is able to produce vanillin from ferulic acid and eugenol 1.1.1.195 cinnamyl-alcohol dehydrogenase synthesis synthesis of cinnamyl alcohol from cinnamaldehyde using the isolated enzyme or by expression of enzyme in Escherichia coli. The reduction of cinnamaldehyde by the isolated enzyme occurrs in 3 h at 50°C with 97% conversion, and yields high purity cinnamyl alcohol with a yield of 88% and a productivity of 50 g/g enzyme. The reduction of 12.5 g/l cinnamaldehyde by whole cells in 6 h, at 37 °C and no requirement of external cofactor occurrs with 97% conversion, 82% yield of 98% pure alcohol and a productivity of 34 mg/g wet cell weight 1.1.1.195 cinnamyl-alcohol dehydrogenase synthesis potential exploitation of rationally engineered forms of CAD2 for the targeted modification of monolignol composition in transgenic plants 1.1.1.198 (+)-borneol dehydrogenase synthesis production of enzyme in the form of inclusion body in Escherichia coli. The refolded BDH1 tends to precipitate. Insoluble recombinant BDH1 is converted into a soluble form by adding glycerol in LB medium 1.1.1.198 (+)-borneol dehydrogenase synthesis recombinant borneol dehydrogenase is found in inclusion bodies when expressed in Escherichia coli. Changing the medium from lysogeny broth to Terrific Broth yield a soluble form of the enzyme 1.1.1.201 7beta-hydroxysteroid dehydrogenase (NADP+) synthesis the enzyme is useful in production of ursodeoxycholic acid, a secondary bile acid, which is used as a drug for the treatment of various liver diseases 1.1.1.202 1,3-propanediol dehydrogenase synthesis Klebsiella pneumoniae is used for production of 1,3-propanediol by the enzyme which is utilized in plastic industry 1.1.1.202 1,3-propanediol dehydrogenase synthesis the enzyme is useful in 1,3-propanediol production from glycerol, a byproduct in biodiesel production, by an enzymatic bioconversion in a membrane reactor in which the NAD+ coenzyme can be regenerated, mathematical description and modelling of the system, overview 1.1.1.202 1,3-propanediol dehydrogenase synthesis constitutive overexpression of 1,3-PD oxidoreductase in Klebsiella pneumoniae leads to a nearly 3fold decrease in peak level of the intermediary metabolite 3-hydroxypropionaldehyde and an increase of 16.5% in yield of 1,3-propanediol with respect to the wild-type strain 1.1.1.202 1,3-propanediol dehydrogenase synthesis expression of the yqhD gene, encoding 3-propanediol oxidoreductase isoenzyme from Escherichia coli and the dhaT gene, encoding 3-propanediol oxidoreductase from Klebsiella pneumoniae individually and concomitantly in Klebsiella pneumoniae using the double tac promoter expression plasmid pEtac-dhaT-tac-yqhD. The three resultant recombinant strains show that the peak values for 3-hydroxypropionaldehyde production in broth of the three recombinant strains are less than 25% of that of the parent strain. Expression of dhaT reduces formation of by-products ethanol and lactic acid and increases molar yield of 1,3-propanediol slightly, while expression of yqhD does not enhance molar yield of 1,3-propanediol, but increases ethanol concentration in broth as NADPH participation in transforming 3-hydroxypropionaldehyde to 1,3-propanediol allows more cellular NADH to be used to produce ethanol. Co-expression of both genes therefore decreases by-products and increases the molar yield of 1,3-propanediol by 11.8%, by catalyzing 3-hydroxypropionaldehyde conversion to 1,3-propanediol using the two cofactors NADH and NADPH 1.1.1.202 1,3-propanediol dehydrogenase synthesis genetic engineering of Klebsiella pneumoniae for production of 1.3-propanediol from glycerol. Constitutive expression of the dhaT gene alone gives the highest yield, fed-batch fermentation results in efficient production of 1,3-propanediol from either pure or crude glycerol, without by-product formation 1.1.1.202 1,3-propanediol dehydrogenase synthesis the enzyme can be used for 1,3-propandiol synthesis for use in resaerch and industrial applications, especially in biodiesel industry, but also as a monomer for polycondensation to manufacture plastics with special properties, i.e., polyesters, polyethers, polyurethanes, and polytrimethylene terephthalate as a monomer for cyclic compounds, and as a polyglycol-type lubricant 1.1.1.202 1,3-propanediol dehydrogenase synthesis Escherichia coli is engineered to produce 1,3-propanediol from glycerol, an inexpensive carbon source. This is done by introducing a synthetic 1,3-propanediol production pathway in recombinant Escherichia coli consisting of glycerol dehydratase complex (dhaB123) and glycerol dehydratase reactivation factors (gdrAB) from Klebsiella pneumoniae and 1,3-propanediol oxidoreductase isoenzyme (yqhD) from Escherichia coli. When 10 mM succinate is added to the medium, the titer of 1,3-propanediol and the glycerol consumption increase to 9.9 and 23.84 g/l, respectively. In addition, the ratio of NADH to NAD+ increases by 43%. Succinate addition is a promising route for the biotechnological production of NADH-dependent microbial metabolites 1.1.1.202 1,3-propanediol dehydrogenase synthesis NADH-dependent 1,3-propanediol oxidoreductase is a key enzyme for the production of 1,3-propanediol in soluble cell extract. Klebsiella pneumoniae J2B shows a high potential for the production of 1,3-propanediol from glycerol. Optimization of the culture conditions and the elimination of lactate synthesis improves 1,3-propanediol production significantly 1.1.1.206 tropinone reductase I synthesis co-expression of putrescine N-methyltransferase and tropinone reductase I genes in Anisodus acutangulus hairy roots significantly improves the yields of tropinone alkaloids and shows higher antioxidant activity than control 1.1.1.206 tropinone reductase I synthesis transgenic hairy root lines expressing both tropinone reductase I and hyoscyamine-6beta-hydroxylase produce significantly higher levels of tropinone alkaloids compared with the control and single gene transformed lines, reaching up to 4.293 mg/g tropinone alkaloids. In addition, the content of anisodine is also greatly improved. The average content of anisodine in double transformed lines is 0.984 mg/g dry weight, about 18fold of control lines 1.1.1.206 tropinone reductase I synthesis enzyme PtTRI is a potential candidate for modifying the TAs biosynthetic pathway on TRI to obtain high yielding scopolamine medicinal plants 1.1.1.207 (-)-menthol dehydrogenase synthesis the enzyme is useful, together with menthone:(+)-neomenthol reductase (EC 1.1.1.208), for production of (1R,2S,5R)-(-)-menthol and (1S,2S,5R)-(+)-neomenthol from pulegone, development of a one-pot synthesis method, overview. Menthol isomers, (1R,2S,5R)-(-)-menthol, (1R,2S,5S)-(+)-isomenthol, (1S,2S,5R)-(+)-neomenthol, and (1R,2R,5R)-(+)-neoisomenthol, and carvone are used as additives in oral hygiene products and flavors in food and beverages, or cosmetics 1.1.1.208 (+)-neomenthol dehydrogenase synthesis the enzyme is useful, together with menthone:(-)-menthol reductase (EC 1.1.1.207), for production of (1R,2S,5R)-(?)-menthol and (1S,2S,5R)-(+)-neomenthol from pulegone, development of a one-pot synthesis method, overview. Menthol isomers, (1R,2S,5R)-(?)-menthol, (1R,2S,5S)-(+)-isomenthol, (1S,2S,5R)-(+)-neomenthol, and (1R,2R,5R)-(+)-neoisomenthol, and carvone are used as additives in oral hygiene products and flavors in food and beverages, or cosmetics 1.1.1.213 3alpha-hydroxysteroid 3-dehydrogenase (Re-specific) synthesis the enzyme is useful in reductive production of steroids. In a coupled-enzyme system comprising HSDH and formate dehydrogenase, a twofold increase in production rate of androsterone is obtained when utilizing 1-butyl-3-methylimidazolium L-lactate with NADH regeneration 1.1.1.214 2-dehydropantolactone reductase (Si-specific) synthesis enzyme is useful for production of chiral alcohols 1.1.1.214 2-dehydropantolactone reductase (Si-specific) synthesis in a continuous feeding reaction, 200 mM ketopantolactone is reduced to (R)-pantolactone with 98% conversion and 99% enantiomeric excess within 2.0 h 1.1.1.214 2-dehydropantolactone reductase (Si-specific) synthesis use of polyketone reductase (CPR), glucose dehydrogenase (GDH) and coenzyme NADP+ in organic-inorganic hybrid nanoflowers (hNFs) for the asymmetric reduction of ketopantolactone to synthesize (R)-(-)-pantolactone. The sodium alginate-coated hNF reactor successfully catalyzes the asymmetric synthesis of (R)-pantolactone, with satisfactory stereoselectivity and reusability in repeated batches 1.1.1.214 2-dehydropantolactone reductase (Si-specific) synthesis whole-cell biotransformation process to produce D-pantolactone in a biphasic reaction system. Recombinant CPR and glucose dehydrogenase are co-expressed in Escherichia coli to simultaneously achieve the synthesis of D-PL and the regeneration of NADPH. Presence of 15% dichloromethane significantly inhibits the hydrolysis of ketopantolactone. In a fed-batch system, the D-pantolactone concentration reaches 0.77 mol per l in the reaction mixture at 7 h, and its enantiomeric excess is 99% 1.1.1.215 gluconate 2-dehydrogenase synthesis ga2dh-overexpressing strain Gluconobacter oxydans_tufB_ga2dh is a useful species for use in 2-dehydro-D-gluconate production 1.1.1.215 gluconate 2-dehydrogenase synthesis pulse addition of hydrogen peroxide to synthetic media is beneficial to improve the conversion rate of glucose to 2-oxo-gluconate by 1.47fold 1.1.1.227 (-)-borneol dehydrogenase synthesis production of enzyme in the form of inclusion body in Escherichia coli. The refolded BDH1 tends to precipitate. Insoluble recombinant BDH1 is converted into a soluble form by adding glycerol in LB medium 1.1.1.243 carveol dehydrogenase synthesis evaluation of large scale production of (-)-carvone by cells of Rhodococcus erythropolis strain DCL14, overview 1.1.1.245 cyclohexanol dehydrogenase synthesis synthesis of lactones from cycloalkanes. A heterologous pathway comprising enzymes with compatible kinetics is designed in Pseudomonas taiwanensis VLB120 enabling in-vivo cascade for synthesizing lactones from cycloalkanes. The respective pathway includes cytochrome P450 monooxygenase (CHX), cyclohexanol dehydrogenase (CDH), and cyclohexanone monooxygenase (CHXON) from Acidovorax sp. CHX100. Resting cells of the recombinant host Pseudomonas taiwanensis VLB120 convert cyclohexane, cyclohexanol, and cyclohexanone to epsilon-caprolactone at 22, 80-100, and 170 U/g cell dry weight, respectively. Cyclohexane (5 mM) is completely converted with a selectivity of 65% for epsilon-caprolactone formation in 2 h without accumulation of intermediate products 1.1.1.247 codeinone reductase (NADPH) synthesis Papaver bracteatum hairy roots expressing CodR gene have a high potential to produce morphinan alkaloids 1.1.1.248 salutaridine reductase (NADPH) synthesis the Papaper somniferum enzymes salutaridine synthase (SAS), salutaridine reductase (SAR) and salutaridinol acetyltransferase (SAT) are functionally co-expressed in Saccharomyces cerevisiae and optimization of the pH conditions allows for productive spontaneous rearrangement of salutaridinol-7-O-acetate and synthesis of thebaine from (R)-reticuline. Upon reconstitution of a 7-gene pathway for the production of codeine and morphine from (R)-reticuline, activity of salutaridine reductase and codeine-O-demethylase likely limit flux to morphine synthesis 1.1.1.250 D-arabinitol 2-dehydrogenase synthesis the enzyme catalyzed oxidation of D-arabitol to D-ribulose which is a rare ketopentose sugar that has numerous industrially applications. D-Arabitol 2-dehydrogenase from Thermotoga maritima has the potential to be a useful biocatalyst for the production of D-ribulose starting from the inexpensive D-arabitol due to its regiospecificity and thermostability 1.1.1.254 (S)-carnitine 3-dehydrogenase synthesis Agrobacterium radiobacter can be used for conversion of the waste product L(-)-carnitine to poly-beta-hydroxybutyrate valuable as a biodegradable and biocompatible plastic 1.1.1.260 sulcatone reductase synthesis preparation of sulcatol enantiomers 1.1.1.270 3beta-hydroxysteroid 3-dehydrogenase synthesis used of mutated 3-ketosteroid reductase (Erg27) to obtain radioactive and non-radioactive intermediates of sterol biosynthesis hardly or not available commercially. The strategies are either incubation of cell homogenates of an Erg27-deletant strain with radioactive lanosterol, or incubation of growing cells of a strain expressing point-mutated 3-ketosteroid reductase with radioactive acetate. Chemical reduction of both radioactive and non-radioactive 3-keto sterones gives the physiological 3-beta-OH sterols, as well as the non-physiological 3-alpha-OH isomers 1.1.1.271 GDP-L-fucose synthase synthesis four genes (manB, manC, gmd, and wcaG) cloned from Escherichia coli are expressed in Lactococcus lactis. The one-pot reaction of ManB, ManC, Gmd, and WcaG with mannose-6-P results in the successful production of GDP-L-fucose 1.1.1.274 2,5-didehydrogluconate reductase (2-dehydro-D-gluconate-forming) synthesis enzyme may be useful for the synthesis of the ascorbate precursor 2-keto-L-gulonate 1.1.1.274 2,5-didehydrogluconate reductase (2-dehydro-D-gluconate-forming) synthesis enzyme can be used in the industrial production of vitamin C 1.1.1.274 2,5-didehydrogluconate reductase (2-dehydro-D-gluconate-forming) synthesis enzymatic production of the vitamin C precursor 2-keto-L-gulonate (2-KLG) from 2,5-diketo-D-gluconate (2,5-DKG) by coupling 2,5-diketo-D-gluconic acid reductase via its coenzyme to glucose dehydrogenase. The bienzymatic process shows complicated inhibition patterns caused by reaction products, NADP+ and NADPH. The key parameters for a fast and efficient conversion are the NADP(H) concentration, the volumetric activity of 2,5-DKG reductase, the ratio of synthetic enzyme activity to regenerate enzyme activity and the glucono-1,5-lactone concentration. By modeling the space-time yield of the process is nearly doubled and the coenzyme concentration reduced threefold 1.1.1.280 (S)-3-hydroxyacid-ester dehydrogenase synthesis ethyl (2R,3S)-3-hydroxy-2-methylbutanoate is a useful starting material for the synthesis of ferroelectric liquid crystal compounds and various other biologically active substances 1.1.1.281 GDP-4-dehydro-6-deoxy-D-mannose reductase synthesis generating GDP-rhamnose for in vitro rhamnosylation of glycoproteins and glycopeptides 1.1.1.281 GDP-4-dehydro-6-deoxy-D-mannose reductase synthesis the functional expression of the Pseudomonas aeruginosa enzyme in Saccharomyces cerevisiae will provide a tool for generating GDP-rhamnose for in vitro rhamnosylation of glycoprotein and glycopeptides 1.1.1.281 GDP-4-dehydro-6-deoxy-D-mannose reductase synthesis generation of GDP-rhamnose by coexpression of enzyme with GDP-mannose-4,6-dehydratase in Saccharomyces cerevisiae 1.1.1.282 quinate/shikimate dehydrogenase [NAD(P)+] synthesis enzyme YdiB displays a clear activity on quinate, with either NADP+ or NAD+ as a cofactor in addition to shikimate, making this enzyme an important candidate for quinate production. Production of shikimate, quinate, 3-dehydroshikimate and other aromatic derivatives in strain PB12.SA22 during batch culture fermentation 1.1.1.283 methylglyoxal reductase (NADPH) synthesis engineeering of Sacharomyces cerevisiae for specific reduction of a variety of carbonyl compounds by altering the levels of fatty acid synthase Fas, aldo-keto reductase Ypr1 and R-acetoxy ketone reductase Gre2. By choosing the appropriate engineered yeast strain, it is possible to prepare many of the alcohols in high stereochemical purities and good chemical yields. For example, in the reductions of some alpha-unsubstituted beta-keto esters, genetically disabling Fas virtually eliminates production of (3R)-alcohols. Overexpressing (3S)-selective reductase Gre2 in the Fas knockout strain gives yeast cells that reduce these ketones to the corresponding (3S)-alcohols in >98% ee in each case 1.1.1.289 sorbose reductase synthesis usage of a substrate-coupled biocatalytic process driven by an NADPH-dependent sorbose reductase from Candida albicans for the asymmetric reduction of ethyl 4-chloro-3-oxobutanoate 1.1.1.298 3-hydroxypropionate dehydrogenase (NADP+) synthesis developments in 3-hydroxypropionate (HP) production involving the enzyme using Saccharomyces cerevisiae as an industrial host. By combining genome-scale engineering tools, malonyl-CoA biosensors and optimization of downstream fermentation, the production of 3-HP in yeast has the potential to reach or even exceed the yield of chemical production 1.1.1.298 3-hydroxypropionate dehydrogenase (NADP+) synthesis the enzyme is very useful for 3-hydroxypropionate biosynthesis and production 1.1.1.304 diacetyl reductase [(S)-acetoin forming] synthesis acetoin(diacetyl) reductase, i.e. 2,3-butanediol dehydrogenase, is one of the key enzymes in the microbial production of 2,3-butanediol, a platform with extensive industrial applications in the production of plastics, printing inks, perfumes, fumigants, spandex, moistening and softening agents, plasticizers, and pharmaceutical carrier 1.1.1.304 diacetyl reductase [(S)-acetoin forming] synthesis the enzyme is used for production of S-acetoin with higher than 99.9% optical purity from diacetyl using whole cells of engineered Escherichia coli 1.1.1.305 UDP-glucuronic acid dehydrogenase (UDP-4-keto-hexauronic acid decarboxylating) synthesis engineering of Escherichia coli to ynthesize the plant-specific flavonoid O-pentosides quercetin 3-O-xyloside and quercetin 3-O-arabinoside. For UDP-xylose biosynthesis, genes UXS (UDP-xylose synthase) from Arabidopsis thaliana and ugd (UDP-glucose dehydrogenase) from E.scherichia coli, are overexpressed. The gene encoding ArnA, which competes with UXS for UDP-glucuronic acid, is deleted. For UDP-arabinose biosynthesis, UXE (UDP-xylose epimerase) is overexpressed. UDP-dependent glycosyltransferases are engineered to ensure specificity for UDP-xylose and UDP-arabinose. The srains thus obtained synthesize approximately 160 mg/liter of quercetin 3-O-xyloside and quercetin 3-O-arabinoside 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis D-xylose is the second most abundant renewable sugar in nature, and its fermentation to ethanol has great economical potential. Unfortunately, Saccharomyces cerevisiae, which has been optimized for ethanol production, cannot utilize xylose efficiently, while D-xylulose, an isomerization product of D-xylose, can be assimilated. A major strategy for constructing xylose-fermenting Saccharomyces cerevisiae is to introduce genes involved in xylose metabolism from other organisms. Xylose reductase and xylitol dehydrogenase (EC 1.1.1.9) from the xylose-fermenting yeast Pichia stipitis are cloned into Saccharomyces cerevisiae to allow xylose fermentation to ethanol. In this case, xylose is converted into xylulose by the sequential actions of two oxidoreductases. First, Pichia stipitis xylose reductase catalyses the reduction of xylose into xylitol with NAD(P)H as co-substrate. Xylitol is then oxidized by PsXDH (Pichia stipitis xylitol dehydrogenase) which uses NAD+ exclusively as co-substrate to yield xylulose. The different coenzyme specificity of the two enzymes xylose reductase and xylitol dehydrogenase, however, creates an intracellular redox imbalance, which results in low ethanol yields and considerable xylitol by-product formation. A mutant is constructed that shows an altered active site that is more unfavorable for NADPH than NADH in terms of both Km and kcat. There are potentials for application of the mutant (K270S/N272P/S271G/R276F) in constructing a more balanced xylose reductase/xylitol dehydrogenase pathway in recombinant xylose-fermenting Saccharomyces cerevisiae strains 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis production of xylitol 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis the cofactor preference of Pichia stipitis xylose reductase is altered by site-directed mutagenesis. When the K270R xylose reductase is combined with a metabolic engineering strategy that ensures high xylose utilization capabilities, a recombinant Saccharomyces cerevisiae strain is created that provides a unique combination of high xylose consumption rate, high ethanol yield and low xylitol yield during ethanolic xylose fermentation 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis this enzyme is one of the most active xylose reductases and may be used for the in vitro production of xylitol 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis expression in Candida glycerinogenes WL2002-5, the recombinant strain produces xylitol from D-xylose using glycerol as a cosubstrate for cell growth and NAD (P) H regeneration. 100 g/L D-xylose is completely converted into xylitol when at least 20 g/l glycerol is used as a co-substrate. The strain accumulates 2.1fold increased xylitol concentration over those cells grown on glucose as co-substrate, with a volumetric productivity of 0.83 g/l/h, and a xylitol yield of 98% xylose 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis the microalga Chlorella sorokiniana and provide a target for genetic engineering to improve D-xylose utilization for microalgal lipid production 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis production of bio-xylitol from D-xylose by an engineered Pichia pastoris expressing a recombinant xylose reductase 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis the enzyme can be used for industrial production of xylitol from waste xylose mother liquor. Production of xylitol from xylose mother liquor by immobilized Escherichia coli cells containing xylose reductase and glucose dehydrogenase, overview. The immobilization of Escherichia coli strain BL21(DE3)/ pCDFDuet-1-XR-GDH cells results in an increase of storage stability which can expand its potential applications in industries 1.1.1.307 D-xylose reductase [NAD(P)H] synthesis Escherichia coli expressing XylA synthesizes 13.3 g/l of xylitol during 20 h of cultivation, when D-xylose (50 g/l) and D-glucose (5 g/l) are added to IPTG-induced cells 1.1.1.311 (S)-1-phenylethanol dehydrogenase synthesis application of (S)-1-phenylethanol dehydrogenase for asymmetric reduction of 42 prochiral ketones and 11 beta-keto esters to enantiopure secondary alcohols. The conversions are carried out in a batch reactor using recombinant Escherichia coli as whole-cell catalysts and isopropanol as reaction solvent and cosubstrate for NADH recovery. Ketones are converted to the respective secondary alcohols with excellent enantiomeric excesses and high productivities 1.1.1.311 (S)-1-phenylethanol dehydrogenase synthesis the enzyme recombinantly expressed in Escherichia coli is used for chemoenzymatic synthesis of enantiomerically enriched (S)- and (R)-gamma-aryl-gamma-butyrolactones, whereby the key step is an enzyme-catalyzed stereoselective reduction of methyl 4-oxo-4-arylbutanoates, (S)-PED-catalyzed bioreduction in preparative scale 1.1.1.320 benzil reductase [(S)-benzoin forming] synthesis asymmetric reduction with Bacillus cereus benzil reductase can be utilized to produce important chiral compounds 1.1.1.321 benzil reductase [(R)-benzoin forming] synthesis the enzyme can be used for the highly enantioselective reduction of benzil to produce (R)-benzoin 1.1.1.327 5-exo-hydroxycamphor dehydrogenase synthesis synthetic route for the synthesis of bifunctional camphor derivatives. The combination of the enzymatic P450cam system with 5-exo-hydroxycamphor dehydrogenase (FdeH) allows an efficient synthesis of 2,5-diketobornane, which can be used for further derivatisation 1.1.1.335 UDP-N-acetyl-2-amino-2-deoxyglucuronate dehydrogenase synthesis enzyme WbpB and the related enzymes of the B-band O-antigen pathway of Pseudomonas aeruginosa lipopolysaccharide, WbpA, WbpE, WbpD and WbpI, can be combined in vitro to generate UDP-ManNAc(3NAc)A in a single reaction vessel, thereby providing supplies of this complex glycosyl donor for future studies of lipopolysaccharide assembly 1.1.1.335 UDP-N-acetyl-2-amino-2-deoxyglucuronate dehydrogenase synthesis synthesis of preparative quantities of 2-acetamido-3-amino-2,3-dideoxy-D-glucuronic acid and 2,3-diacetamido-2,3-dideoxy-D-glucuronic acid by coupled reaction of enzyme WbpB and transaminase WpbE 1.1.1.337 L-2-hydroxycarboxylate dehydrogenase (NAD+) synthesis the enzyme can be applied in an industrial process for the production of L-amino acids 1.1.1.345 D-2-hydroxyacid dehydrogenase (NAD+) synthesis the enzyme can be utilized for preparation of enantiomerically pure (R)-2-hydroxy-4-methylpentanoate in 88% yield, using formate dehydrogenase recycling of the NADH coenzyme 1.1.1.346 2,5-didehydrogluconate reductase (2-dehydro-L-gulonate-forming) synthesis 2,5-DKG reductase is an integral part of several industrial processes designed to synthesize 2-dehydro-L-gulonate based on the 2,5-diketo-D-gluconic acid pathway 1.1.1.358 2-dehydropantolactone reductase synthesis the enzyme has a potential application in the asymmetric synthesis of optically active (R)-pantothenate, synthetic method of (R)-pantothenate production through the stereoselective reduction of oxopantoyl lactone (KPL) by aldo-keto reductase (AKR). (R)-Pantolactone (PL) is a key chiral building block for the synthesis of calcium (R)-pantothenate (vitamin B5), (R)-panthenol, and (R)-pantetheine, which are used as food and feed additives, as well as ingredients in pharmaceutical and cosmetic compositions 1.1.1.358 2-dehydropantolactone reductase synthesis the recombinant CduCPR from Candida dubliniensis exhibits potential application in the asymmetric synthesis of (R)-pantolactone 1.1.1.363 glucose-6-phosphate dehydrogenase [NAD(P)+] synthesis bacitracin, a kind of cyclic peptide antibiotic mainly produced by Bacillus, has wide ranges of applications. The bacitracin production is enhanced by by NADPH generation via overexpressing glucose-6-phosphate dehydrogenase Zwf in Bacillus licheniformis 1.1.1.365 D-galacturonate reductase synthesis L-galactonate shows potential applications as an additive to nutrients and cosmetics. AnGar1 variants are developed that represent the a GalA reductases with a higher preference for NADH compared to NADPH. The altered cofactor-specificity enables the coupling of GalA reduction to glycolysis, resulting in higher yields of GalOA when glucose is used as a redox donor. The engineered AnGar1 should prove valuable for D-galacturonic acid utilization in pectin-rich hydrolysates, which contain neutral sugars such as glucose, galactose, or arabinose, all of which are funneled into glycolysis. The NADH-dependent GalA reductases can facilitate the coupling of L-galactonate production to the oxidation of glycerol, an abundant waste product that can be supplemented to pectin-rich hydrolysates 1.1.1.379 (R)-mandelate dehydrogenase synthesis D-mandelate dehydrogenase (DMDH) has the potential to convert D-mandelic acid to phenylglyoxylic acid (PGA), which is a key building block in the field of chemical synthesis and is widely used to synthesize pharmaceutical intermediates or food additives. Development of an alternative strategy for the chiral resolution of racemic mandelic acid and the biosynthesis of PGA 1.1.1.406 galactitol 2-dehydrogenase (L-tagatose-forming) synthesis preparation of optically pure aliphatic diols by enzymic oxidation of one enantiomer or stereospecific reduction of keto-alcohols or diketones 1.1.1.406 galactitol 2-dehydrogenase (L-tagatose-forming) synthesis as an enzyme capable of the stereo- and regioselective modification of carbohydrates, GatDH exhibits a high potential for application in biotechnology as a biocatalyst, e.g. preparation of several (R)-1,2-diols by racemic resolution with GatDH as well as the synthesis of several S-configured aliphatic alcohols by reducing corresponding prochiral ketones 1.1.1.406 galactitol 2-dehydrogenase (L-tagatose-forming) synthesis synthesis of (S)-1,2-propanediol and L-tagatose starting from hydroxyacetone and galactitol using an immobilized enzyme system. One-pot purification of His6-tagged GatDH and FDH followed by the production of rare sugar and chiral diol by use of affinity magnetic nanoparticles 1.1.1.406 galactitol 2-dehydrogenase (L-tagatose-forming) synthesis galactitol dehydrogenase is coupled with water-forming NADH oxidase for efficient enzymatic synthesis of L-tagatose (a building block in the production of many value-added chemicals) 1.1.1.406 galactitol 2-dehydrogenase (L-tagatose-forming) synthesis the two oxidoreductases, xylose reductase and galactitol dehydrogenase are functionally expressed in the engineered yeast (EJ2g_pXpG) and enable direct production of tagatose from lactose. The expression levels of the enzymes are adjusted to maximize tagatose production. The resulting engineered yeast produces 37.69 g/L of tagatose from lactose with a tagatose and galactose ratio of 9:1 in the reaction broth 1.1.1.415 noscapine synthase synthesis complete biosynthesis of noscapine and halogenated alkaloids in yeast and optimizing noscapine production toward scalable manufacturing. Engineered strain contains 25 heterologous plant, bacteria, and mammalian genes and 6 mutant or overexpressed yeast genes. The noscapine biosynthetic pathway incorporates seven endomembrane-localized plant enzymes, highlighting the ability of the yeast to functionally express and properly localize large numbers of heterologous enzymes into the endoplasm reticulum. Noscapine titers were improved by 18000fold (to low mg/l levels) via a combination of enzyme engineering, pathway and strain engineering, and fermentation optimization. Microbial fermentation can be used to produce halogenated alkaloid derivatives, which can ultimately serve as potential drug leads, through feeding amino acid derivatives to strains 1.1.1.422 pseudoephedrine dehydrogenase synthesis the wide substrate spectrum of these dehydrogenases, combined with their regio- and enantioselectivity, suggests a high potential for the industrial production of valuable chiral compounds 1.1.1.422 pseudoephedrine dehydrogenase synthesis the enzyme catalyzes the oxidation of an isomers of ephedrine and the regio- and enantioselective reduction of sterically demanding substrate 1-phenyl-1,2-propanedione to give (S)-phenylacetylcarbinol. (S)-phenylacetylcarbinol can serve as a precursor in the synthesis of many pharmaceuticals, such as (+)-(S,S)-pseudoephedrine 1.1.1.422 pseudoephedrine dehydrogenase synthesis the wide substrate spectrum of the dehydrogenase, combined with its regio- and enantioselectivity, suggests a high potential for the industrial production of valuable chiral compounds 1.1.1.423 (1R,2S)-ephedrine 1-dehydrogenase synthesis the wide substrate spectrum of these dehydrogenases, combined with their regio- and enantioselectivity, suggests a high potential for the industrial production of valuable chiral compounds 1.1.1.423 (1R,2S)-ephedrine 1-dehydrogenase synthesis the enzyme catalyzes the oxidation of an isomer of ephedrine and the regio- and enantioselective reduction of sterically demanding substrate 1-phenyl-1,2-propanedione to give (R)-phenylacetylcarbinol 1.1.1.430 D-xylose reductase (NADH) synthesis D-xylose is the second most abundant renewable sugar in nature, and its fermentation to ethanol has great economical potential. Unfortunately, Saccharomyces cerevisiae, which has been optimized for ethanol production, cannot utilize xylose efficiently, while D-xylulose, an isomerization product of D-xylose, can be assimilated. A major strategy for constructing xylose-fermenting Saccharomyces cerevisiae is to introduce genes involved in xylose metabolism from other organisms. Xylose reductase and xylitol dehydrogenase (EC 1.1.1.9) from the xylose-fermenting yeast Pichia stipitis are cloned into Saccharomyces cerevisiae to allow xylose fermentation to ethanol. In this case, xylose is converted into xylulose by the sequential actions of two oxidoreductases. First, Pichia stipitis xylose reductase catalyses the reduction of xylose into xylitol with NAD(P)H as co-substrate. Xylitol is then oxidized by PsXDH (Pichia stipitis xylitol dehydrogenase) which uses NAD+ exclusively as co-substrate to yield xylulose. The different coenzyme specificity of the two enzymes xylose reductase and xylitol dehydrogenase, however, creates an intracellular redox imbalance, which results in low ethanol yields and considerable xylitol by-product formation. A mutant is constructed that shows an altered active site that is more unfavorable for NADPH than NADH in terms of both Km and kcat. There are potentials for application of the mutant (K270S/N272P/S271G/R276F) in constructing a more balanced xylose reductase/xylitol dehydrogenase pathway in recombinant xylose-fermenting Saccharomyces cerevisiae strains 1.1.1.430 D-xylose reductase (NADH) synthesis dual specific xylose reductase (dsXR) has an about 4fold higher specificity for NADH than NADPH. This fact could make this enzyme an interesting candidate to be used in metabolic engineering of the yeast xylose metabolism, likely in Saccharomyces cerevisiae. Increased levels of dsXR activity could contribute to an improvement of ethanol production from D-xylose by reducing the cofactor imbalance of the initial catabolic pathway 1.1.1.430 D-xylose reductase (NADH) synthesis fermentation of mixed glucose-xylose substrates in Saccharomyces cerevisiae strains BP10001 and BP000, expressing Candida tenuis xylose reductase in mutated NADH-preferring form and NADPH-preferring wild-type form, respectively. Glucose and xylose, each at 10 g/l, are converted sequentially. The distribution of fermentation products from glucose is identical for both strains whereas when using xylose, BP10001 shows enhanced ethanol yield and decreased yields of xylitol and glycerol as compared to BP000. Increase in xylose concentration from 10 to 50 g/l results in acceleration of substrate uptake by BP10001 and reduction of the xylitol yield. In mixed substrate batches, xylose is taken up at low glucose concentrations and up to 5fold enhanced xylose uptake rate is found towards glucose depletion 1.1.1.430 D-xylose reductase (NADH) synthesis expression of wild-type Xyl1 or an NADH-specific mutant in Saccharomyces cerevisiae. The Xyl1 mutant decreases the biocatalyst’s performance, suggesting use of the NADPH-preferring wild-type enzyme when (semi-)aerobic conditions are applied. In a bioreactor process, the best-performing strain converts 40 g/l xylose with an initial productivity of 1.16 g/l/h and a xylitol yield of 100% 1.1.1.430 D-xylose reductase (NADH) synthesis in fermentation for citric acid production and xylitol accumulation by using D-xylose as the sole carbon source, a sttrain carrying mutant K274R shows a 2.8fold reduction in xylitol accumulation and 4.5fold increase in citric acid production compared to the wild-type strain 1.1.1.431 D-xylose reductase (NADPH) synthesis dual specific xylose reductase (dsXR) has an about 4fold higher specificity for NADH than NADPH. This fact could make this enzyme an interesting candidate to be used in metabolic engineering of the yeast xylose metabolism, likely in Saccharomyces cerevisiae. Increased levels of dsXR activity could contribute to an improvement of ethanol production from D-xylose by reducing the cofactor imbalance of the initial catabolic pathway 1.1.1.431 D-xylose reductase (NADPH) synthesis the enzyme is useful for xylitol bioproduction, profiles, overview 1.1.1.431 D-xylose reductase (NADPH) synthesis fermentation of mixed glucose-xylose substrates in Saccharomyces cerevisiae strains BP10001 and BP000, expressing Candida tenuis xylose reductase in mutated NADH-preferring form and NADPH-preferring wild-type form, respectively. Glucose and xylose, each at 10 g/l, are converted sequentially. The distribution of fermentation products from glucose is identical for both strains whereas when using xylose, BP10001 shows enhanced ethanol yield and decreased yields of xylitol and glycerol as compared to BP000. Increase in xylose concentration from 10 to 50 g/l results in acceleration of substrate uptake by BP10001 and reduction of the xylitol yield. In mixed substrate batches, xylose is taken up at low glucose concentrations and up to 5fold enhanced xylose uptake rate is found towards glucose depletion 1.1.1.431 D-xylose reductase (NADPH) synthesis expression in Phanerochaete sordida YK-624 results in increased xylitol production and markedly higher xylose reductase activities than in the wild-type strain 1.1.1.431 D-xylose reductase (NADPH) synthesis in fermentation for citric acid production and xylitol accumulation by using D-xylose as the sole carbon source, a sttrain carrying mutant K274R shows a 2.8fold reduction in xylitol accumulation and 4.5fold increase in citric acid production compared to the wild-type strain 1.1.1.431 D-xylose reductase (NADPH) synthesis overexpression of D-xylose reductase Xyl1 gene and antisense inhibition of D-xylulokinase XyiH gene result in increase in xylitol production from 22.8 mM to 24.8 mM 1.1.1.431 D-xylose reductase (NADPH) synthesis expression of wild-type Xyl1 or an NADH-specific mutant in Saccharomyces cerevisiae. The Xyl1 mutant decreases the biocatalyst's performance, suggesting use of the NADPH-preferring wild-type enzyme when (semi-)aerobic conditions are applied. In a bioreactor process, the best-performing strain converts 40 g/l xylose with an initial productivity of 1.16 g/l/h and a xylitol yield of 100% 1.1.2.3 L-lactate dehydrogenase (cytochrome) synthesis the enzyme is potentially important for bioanalytical technologies for highly selective assays of L-lactate in biological fluids and foods 1.1.2.4 D-lactate dehydrogenase (cytochrome) synthesis Pseudomonas stutzeri containing D-lactate dehydrogenase can act as a novel biocatalyst for pyruvate production from DL-lactate 1.1.3.2 L-lactate oxidase synthesis biocascade synthesis of L-tyrosine derivatives by coupling a thermophilic tyrosine phenol-lyase and L-lactate oxidase. o-Phenol derivatives are transformed into the corresponding L-tyrosine derivatives with excellent stereoselectivity and high yields using an efficient one-pot, two-step cascade containing thermophilic tyrosine phenol-lyase mutants from Symbiobacterium toebii and L-lactate oxidase from Aerococcus viridans 1.1.3.2 L-lactate oxidase synthesis enzymatic preparation of pyruvate by a whole-cell biocatalyst coexpressing L-lactate oxidase and catalase. Under the optimized transformation conditions, pyruvate is produced at a titer of 59.9 g/l and a yield of 90.8% in a substrate fed-batch process, promising an alternative route for the green production of pyruvate 1.1.3.4 glucose oxidase synthesis preparative production of hydroquinone using a column packed with the enzyme immobilized onto alumina, O2 is replaced with benzoquinone 1.1.3.4 glucose oxidase synthesis enzymatic biotransformation of (4R)-limonene to carvone involves addition of glucose oxidase and peroxidase to the biotransformation medium 1.1.3.4 glucose oxidase synthesis AldO is an enantioselective biocatalyst for the kinetic resolution of racemic 1,2-diols 1.1.3.4 glucose oxidase synthesis GOD is used as a commercial source of gluconic acid, which can be produced by the hydrolysis of delta-glucono-1, 5-lactone, the endproduct of GOD catalysis 1.1.3.4 glucose oxidase synthesis utilization of recombinant enzyme expressed in the periplasm or on the cell surface of Escherichia coli as biocatalyst in a non-laborious and non-costly whole-cell application for reacting on towards different polyols such as xylitol and sorbitol 1.1.3.4 glucose oxidase synthesis recombinant Aga2-GOx fusion proteins in the Saccharomyces cerevisiae cell wall can be used as immobilized catalysts for the production of gluconic acid 1.1.3.4 glucose oxidase synthesis the enzyme is used in the production of gluconic acid 1.1.3.4 glucose oxidase synthesis the addition of ferrous ions (Fe2+) induces the formation of hydroxyl radicals from the hydrogen peroxide, which act as initiating species for the microgel synthesis. Poly(N-vinyl)caprolactam (PVCL) microgels are synthesized by precipitation polymerization initiated by the enzyme and a conventional azo-initiator 2,2'-azobis(N-(2-carboxyethyl)-2-methylpropionamidine) tetrahydrate. The use of enzymes in precipitation polymerization leads to the encapsulation of enzymes in formed microgels. After the GOx-induced polymerization and purification of the microgels, high enzyme activity could be determined in the microgels, enabling facile synthesis of core-shell microgels. The glucose oxidase-based initiator system is a powerful and promising alternative to azo- or peroxide-initiated polymerization, leading to the formation of polymers at low synthesis temperatures 1.1.3.B5 eugenol oxidase synthesis fusion of eugenol oxidase and 5-hydroxymethylfurfural oxidase to be used for dioxygen-driven, one-pot, two-step cascade reactions to convert vanillyl alcohol into divanillin and eugenol into lignin oligomers 1.1.3.B5 eugenol oxidase synthesis synthesis of syringaresinol in a one-pot conversion containing eugenol oxidase (EUGO) and horseradish peroxidase (HRP) using 2,6-dimethoxy-4-allylphenol as a substrate. The hydrogen peroxide generated from the reaction of EUGO with the substrate is utilized by the HRP to convert the formed sinapyl alcohol into syringaresinol. Mutant I427A together with HRP are capable of efficiently producing syringaresinol as a major product. After optimization and upscaling to a semipreparative scale (1 gr), syringaresinol is obtained in 81% yield 1.1.3.6 cholesterol oxidase synthesis the enzyme is used for the production of steroid drugs and insecticides 1.1.3.7 aryl-alcohol oxidase synthesis due to hydride-transfer stereoselectivity and specificity on substituted aldehydes, the enzyme is useful for flavor production and in production of chiral compounds. Flavor synthesis and enzyme stereoselectivity, overview 1.1.3.7 aryl-alcohol oxidase synthesis AAO is able to produce 2,5-furandicarboxylic acid from formylfurancarboxylic acid, allowing full oxidation of 5-hydroxymethylfurfural. During 5-hydroxymethylfurfural reactions, an inhibitory effect of the H2O2 produced in the first two oxidation steps is the cause of the lack of AAO activity on formylfurancarboxylic acid. 5-Hydroxymethylfurfural is successfully converted into 2,5-furandicarboxylic acid when the AAO reaction is carried out in the presence of catalase 1.1.3.7 aryl-alcohol oxidase synthesis biooxidation of benzylic alcohols in the presence of various organic (co)solvents. The enzyme activity decreases at elevated concentrations of water-miscible polar solvents, while the presence of (halogenated) hydrocarbons is tolerated up to 90% (v/v), which leads to drastically improved conversions of up to >99% in case of hexafluorobenzene. This effect is correlated with the improved solubility of O2 in the employed solvents 1.1.3.7 aryl-alcohol oxidase synthesis construction of fusion proteins with Saccharomyces cerevisiae unspecific peroxidase, EC 1.11.2.1, using different peptide linkers. The reaction system is optimized to control the aromatic alcohol transformation rate, and therefore the H2O2 supply, to achieve total turnover numbers of 62000 for the biocatalytic synthesis of dextrorphan, a metabolite of the antitussive drug dextromethorphan 1.1.3.7 aryl-alcohol oxidase synthesis enantioselective oxidation of sec-allylic alcohols using variants of the berberine bridge enzyme analogue from Arabidopsis thaliana (AtBBE15) and the 5-(hydroxymethyl)furfural oxidase (HMFO) and its variants V465T, V465S, V465T/W466H and V367R/W466F. The enantioselectivity can be tuned by applying either pressure or by the addition of cosolvents 1.1.3.7 aryl-alcohol oxidase synthesis expression and secretion of AAO with yield of 315 mg/l using Pichia pastoris 1.1.3.7 aryl-alcohol oxidase synthesis expression variant carrying 4 mutations in the chimeric signal peptide (prealphaproK), plus mutations H91N/L170M in the mature protein, shows 350fold improved secretion (4.5 mg/l) in Saccharomyces cerevisiae and is stable. Expression in Pichia pastoris and fermentation in a fed-batch bioreactor enhances production to 25 mg/l. Both recombinant AAO from Saccharomyces cerevisiae and Pichia pastoris are subjected to the same N-terminal processing and have a similar pH activity profile, they differ in their kinetic parameters and thermostability 1.1.3.7 aryl-alcohol oxidase synthesis high-yield production of AAO in submerged culture using a recombinant Aspergillus nidulans strain grown on corn steep liquor. The optimum medium component concentrations are 61.0 g/l maltose, 26.4 g/L corn steep liquor, and 13.8 g/l NaNO3. The greatest AAO activity achieved is 1021 U/l with a protein concentration of 0.75 g/l 1.1.3.7 aryl-alcohol oxidase synthesis Mn(OAc)3 functions as a suitable activator for several commercially available variants of GOase with a series of alcohol substrates. Use of the Mn(OAc)3 additive is also compatible with biocatalytic synthesis of islatravir and subsequent biocatalytic steps in the islatravir-forming cascade 1.1.3.7 aryl-alcohol oxidase synthesis production of AAO using Aspergillus nidulans in a stirred-tank bioreactor. pH control significantly affects protein production and increasing dissolved oxygen level stimulates AAO production. The greatest AAO activity (1906 U/l) and protein concentration (1.19 g/l) are achieved in 48 h at 60% dissolved oxygen with pH controlled at 6.0. The yield and productivity (in 48 h) are 31.2 U/g maltose and 39.7 U/l/h, respectively 1.1.3.7 aryl-alcohol oxidase synthesis selective oxidation of (2E)-hex-2-en-1-ol to the corresponding aldehyde using recombinant aryl alcohol oxidase. The application of a two liquid phase system to overcome solubility and product inhibition issues yields more than 2200000 catalytic turnovers for the enzyme as well as molar product concentrations 1.1.3.9 galactose oxidase synthesis combination of UDP-Glc(NAc) 4'-epimerase and galactose oxidase in a one-pot synthesis of biotinylated nucleotide sugars. The enzymatic epimerization of uridine 5’-diphospho-alpha-D-glucose and uridine 5’-diphospho-N-acetyl-alpha-D-glucosamine and the subsequent oxidation of uridine 5’-diphospho-alpha-D-galactose and uridine 5’-diphospho-N-acetyl-alpha-D-galactosamine are combined with chemical biotinylation with biotin-epsilon-amidocaproylhydrazide in a one-pot synthesis. A mixture (1.0:1.4) of the biotinylated nucleotide sugars uridine 5’-diphospho-6-biotin-epsilon-amidocaproylhydrazino-alpha-D-galactose and uridine 5’-diphospho-6-biotin-epsilon-amidocaproylhydrazino-alpha-D-glucose, is produced in a reaction started with uridine 5’-diphospho-alpha-D-glucose. One product, uridine 5’-diphospho-6-biotin-epsilon-amidocaproylhydrazino-N-acetyl-alpha-D-galactosamine is formed when the reaction is initiated with uridine 5’-diphospho-N-acetyl-alpha-D-glucosamine 1.1.3.9 galactose oxidase synthesis galactose oxidase is used as a catalyst to oxidize selectively the C-6 hydroxyls of terminal galactose to carbonyl groups, e.g. polysaccharides studied included spruce galactoglucomannan, guar galactomannan, larch arabinogalactan, corn fiber arabinoxylan, and tamarind seed xyloglucan, with terminal galactose. A multienzyme system is used, with catalase and horseradish peroxidase to enhance the action of galactose oxidase, GC-MS analysis, overview 1.1.3.9 galactose oxidase synthesis galactose oxidase (GaO) selectively oxidizes the primary hydroxyl of galactose to a carbonyl, facilitating targeted chemical derivatization of galactose-containing polysaccharides, leading to renewable polymers with tailored physical and chemical properties. Increased substrate binding impeded the action of GaO fusions on more concentrated preparations of galactomannan, galactoglucomannan, and galactoxyloglucan 1.1.3.9 galactose oxidase synthesis galactose oxidase is a promising biocatalyst for the oxidation of primary and secondary alcohols to their corresponding aldehydes and ketones, respectively. For this purpose, GOase requires a number of additives to sustain its catalytic function, such as the enzyme catalase for degradation of the byproduct hydrogen peroxide as well as single-electron oxidants to reactivate the enzyme upon loss of the amino acid radical in its active site. GOase can be used to modify naturally occurring polysaccharides with terminal galactose moieties (or other saccharides through GOase mutants) by oxidizing the C6 hydroxyl groups and enabling further chemical or enzymatic modifications of the aldehyde such as amination. And GOase can be applied in the synthesis of a range of industrially relevant compounds containing ketones and aldehydes, such as diformylfuran obtained by selective oxidation of 5-hydroxymethylfurfural. GOase mutants able to enantioselectively oxidize secondary alcohols enable the use of the enzyme for kinetic resolution of racemic mixtures of secondary alcohols 1.1.3.9 galactose oxidase synthesis the enzyme can be used in the synthesis of small molecules, alcohols or amines, the production of H2O2 and reactive oxygen, and the production of O-glycosylated proteins 1.1.3.9 galactose oxidase synthesis Mn(OAc)3 functions as a suitable activator for several commercially available variants of GOase with a series of alcohol substrates. Use of the Mn(OAc)3 additive is also compatible with biocatalytic synthesis of islatravir and subsequent biocatalytic steps in the islatravir-forming cascade 1.1.3.10 pyranose oxidase synthesis oxidation of 2-keto-aldoses, formation of the methyl pentulose 1-deoxy-D-xylulose, whose phosphorylated form is the precursor of the vitamins thiamine and pyridoxol 1.1.3.10 pyranose oxidase synthesis industrial production of fructose 1.1.3.10 pyranose oxidase synthesis the enzyme produces the beta-pyrone antibiotic cortalcerone from D-glucose 1.1.3.10 pyranose oxidase synthesis hydrogenation of 2-keto-aldoses, preparation of D-tagatose, which is used as a building block for the synthesis of bioactive amino sugars and as low-calory sweetener 1.1.3.10 pyranose oxidase synthesis application of the engineered enzyme K312E in bioconversion of L-sorbose to 5-keto-D-fructose 1.1.3.10 pyranose oxidase synthesis enzyme P2O is a useful biocatalyst in several biotechnological applications, including biotransformation of carbohydrates such as D-glucose and D-galactose to generate 2-oxo-sugars that can be further reduced at the C1 position to yield D-fructose and D-tagatose, respectively 1.1.3.10 pyranose oxidase synthesis immobilization of enzyme to a glass-beaded support with activity yields of 10%-23.3%. after 4,800 measurement cycles carried out over 5 days, 58-62% of activity remains 1.1.3.12 pyridoxine 4-oxidase synthesis coexpression of enzyme with GroEL/Es genes in Escherichia coli under cold stress at 23°C results in 88fold higher specific activity than that of Microbacterium luteolum 1.1.3.12 pyridoxine 4-oxidase synthesis development of a simple and efficient synthesis for 4-pyridoxolactone starting with pyridoxine and using a whole-cell biotransformation by two transformed Escherichia coli cell type expressing the pyridoxine oxidase and the catalase, and chaperonin, while the second set expressed pyridoxal 4-dehydrogenase, overview. Pyridoxine is first oxidized to pyridoxal, which is then dehydrogenated to 4-pyridoxolactone by pyridoxine 4-oxidase and pyridoxal 4-dehydrogenase, respectively 1.1.3.12 pyridoxine 4-oxidase synthesis bioconversion of pyridoxine to pyridoxal. Approximately 450 mM pyridoxal is produced from 500 mM pyridoxine using recombinant Rhodococcus erythropolis expressing the pyridoxine 4-oxidase gene. Pyridoxal is converted to pyridoxamine by expression of the pyridoxamine-pyruvate aminotransferase gene derived from Rhizobium loti, and pyridoxine to pyridoxamine through pyridoxal using coexpression of the genes for pyridoxine 4-oxidase and pyridoxamine-pyruvate aminotransferase 1.1.3.13 alcohol oxidase synthesis enantioselective oxidation of thioanisole with a bienzymatic couple alcohol oxidase/peroxidase 1.1.3.13 alcohol oxidase synthesis expression of alcohol dehydrogenase with a thermostable NADPH-oxidase fusion partner (phenylacetone monooxygenase C65D). The resulting bifunctional biocatalysts retains the catalytic properties of the individual enzymes, and acts essentially like alcohol oxidases, while merely requiring a catalytic amount of NADP+ 1.1.3.13 alcohol oxidase synthesis the combination of alcohol oxidase and catalase is most effective in converting over 97% 5-hydroxymethylfurfural to 2,5-diformylfuran in 72 h 1.1.3.13 alcohol oxidase synthesis when covalently immobilized onto barium ferrite (BaFe12O19) magnetic microparticles, an immobilization efficiency of 71.0 % and catalytic activity of 34.6 U/g can be obtained. The immobilized OthAOX works optimally at 55°C and pH 8.0. More than 65% of the initial immobilized enzyme activity is retained after 24 h pre-incubation at 45°C. The immobilized enzyme shows a greater catalytic efficiency for oxidation of methanol and ethanol than free enzyme and can be recovered by magnetization and recycled for at least three consecutive batches, after which 70% activity remains 1.1.3.15 (S)-2-hydroxy-acid oxidase synthesis - 1.1.3.15 (S)-2-hydroxy-acid oxidase synthesis production of glyoxylate in biocatalysis of co-produced glycolate oxidase and catalase T 1.1.3.15 (S)-2-hydroxy-acid oxidase synthesis optimization of enzyme expression in Escherichia coli for production glyoxylate in culture medium 1.1.3.15 (S)-2-hydroxy-acid oxidase synthesis optimization of production of lactate oxidase by cultivating strains under high aeration in a medium with 0.5% glucose and 2% lactate for 1 day results in activities of 130-140 U/l 1.1.3.15 (S)-2-hydroxy-acid oxidase synthesis coexpression of N-demethylase NdmB gene from Pseudomonas putida CBB5 and glycolate oxidase gene in Escherichia coli. By two-step purification of Ni affinity chromatography and Q-Sepharose chromatography, the coexpressed NdmB and GO are separated and result in a 15.8fold purification with 8.7% yield and 12.8fold purification with 7.2% yield, respectively 1.1.3.17 choline oxidase synthesis a bienzymatic cascade for selective sulfoxidation uses an evolved recombinant peroxygenase from Agrocybe aegeritra which catalyzes the enantioselective sulfoxidation of thioanisole whereas the choline oxidase from Arthrobacter nicotianae provides the H2O2 necessary via reductive activation of ambient oxygen. The reactions are performed in choline chloride-based deep eutectic solvents serving as cosolvent and stoichiometric reductant at the same time. Product concentrations of up to 15 mM enantiopure sulfoxide and turnover numbers of 150,000 and 2100 for the peroxygenase and oxidase, respectively have been achieved 1.1.3.18 secondary-alcohol oxidase synthesis enzyme cartalyzes selective oxidation of several secondary (S)-alcohols, so the (R)-alcohols are obtained from racemic mixtures in high enantiopurity 1.1.3.21 glycerol-3-phosphate oxidase synthesis synthesis of dihydroxyacetone phosphate, enzyme coimmobilized with catalase on oxirane activated acrylic polymer beads, Eupergit C 250 1.1.3.38 vanillyl-alcohol oxidase synthesis bienzymatic route from capsaicin to vanillin for biocatalytic production of natural vanillin with carboxyesterase and vanillyl-alcohol oxidase 1.1.3.38 vanillyl-alcohol oxidase synthesis synthesis of natural vanillin from capsaicin using a two-enzyme system of carboxylesterase from liver and vanillyl-alcohol oxidase 1.1.3.38 vanillyl-alcohol oxidase synthesis fusion of eugenol oxidase and 5-hydroxymethylfurfural oxidase to be used for dioxygen-driven, one-pot, two-step cascade reactions to convert vanillyl alcohol into divanillin and eugenol into lignin oligomers. At pH 5.5, the fusdion protein converts 92% of vanillyl alcohol into vanillin (53%) and a higher amount of oligomers (39 %), of which the most dominant product is divanillin 1.1.3.41 alditol oxidase synthesis AldO is an enantioselective biocatalyst for the kinetic resolution of racemic 1,2-diols 1.1.3.41 alditol oxidase synthesis utilization of recombinant enzyme expressed in the periplasm or on the cell surface of Escherichia coli as biocatalyst in a non-laborious and non-costly whole-cell application for reacting on towards different polyols such as xylitol and sorbitol 1.1.3.41 alditol oxidase synthesis biosynthetic production of glycolate from glycerol using a variant of alditol oxidase, 2-hydroxyglutarate-pyruvate transhydrogenase from Saccharomyces cerevisiae, alpha-ketoisovalerate decarboxylase from Lactococcus lactis, and aldehyde dehydrogenase from Escherichia coli in an artificial operon expressed in Escherichia coli. To redirect glycerol flux toward glycolate synthesis, key genes of the native glycerol assimilation pathways are deleted and a second plasmid expressing Dld3 to reduce the accumulation of the intermediate D-glycerate is introduced. The final engineered strain produces 0.64 g/l glycolate in shake flasks, which is increased to 4.74 g/l in fed-batch fermentation 1.1.3.41 alditol oxidase synthesis one-pot biocatalytic system converting a range of triacylglycerols/natural oils into alpha-olefins. The system consists of CRL from Candida rugosa (for triacylglycerol hydrolysis to provide free fatty acids and glycerol), AldO (for in situ H2O2 generation upon glycerol oxidation), and OleTJE (for free fatty acid decarboxylation using H2O2 as cofactor) and is independent of exogenous addition of H2O2. The reaction system achieves a 68.5% total alkene yield from 500 microM coconut oil. About 0.5 g/l of alpha-olefins are produced from coconut oil (1500 microM) upon some reaction optimization 1.1.3.41 alditol oxidase synthesis production of ethylene glycol from glycerol by an artificial enzymatic cascade comprised of alditol oxidase, catalase, glyoxylate/hydroxypyruvate reductase, pyruvate decarboxylase and lactaldehyde:propanediol oxidoreductase. The NADH generated during the dehydrogenation of the glycerol oxidation product D-glycerate can be used as the reductant to support the ethylene glycol production. Using this in vitro synthetic system with self-sufficient NADH recycling, 7.64 mM ethylene glycol is produced from 10 mM glycerol in 10 h, with a yield of 0.515 g/g 1.1.3.41 alditol oxidase synthesis synthesis of rare ketoses from glycerol and D-/L-glyceraldehyde in a one-pot multienzyme fashion in which the only carbon source is glycerol. Glycerol is phosphorylated and then oxidized at C2 to afford dihydroxyacetone phosphate. The primary alcohol of glycerol is also oxidized to give the acceptor molecule glycerol aldehyde in situ (D- or L-isomer can be formed stereospecifically with either alditol oxidase or horse liver alcohol dehydrogenase). Different dihydroxyacetone phosphate-dependent aldolases are used to generate the aldol adducts (rare ketohexose phosphates) with various stereoconfigurations and diastereomeric ratios 1.1.3.46 4-hydroxymandelate oxidase synthesis whole-cell bioconversion of L-tyrosine into protocatechuic acid using artificial enzymatic cascades engineered in Escherichia coli. The first biocatalytic route comprises L-amino acid deaminase (LAAD) from Proteus mirabilis, hydroxymandelate synthase (HmaS) from Amycolatopsis orientalis, two-component flavin-dependent monooxygenase (HpaBC) from Escherichia coli, hydroxymandelate oxidase (HMO) from Streptomyces coelicolor, benzoylformate decarboxylase (BFD) from Pseudomonas putida, and aldehyde dehydrogenase (ALDH) from Saccharomyces cerevisiae. Combining LAAD/HmaS/HpaBC results in efficient synthesis of 3,4-dihydroxymandelate, which can be further converted to protocatechuic acid by HMO/BFD/ALDH to a final conversion of 64.4%. The recombinant E. coli catalyzes >99% conversion of L-tyrosine into 4-hydroxybenzoate within 12 h, further incorporation of 4-hydroxybenzoate hydroxylase PobA results in complete conversion of 4-hydroxybenzoate into protocatechuic acid, reaching >99% conversion 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis biocatalytic production of furan-2,5-dicarboxylate, a biobased platform chemical for the production of polymers 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis AAO is able to produce 2,5-furandicarboxylic acid from formylfurancarboxylic acid, allowing full oxidation of 5-hydroxymethylfurfural. During 5-hydroxymethylfurfural reactions, an inhibitory effect of the H2O2 produced in the first two oxidation steps is the cause of the lack of AAO activity on formylfurancarboxylic acid. 5-Hydroxymethylfurfural is successfully converted into 2,5-furandicarboxylic acid when the AAO reaction is carried out in the presence of catalase 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis biooxidation of benzylic alcohols in the presence of various organic (co)solvents. The enzyme activity decreases at elevated concentrations of water-miscible polar solvents, while the presence of (halogenated) hydrocarbons is tolerated up to 90% (v/v), which leads to drastically improved conversions of up to >99% in case of hexafluorobenzene. This effect is correlated with the improved solubility of O2 in the employed solvents 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis enantioselective oxidation of sec-allylic alcohols using variants of the berberine bridge enzyme analogue from Arabidopsis thaliana (AtBBE15) and the 5-(hydroxymethyl)furfural oxidase (HMFO) and its variants V465T, V465S, V465T/W466H and V367R/W466F. The enantioselectivity can be tuned by applying either pressure or by the addition of cosolvents 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis expression of HMFO in Pseudomonas putida S12 for the biocatalytic conversion of 5-hydroxymethylfurfural to FDCA. 35.7 mM 2,5-furandicarboxylic acid is produced from 50 mM 5-hydroxymethylfurfural in 24 h without notable inhibition. When the initial 5-ydroxymethylfurfural concentration is elevated to 100 mM, remarkable inhibition on 2,5-furandicarboxylic acid production is observed. Increasing the inoculum density solves the substrate inhibition. Using a fed-batch strategy, 545 mM of 2,5-furandicarboxylic acid can be accumulatively produced after 72 h 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis HMFO is used to convert 5-hydroxymethylfurfural to 2,5-diformylfuran and 5-formylfuroic acid (FFA), which is consecutively transformed to 2,5-furandicarboxylic acid by lipase Novozym 435. To facilitate the purification, a coupled alkali precipitation was developed to recover 2,5-furandicarboxylic acid from organic solvent with an improved purity from 84.4 to 99.0% and recovery of 78.1% 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis one-pot synthetic pathway to yield 2,5-furandicarboxylic acid from furfural. An oxidase and a prenylated flavin mononucleotide-dependent reversible decarboxylase, catalyze furfural oxidation and carboxylation of 2-furoic acid, respectively. The reversible decarboxylase is identified in Paraburkholderia fungorum KK1, whereas hydroxymethylfurfural oxidase from Methylovorus sp. MP688 exhibits furfural oxidation activity 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis oxidative kinetic resolution of racemic sec-thiols by enzyme variants, yielding the corresponding thioketones and nonreacted R-configured thiols with excellent enantioselectivities (E+200) 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis production of 2,5-furandicarboxylic acid by biotransformation of 5-hydroxymethylfurfural. Genes encoding 5-hydroxymethylfurfural oxidase and 5-hydroxymethylfurfural/furfural oxidoreductase from Cupriavidus basilensis HMF14 are introduced into Raoultella ornithinolytica BF60. The 2,5-furandicarboxylic acid production in the engineered whole-cell biocatalyst increases from 51.0 to 93.6 mM, and the molar conversion ratio of 5-hydroxymethylfurfural to 2,5-furandicarboxylic acid increases from 51.0 to 93.6% 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis synthetic pathway to yield 2,5-furandicarboxylic acid from furfural, produced from lignocellulosic biomass. The pathway consists of an oxidase and a prenylated flavin mononucleotide (prFMN)-dependent reversible decarboxylase, catalyzing furfural oxidation and carboxylation of 2-furoic acid, respectively. Upon coexpression in Escherichia coli, as well as a flavin prenyltransferase, 2,5-furandicarboxylic acid can be produced from furfural via 2-furoic acid in one pot 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis the combination of alcohol oxidase and catalase is most effective in converting over 97% 5-hydroxymethylfurfural to 2,5-diformylfuran in 72 h 1.1.3.47 5-(hydroxymethyl)furfural oxidase synthesis utilization of whole-cell Paraburkholderia azotifigens F18 for selective reduction of 5-hydroxymethylfurfural to 2,5-bis(hydroxymethyl)furan (BHMF) or oxidation to 5-hydroxymethyl-2-furancarboxylic acid (HMFCA). The whole-cell system can proceed an efficient hydrogenation reaction toward 5-hydroxymethylfurfural with a good selectivity of 97.6% to yield the BHMF at 92.2%. BHMF can be further oxidized to HMFCA and 2,5-furandicarboxylic acid (FDCA). The genes encoding HMF oxidoreductase/oxidase of whole-cell F18 are then deleted to prevent the further conversion of HMFCA to FDCA, which leads to a 10-fold decrease of FDCA. An 5-hydroxymethylfurfural conversion of 100% with an HMFCA yield of 98.3% is finally achieved, with an selectivity of 96.3% and a yield of 85.1% even at a high substrate concentration of up to 200 mM 1.1.5.14 fructose 5-dehydrogenase synthesis Gluconobacter frateurii CHM 43 have D-mannitol dehydrogenase (quinoprotein glycerol dehydrogenase) and flavoprotein D-fructose dehydrogenase in the membranes. When the two enzymes are functional, D-mannitol is converted to 5-keto-D-fructose with 65% yield when cultivated on D-mannitol. 5-Keto-D-fructose production with almost 100% yield is realized with the resting cells 1.1.99.6 D-lactate dehydrogenase (acceptor) synthesis production of pyruvate in an enzyme-membrane reactor 1.1.99.B9 (S)-2-hydroxycarboxylate dehydrogenase synthesis coupling of asymmetric oxidation of racemic 2-hydroxyacids with the opposite stereoselective reduction of 2-ketoacids for the stereoselective oxidoreductive deracemization to (R)-2-hydroxyacids, using a one-pot biocatalysis by resting cells of Saccharomyces cerevisiae strain ZJB5074 and Pseudomonas aeruginosa CCTCC M 2011394. Substituted racemic hydroxy(phenyl)acetic acids are converted to (R)-2-hydroxy-2-phenyl acetic acids with 55-98% conversion rate and 99.9% enantiomeric excess 1.1.99.B9 (S)-2-hydroxycarboxylate dehydrogenase synthesis enzyme can be used for the concurrent obtaining of aromatic (R)-2-hydroxyacids and aromatic 2-ketoacids by oxidation of aromatic 2-hydroxyacids in one-step biotransformation 1.1.99.18 cellobiose dehydrogenase (acceptor) synthesis the enzyme contains a protease-sensitive linker region, can be cleaved by endogenous proteases into a catalytically active flavin fragment and an inactive haem domain. Cleavage can be prevented by using increased concentrations of peptone or certain amino acids such as Val or Leu. These concentrations of up to 80 g peptone per l are not realistic for large scale-fermentations. Yet high levels of the enzyme produced under these conditions should stimulate both basic studies on the enzyme and studies on potential technological applications, e.g. for biosensors, bioremediation, or biocatalysis 1.1.99.21 D-sorbitol dehydrogenase (acceptor) synthesis production of 6-amino-L-sorbose, which is an intermediate in the miglitol production and an intermediate for oral alpha-glucosidase-inhibitors 1.1.99.21 D-sorbitol dehydrogenase (acceptor) synthesis the most prominent industrial method of producing L-sorbose is the biotransformation of D-sorbitol to L-sorbose in Gluconobacter species or Acetobacter species. L-sorbose is an important carbohydrate that is predominantly used as a starting material in the biosynthesis of L-ascorbic acid 1.1.99.21 D-sorbitol dehydrogenase (acceptor) synthesis biosynthesis of miglitol intermediate 6-(N-hydroxyethyl)-amino-6-deoxy-alpha-L-sorbofuranose by an improved D-sorbitol dehydrogenase from Gluconobacter oxydans. Miglitol (N-hydroxyethyl-1-deoxynojirimycin) is a pseudomonosaccharide alpha-glucosidase inhibitor in the treatment of non-insulin-dependent mellitus 1.1.99.28 glucose-fructose oxidoreductase synthesis glucose-fructose oxidoreductase and glucono-delta-lactonase are involved in bioconversion by cell of Zymomonas mobilis, production of sorbitol and gluconic acid. High initial glucose concentration leads to decreasing activities of in fresh cells due to changes in cell wall 1.1.99.28 glucose-fructose oxidoreductase synthesis the enzyme is used for large scale production of gluconic acid in a bioreactor system with immobilized enzyme, which is regenerated in situ, method development, overview 1.1.99.28 glucose-fructose oxidoreductase synthesis lactobionic acid production by a glucose-fructose oxidoreductase (GFOR)/glucono-delta-lactonase (GL) enzyme complex 1.1.99.29 pyranose dehydrogenase (acceptor) synthesis PDH catalysis with 1,4-benzoquinone as an oxidant provides biocatalytic sugar chemistry with a new convenient tool for high yield production of 3-keto-oligosaccharides and 3-keto-glycosides 1.1.99.29 pyranose dehydrogenase (acceptor) synthesis possible use in the preparation of rare di- and tricarbonyl sugar derivatives 1.1.99.29 pyranose dehydrogenase (acceptor) synthesis pyranose dehydrogenase is a promising candidate for the production of di- and tri-carbonyl sugar derivatives as chiral intermediates for the synthesis of rare sugars, novel drugs and fine chemicals 1.1.99.30 2-oxo-acid reductase synthesis enzyme can be used as a stereospecific biocatalyst for production of a wide range of 2-hydroxy-carboxylate compounds at preparative scale, with cofactor methyl or benzyl viologen being more stable than NAD(P)H 1.1.99.30 2-oxo-acid reductase synthesis enzyme can be used as a stereospecific biocatalyst for production of a wide range of compounds, e.g. 3-enoic-, 3,5-dienoic-, and 4-oxo-3R,S-hydroxyacids 1.1.99.30 2-oxo-acid reductase synthesis large scale microbial production of D-(S)-chloroacetic acid 1.1.99.30 2-oxo-acid reductase synthesis production of pyruvate from (R)-lactate in an enzyme-membrane reactor with coupled electrochemical regeneration of the artificial mediator anthraquinone-2,6-disulfonate 1.1.99.32 L-sorbose 1-dehydrogenase synthesis the enzyme (SDH) can be used to directly produce 2-keto-L-gulonic acid from L-sorbose with high substrate/product specificity. A platform strain suitable for highthroughput screening of SDH is constructed 1.1.99.35 soluble quinoprotein glucose dehydrogenase synthesis expression of enzyme in Pichia pastoris using the Saccharomyces cerevisiae alpha-factor signal sequence for secretion. The productivity of secreted PQQGDH-B achieves 218 kU/liter, i.e. 43 mg/liter. The secreted PQQGDH-B in Pichia pastoris is glycosylated but shows similar enzymatic properties as compared with those of recombinant PQQGDH-B produced in Escherichia coli 1.1.99.35 soluble quinoprotein glucose dehydrogenase synthesis production of recombinant soluble isoform by expression in Klebsiella pneumoniae at about 18 000 U per l, equal to that achieved in recombinant Escherichia coli. The signal sequence of recombinant PQQGDH-B produced by Klebsiella pneumoniae is correctly processed 1.1.99.36 alcohol dehydrogenase (nicotinoprotein) synthesis enzyme catalyzes the asymmetric reduction of ketones using cheap reductants, such as ethanol, with high stereoselectivity, but the reaction is too slow to obtain good yields. For developing biotransformations of industrial interest using nicotinoprotein alcohol dehydrogenases, the attention should be focused on enzymes with a higher reactivity towards prochiral ketones and secondary alcohols 1.2.1.3 aldehyde dehydrogenase (NAD+) synthesis synthesis of 3-hydroxypropionic acid. UTR engineering is used to maximally increase the activities of glycerol dehydratase and aldehyde dehydrogenase for the high conversion of glycerol to 3-hydroxypropionic acid. Thereafter, the activity of glycerol dehydratase is precisely controlled to avoid the accumulation of 3-hydroxypropionaldehyde by varying expression of dhaB1, a gene encoding a main subunit of glycerol dehydratase. The optimally balanced Escherichi coli HGL_DBK4 shows a substantially enhanced 3-hydroxypropionic acid titer and productivity compared with the parental strain. The yield on glycerol is 0.97 g 3-hydroxypropionic acid/g glycerol, in a fed-batch experiment 1.2.1.4 aldehyde dehydrogenase (NADP+) synthesis the aldehyde dehydrogenase from Thermoplasma acidophilum implemented as a key enzyme in a synthetic cellfree reaction cascade for the production of alcohols. Thermoplasma acidophilum enzyme TaALDH matches the cascade equirements regarding temperature stability, efficient heterologous expression in Escherichia coli, and exclusive activity for D-glyceraldehyde (not active on acetaldehyde). The remaining drawback is its cofactor preference toward NADP+ resulting in high KM for NAD+ and a rather low overall activity under cascade conditions 1.2.1.5 aldehyde dehydrogenase [NAD(P)+] synthesis titer and yield of 3-hydroxypropionic acid are improved to 54.7 mmol/l and 67% (mol/mol), respectively, when an ALDH gene (puuC) from Klebsiella pneumoniae is overexpressed in Pseudomonas denitrificans strain ATCC 13867 1.2.1.5 aldehyde dehydrogenase [NAD(P)+] synthesis the enzyme can be useful in the production of retinoic acid and related high-end products 1.2.1.10 acetaldehyde dehydrogenase (acetylating) synthesis 50 microg of alcohol dehydrogenase AdhA, EC 1.1.1.2, and 50 microg actaldehyde dehydrogenase AldH, EC 1.2.1.10, in buffer solution (pH 8.0) containing NADPH, NADH and acetyl-CoA at 60°C, produce 1.6 mM ethanol from 3 mM acetyl-CoA after 90 min 1.2.1.10 acetaldehyde dehydrogenase (acetylating) synthesis construction of a bypassed pyruvate decarboxylation pathway, through which pyruvate can be converted to acetyl-CoA, by using a coupled enzyme system consisting of pyruvate decarboxylase from Acetobacter pasteurianus and the CoA-acylating aldehyde dehydrogenase from Thermus thermophilus. A cofactor-balanced and CoA-recycling synthetic pathway for N-acetylglutamate production is designed by coupling the bypassed pathway with the glutamate dehydrogenase from Thermus thermophilus and N-acetylglutamate synthase from Thermotoga maritima. N-Acetylglutamate can be produced from an equimolar mixture of pyruvate and alpha-ketoglutarate with a molar yield of 55% through the synthetic pathway consisting of a mixture of four recombinant Escherichia coli strains having either one of the thermostable enzymes. The overall recycling number of CoA is 27 1.2.1.10 acetaldehyde dehydrogenase (acetylating) synthesis synthetic pathway for n-butanol production from acetyl coenzyme at 70°C, using beta-ketothiolase Thl, 3-hydroxybutyryl-CoA dehydrogenase Hbd, and 3-hydroxybutyryl-CoA dehydratase Crt from Caldanaerobacter subterraneus subsp. tengcongensis, trans-2-enoyl-CoA reductase Ter from Spirochaeta thermophila and bifunctional aldehyde dehydrogenase AdhE and and butanol dehydrogenase in vitro. n-Butanol is produced at 70°C, but with different amounts of ethanol as a coproduct, because of the broad substrate specificities of AdhE, Bad, and Bdh. A reaction kinetics model, validated via comparison to in vitro experiments, is used to determine relative enzyme ratios needed to maximize n-butanol production. By using large relative amounts of Thl and Hbd and small amounts of Bad and Bdh, >70% conversion to n-butanol is observed in vitro, but with a 60% decrease in the predicted pathway flux 1.2.1.10 acetaldehyde dehydrogenase (acetylating) synthesis synthetic pathway for n-butanol production from acetyl-CoA at 70°C, using beta-ketothiolase Thl, 3-hydroxybutyryl-CoA dehydrogenase Hbd, and 3-hydroxybutyryl-CoA dehydratase Crt from Caldanaerobacter subterraneus subsp. tengcongensis, trans-2-enoyl-CoA reductase Ter from Spirochaeta thermophila and bifunctional aldehyde dehydrogenase AdhE and and butanol dehydrogenase in vitro. n-Butanol is produced at 70°C, but with different amounts of ethanol as a coproduct, because of the broad substrate specificities of AdhE, Bad, and Bdh. A reaction kinetics model, validated via comparison to in vitro experiments, is used to determine relative enzyme ratios needed to maximize n-butanol production. By using large relative amounts of Thl and Hbd and small amounts of Bad and Bdh, >70% conversion to n-butanol is observed in vitro, but with a 60% decrease in the predicted pathway flux 1.2.1.11 aspartate-semialdehyde dehydrogenase synthesis the modofied enzyme with altered substrate specificity using NAD(H) is preferred in biotechnological production of amino acids due to lower costs and higher stability 1.2.1.12 glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) synthesis generation of a de novo NADPH generation pathway by altering the coenzyme specificity of native NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH) to NADP, to produce additional NADPH in the glycolytic pathway. Increasing the catalytic efficiency of GAPDH towards NADP enhances lysine production in all of the tested mutants, the most significant improvement of lysine production (60%) is achieved with the mutant showing similar preference towards both NAD and NADP. There is no significant change of flux towards the pentose phosphate pathway and the increased lysine yield is mainly attributed to the NADPH generated by the mutated GAPDH 1.2.1.12 glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) synthesis engineering the coenzyme specificity of (GAPDH) as a promising NADPH source is of interest for the metabolic engineering of NADPH-dependent bioproduction systems, e.g. for lysine production 1.2.1.19 aminobutyraldehyde dehydrogenase synthesis the enzyme is useful for GABA production in an engineered strain of Corynebacterium glutamicum produces 5.3 g/l of GABA 1.2.1.29 aryl-aldehyde dehydrogenase synthesis the enzyme can be used for production of bio-vanillin from vanillic acid 1.2.1.30 carboxylate reductase (NADP+) synthesis by combining the carboxylic acid reductase-dependent pathway with an exogenous fatty acid-generating lipase, natural oils (coconut oil, palm oil, and algal oil bodies) can be enzymatically converted into fatty alcohols across a broad chain length range (C8-C18) 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases (CARs) catalyze the conversion of carboxylic acids to aldehydes, which are a valuable class of chemicals for many consumer and industrial applications. CARs generally exhibit broad substrate specificity that encompasses aromatic, aliphatic, and di/tri-carboxylic acids, enabling the development of biosynthetic pathways to a wide array of potential aldehyde products. De novo biosynthesis utilizing CARs have produced industrially relevant products including aromatic aldehydes, fatty and aromatic alcohols, and alkanes. De novo synthetic pathways implementing CARs have enabled the production of sustainable aldehyde products or utilized highly reactive aldehydes as intermediates in the production of chemicals including amines, alcohols, and alkanes. Aromatic aldehydes, such as vanillin, benzaldehyde, and cinnamaldehyde are particularly valuable in the fragrance and flavoring industries and are produced from petroleum feedstocks in large quantities. Aldehydes as reactive intermediates in biosynthetic pathways, overview 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases (CARs) catalyze the conversion of carboxylic acids to aldehydes, which are a valuable class of chemicals for many consumer and industrial applications. CARs generally exhibit broad substrate specificity that encompasses aromatic, aliphatic, and di/tri-carboxylic acids, enabling the development of biosynthetic pathways to a wide array of potential aldehyde products. De novo biosynthesis utilizing CARs have produced industrially relevant products including aromatic aldehydes, fatty and aromatic alcohols, and alkanes. De novo synthetic pathways implementing CARs have enabled the production of sustainable aldehyde products or utilized highly reactive aldehydes as intermediates in the production of chemicals including amines, alcohols, and alkanes. Aromatic aldehydes, such as vanillin, benzaldehyde, and cinnamaldehyde are particularly valuable in the fragrance and flavoring industries and are produced from petroleum feedstocks in large quantities. Recombinant enzyme expression in Saccharomyces cerevisiae and Saccharomyces pombe and an engineered aldehyde-accumulating Escherichia coli strain for de novo production of vanillin from glucose. Aldehydes as reactive intermediates in biosynthetic pathways, overview 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases (CARs) catalyze the conversion of carboxylic acids to aldehydes, which are a valuable class of chemicals for many consumer and industrial applications. CARs generally exhibit broad substrate specificity that encompasses aromatic, aliphatic, and di/tri-carboxylic acids, enabling the development of biosynthetic pathways to a wide array of potential aldehyde products. De novo biosynthesis utilizing CARs have produced industrially relevant products including aromatic aldehydes, fatty and aromatic alcohols, and alkanes. De novo synthetic pathways implementing CARs have enabled the production of sustainable aldehyde products or utilized highly reactive aldehydes as intermediates in the production of chemicals including amines, alcohols, and alkanes. Aromatic aldehydes, such as vanillin, benzaldehyde, and cinnamaldehyde are particularly valuable in the fragrance and flavoring industries and are produced from petroleum feedstocks in large quantities. The CAR from Mycobacterium marinum (mmCAR) reduces a number of aliphatic acids ranging from C3 to C18, expanding the potential of CARs in synthetic pathways. Aldehydes as reactive intermediates in biosynthetic pathways, overview 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases are important enzymes in the toolbox for sustainable chemistry and provide specific use as biocatalysts 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases are important enzymes in the toolbox for sustainable chemistry and provide specific use as biocatalysts, application for the reduction of fatty acids to fatty alcohols 1.2.1.30 carboxylate reductase (NADP+) synthesis carboxylic acid reductases are important enzymes in the toolbox for sustainable chemistry and provide specific use as biocatalysts. The reduction of racemic ibuprofen by whole Nocardia iowensis cells gives an enantiomeric excess (ee) of 61.2%, which is attributed to enantioselectivity by niCAR based on kinetic data for its reduction of (S)-(+)- and (R)-(-)-ibuprofen enantiomers 1.2.1.30 carboxylate reductase (NADP+) synthesis the enzyme can be useful for aromatic aldehyde synthesis on industrial level. The product selectivity is an essential asset of the enzyme if it is used for the biocatalytic synthesis of organic molecules on the preparative level 1.2.1.36 retinal dehydrogenase synthesis the enzyme can be useful in the production of retinoic acid and related high-end products 1.2.1.44 cinnamoyl-CoA reductase synthesis potential exploitation of rationally engineered forms of CCR and CAD2 for the targeted modification of monolignol composition in transgenic plants 1.2.1.59 glyceraldehyde-3-phosphate dehydrogenase (NAD(P)+) (phosphorylating) synthesis engineering the coenzyme specificity of (GAPDH) as a promising NADPH source is of interest for the metabolic engineering of NADPH-dependent bioproduction systems, e.g. for lysine production 1.2.1.67 vanillin dehydrogenase synthesis isolate P2 can tolerate high concentrations of ferulic acid and vanillin, thus can be an excellent host for the high level production of vanillin and other phenolics by introducing metabolic pathway 1.2.1.67 vanillin dehydrogenase synthesis microbial vanillin production from ferulic acid precursor rice bran by employing vanillin-resistant Pediococcus acidilactici BD16, a natural lactic acid bacteria isolate. The vanillin-resistant organism can overcome the major drawback of microbial vanillin production from ferulic acid. which is the degradation and biotransformation of toxic vanillin to other phenolic derivatives. The strain resists more than 5 g/l vanillin in the medium 1.2.1.67 vanillin dehydrogenase synthesis the Gram-positive bacterium Amycolatopsis sp. strain ATCC 39116 is used for the fermentative production of natural vanillin from ferulic acid on an industrial scale. The strain is known for its outstanding tolerance to this toxic product vanillin 1.2.1.70 glutamyl-tRNA reductase synthesis the recombinant engineered GluTR variant R1 can be used for improvement of the C5 pathway to enhance 5-aminolevulinic acid and other products 1.2.1.75 malonyl-CoA reductase (malonate semialdehyde-forming) synthesis MCR is of biotechnological interest for the synthesis of polyester building blocks 1.2.1.75 malonyl-CoA reductase (malonate semialdehyde-forming) synthesis integration of multiple copies of malonyl-CoA reductase MCR and of phosphorylation-deficient acetyl-CoA carboxylase ACC1 genes into the genome of yeast increases 3-hydroxypropionic acid titer fivefold in comparison with single integration. Optimizing the supply of acetyl-CoA by overexpressing native pyruvate decarboxylase PDC1, aldehyde dehydrogenase ALD6, and acetyl-CoA synthase from Salmonella enterica SEacsL641P engineering the cofactor specificity of the glyceraldehyde-3-phosphate dehydrogenase to increase the intracellular production of NADPH at the expense of NADH improves 3-hydroxypropionic acid production and reduces formation of glycerol as by-product. The final strain produces 9.8 g per L 3-hydroxypropionic acid with a yield of 13% C-mol per C-mol glucose after 100 h in carbon-limited fed-batch cultivation at pH 5 1.2.1.75 malonyl-CoA reductase (malonate semialdehyde-forming) synthesis the combination of Escherichia coli BL21(DE3) and pET28a carrying heterogeneous acetyl-CoA carboxylase (Acc) from Corynebacterium glutamicum and codon-optimized malonyl-CoA reductase (MCR) from Chloroflexus aurantiacus is the most efficient host-vector system for 3-hydroxypropionic acid production, and the highest concentration of 3-hydroxypropionic attained in shake flask cultivation reaches 1.80 g/l with induction at 0.25 mM IPTG and 25°C, and supplementation of NaHCO3 and biotin 1.2.1.75 malonyl-CoA reductase (malonate semialdehyde-forming) synthesis developments in 3-hydroxypropionate (HP) production involving the enzyme using Saccharomyces cerevisiae as an industrial host. By combining genome-scale engineering tools, malonyl-CoA biosensors and optimization of downstream fermentation, the production of 3-HP in yeast has the potential to reach or even exceed the yield of chemical production 1.2.1.75 malonyl-CoA reductase (malonate semialdehyde-forming) synthesis the enzyme is very useful for 3-hydroxypropionate biosynthesis and production 1.2.1.89 D-glyceraldehyde dehydrogenase (NADP+) synthesis the enzyme accept NAD+ as cofactor under technically relevant conditions, making it suitable for application in the artificial glycolysis by synthetic cascade biomanufacturing 1.2.1.99 4-(gamma-glutamylamino)butanal dehydrogenase synthesis 3-hydroxypropionic acid is an important compound from which several commodity and specialty chemicals can be generated. The DELTAdhaT mutant (deletion in gene dhaT, encoding NADH-dependent 1,3-propanediol oxidoreductase) overexpressing PuuC has a potential to coproduce two commercially valuable products, 3-hydroxypropionic acid and 1,3-propanediol 1.2.1.99 4-(gamma-glutamylamino)butanal dehydrogenase synthesis 3-hydroxypropionic acid is an important compound from which several commodity and specialty chemicals can be generated. The recombinant strain SH254 producing 3-hydroxypropionate from glycerol is developed by cloning of dhaB and aldH genes in Escherichia coli BL21 under different regulatory promoters. The recombinant SH254 can accumulate 3-hydroxypropionic acid from glycerol at 6.5 mmol/l in 30 h 1.2.1.99 4-(gamma-glutamylamino)butanal dehydrogenase synthesis PuuC is an active enzyme that can be utilized effectively for several purposes, including biological production of 3-hydroxypropionate 1.2.1.99 4-(gamma-glutamylamino)butanal dehydrogenase synthesis 3-hydroxypropionic acid is an important platform chemical that can be used to synthesize a range of chemical compounds, such as acrylic acid, 1,3-propanediol, methyl acrylate, malonic acid, acrylamide, and hydroxyamides 1.2.1.104 pyruvate dehydrogenase system synthesis expression of pyruvate decarboxylase and alcohol dehydrogenase in Clostridium thermocellum DSM 1313. Though both enzymes are functional in Clostridium thermocellum, the presence of alcohol dehydrogenase severely limits the growth of the recombinant strains, irrespective of the presence or absence of the pyruvate decarboxylase gene. The recombinant strain expressing pyruvate decarboxylase shows two-fold increase in pyruvate carboxylase activity and ethanol production when compared with the wild type strain 1.2.1.104 pyruvate dehydrogenase system synthesis metabolic engineering of Geobacillus thermoglucosidasius to divert the fermentative carbon flux from a mixed acid pathway to one in which ethanol becomes the major product, involving elimination of the lactate dehydrogenase and pyruvate formate lyase pathways by disruption of the ldh and pflB genes, respectively, and upregulation of expression of pyruvate dehydrogenase. Pyruvate dehydrogenase is active under anaerobic conditions, but expressed suboptimally for a role as the primary fermentation pathway. Strains with all three modifications form ethanol efficiently and rapidly at temperatures in excess of 60°C in yields in excess of 90% of theoretical. The strains also efficiently ferment cellobiose and a mixed hexose and pentose feed 1.2.1.104 pyruvate dehydrogenase system synthesis coexpression of pyruvate dehydrogenase (PDH) E1 alpha and E1 beta subunits in Escherichia coli leads to fully active E1 protein. The production of E1 alpha alone results in a catalytically inactive protein. The PDH E1 protein produced in Escherichia coli is capable of being phosphorylated by PDH-specific kinase 1.2.1.104 pyruvate dehydrogenase system synthesis exchange of the native Corynebacterium glutamicum promoter of the AceE gene, with mutated DapA promoter variants leads to a series of strains with gradually reduced growth rates and pyruvate dehydrogenase complex activities. Upon overexpression of the L-valine biosynthetic genes IlvBNCE, all strains produce L-valine. Additional deletion of the Pqo and Ppc genes, encoding pyruvate:quinone oxidoreductase and phosphoenolpyruvate carboxylase enables production of up to 738mM (i.e., 86.5 g/liter). Inactivation of the transaminase B gene (IlvE) and overexpression of IlvBNCD instead of ilvBNCE transform the L-valine-producing strain into a 2-ketoisovalerate producer, excreting up to 303mM (35 g/liter) 2-ketoisovalerate. The replacement of the AceE promoter by the DapA-A16 promoter improves the production by 100% and 44%, respectively 1.2.1.104 pyruvate dehydrogenase system synthesis production of the functional pyruvate dehydrogenase complex. All components are coexpressed in the cytoplasm of baculovirus-infected SF9 cells by deletion of the mitochondrial localization signal sequences and E1a is FLAG-tagged to facilitate purification. The protein complex is purified using FLAG-M2 affinity resin, followed by Superdex 200 sizing chromatography. The E2 and E3BP components are then lipoylated in vitro. The resulting complex is over 90% pure and homogenous 1.2.1.105 2-oxoglutarate dehydrogenase system synthesis expression of the structural genes for all three subunits of the Escherichia coli 2-oxoglutarate dehydrogenase complex in Saccharomyces cerevisiae. The Escherichia coli lipoic-acid scavenging enzyme is coexpressed to enable cytosolic lipoylation of the complex, which is required for its enzymatic activity. In vivo cytosolic activity of the complex is tested byconstructing a reporter strain in which the essential metabolite 5-aminolevulinic acid can only be synthetized from cytosolic, and not mitochondrial, succinyl-CoA. In the resulting strain, complementation of 5-aminolevulinic acid auxotrophy depends on activation of the 2-oxoglutarate dehydrogenase complex by lipoic acid addition 1.2.3.1 aldehyde oxidase synthesis in shake-flask cultures, recombinant Klebsiella pneumoniae expressing the enzyme produces 0.89 g 3-hydroxypropionate per l, i.e. 3 g 3-hydroxypropionate per l during 24 h fed-batch cultivation in a 5 l bioreactor 1.2.3.3 pyruvate oxidase synthesis constitutive overproduction of enzyme, acetate becomes the only fermentation end product 1.2.3.3 pyruvate oxidase synthesis effect of acettae pathway mutations on production of pyruvate, pH value of 7.0 and 32°C favor greatest pyruvate generation 1.2.3.3 pyruvate oxidase synthesis high-level expression of enzyme is achieved in recombinant Escherichia coli by optimizing the expression system and induction conditions. Optimization leads to the highest pyruvate oxidase yield (4106.9 U/l) under conditions of 25°C, 0.5 mM IPTG, 0.5 OD600, after 24 h of induction 1.2.3.3 pyruvate oxidase synthesis pH stress during recombinant Escherichia coli cultivation leads to great losses of enzyme. Residual POD activity (3.1%) is increased to 43.3, 88.0 and 61.5% by maintaining the pH at 5.0, 6.0 and 7.0 respectively during the induction phase, which decreases inclusion body formation and increases production of soluble POD. Glycerol addition increases the cell density and volumetric POD activity to 21243.3 U/l at pH 6.0 after 64 h induction 1.2.3.3 pyruvate oxidase synthesis POD expression in Escherichia coli is achieved by coexpression of chaperone SecB under three promoters (T7, lac, bla). The weakest promoter, bla, when compared with T7 and lac promoters, provides the optimum extracellular POD activity among these three. After optimization of cultivation conditions, the extracellular POD yield increases to 795.7 U/l. By using glycine to disrupt recombinant Escherichia coli cell wall and Cu2+ ions as POD stabilizer, the final extracellular POD yield reaches 2926.3 U/l 1.2.3.3 pyruvate oxidase synthesis production of enzyme in recombinant Escherichia coli constitutively expressing paruvate oxidase. A high-cell density fed-batch cultivation with gradient temperature decrease strategy is achieved, under which the biomass (OD600) of recombinant E. coli can reach to 71 and the highest PyOD activity in broth can reach to 3307 U/l in 26 h fermentation 1.2.3.4 oxalate oxidase synthesis optimum culture conditions for production of oxalate oxidase are sucrose at 20 g/l, NH4Cl at 2 g/l, MnSO4 at 0.05 g/l and sodium oxalate as inducer at 5 g/l, fill up volume in shake flask at one tenth, incubation temperature of 30°C and initial medium pH of 6.5 and inoculum age of 18 h 1.2.3.15 (methyl)glyoxal oxidase synthesis efficient expression in Pichia pastoris and Trichoderma reesei with subsequent purification by anion exchange and hydrophobic interaction chromatography. Both processes are suitable for the production of the target protein at high levels. GLOX produced in T. reesei carries mainly Man5 glycosylation while the enzyme produced in P. pastoris exhibits the typical high-mannose type N-glycosylation. The enzyme expressed in P. pastoris shows slightly higher specific activities which correlates with the higher copper loading of 65.5 % compared to 51.9 % for the protein from T. reesei 1.2.4.1 pyruvate dehydrogenase (acetyl-transferring) synthesis engineering of the wild type of Corynebacterium glutamicum for the growth-decoupled production of 2-ketoisovalerate from glucose by deletion of the aceE gene encoding the E1p subunit of the pyruvate dehydrogenase complex, deletion of the transaminase B gene ilvE, and additional overexpression of the ilvBNCD genes, encoding the L-valine biosynthetic enzymes acetohydroxyacid synthase (AHAS), acetohydroxyacid isomeroreductase, and dihydroxyacid dehydratase. 2-Ketoisovalerate production is further improved by deletion of the pyruvate:quinone oxidoreductase gene pqo. In fed-batch fermentations at high cell densities, the newly constructed strains produce up to 188 mM (21.8 g/liter) 2-ketoisovalerate and showd a product yield of about 0.47 mol per mol (0.3 g/g) of glucose and a volumetric productivity of about 4.6 mM (0.53 g/liter) 2-ketoisovalerate per h in the overall production phase 1.2.4.1 pyruvate dehydrogenase (acetyl-transferring) synthesis coexpression of pyruvate dehydrogenase (PDH) E1 alpha and E1 beta subunitsin Escherichia coli leads to fully active E1 protein. The production of E1 alpha alone results in a catalytically inactive protein. The PDH E1 protein produced in Escherichia coli is capable of being phosphorylated by PDH-specific kinase 1.2.4.1 pyruvate dehydrogenase (acetyl-transferring) synthesis production of pyruvate in an Escherichia coli strain with central metabolic pathways modified. Genes ldhA, pflB, pta-ackA, poxB, ppc, frdBC are knocked out sequentially and full pyruvate dehydrogenase is retained. In batch fermentation with M9 medium, pyruvate yield and production rate reach 0.63 g/g glucose and 1.89 g/(1 h), respectively. The production of acetate, succinate, and other carboxylates is effectively controlled, while the pathways of convertion of pyruvate to phosphoenol pyruvate and acetyl CoA are enhanced 1.2.4.1 pyruvate dehydrogenase (acetyl-transferring) synthesis when recombinant subunits PDHA and PDHB are mixed at a 1:1 molar ratio, they exhibit strong enzymatic activity. Alone, recombinant PDHA and PDHB exhibit no or weak enzymatic activity 1.2.4.2 oxoglutarate dehydrogenase (succinyl-transferring) synthesis expression of the structural genes for all three subunits of the Escherichia coli 2-oxoglutarate dehydrogenase complex in Saccharomyces cerevisiae. The Escherichia coli lipoic-acid scavenging enzyme is coexpressed to enable cytosolic lipoylation of the complex, which is required for its enzymatic activity. In vivo cytosolic activity of the complex is tested by constructing a reporter strain in which the essential metabolite 5-aminolevulinic acid can only be synthetized from cytosolic, and not mitochondrial, succinyl-CoA. In the resulting strain, complementation of 5-aminolevulinic acid auxotrophy depends on activation of the 2-oxoglutarate dehydrogenase complex by lipoic acid addition 1.2.4.4 3-methyl-2-oxobutanoate dehydrogenase (2-methylpropanoyl-transferring) synthesis functional expression of the branched chain alpha-keto acid dehydrogenase complex in Escherichia coli by independent and selective optimization of individual subunit genes of the complex. Codon optimization significantly improves the expression of complex component proteins BkdH and LpdA1 but expression of dehydrogenase E1 alpha subunit BkdF and dehydrogenase E1 beta subunit BkdG depends on coexpression of the BkdH gene. The optimized branched chain alpha-keto acid dehydrogenase complex supplies sufficient short branched-chain acyl-CoA to synthesize phlorisovalerophenone 1.2.5.2 aldehyde dehydrogenase (quinone) synthesis application in direct bioelectrocatalysis via site specific immobilization to form a monolayer of biocatalysts with a uniform orientation toward the gold electrode. Six-histidine tags at the N- or C-terminus of each of the three subunits are utilized as linking sites to performsite specific immobilization. The orientation of multi-subunit enzymes can affect direct electron transfer greatly by varying the electron tunneling distances. The favorable orientation allowing for a minimal heme c electron transfer distance shows a direct electron transfer rate 6.6fold higher than that with the orientation closest to the active site of the enzyme 1.2.5.2 aldehyde dehydrogenase (quinone) synthesis bioelectrooxidation of ethanol using pyrroloquinoline quinone-dependent alcohol and aldehyde dehydrogenase enzymes for biofuel cell applications. A modified Nafion membrane provides the best electrical communication between enzymes and the electrode surface,with catalytic currents as high as 16.8 mA/cm2. Direct electron transfer using the pyrroloquinoline quinone-dependent alcohol and aldehyde dehydrogenase still lacks high current density, while the bioanodes that operate via mediated electron transfer employing ferrocene-modified linear polyethyleneimine redox polymers show efficient energy conversion capability in ethanol/air biofuelcells 1.2.5.3 aerobic carbon monoxide dehydrogenase synthesis establishment of a functional heterologous expression system of Moco-free apo-CODH in Escherichia coli. The expression of the CoxMSL genes alone results in a colorless and inactive protein that lacks the cofactors. Expression of an active protein requires coexpression of CoxI 1.2.7.1 pyruvate synthase synthesis engineering of Thermoanaerobacterium saccharolyticum to produce ethanol at high yield. Knockout of genes involved in organic acid formation (acetate kinase, phosphate acetyltransferase, and L-lactate dehydrogenase) results in a strain able to produce ethanol as the only detectable organic product and substantial changes in electron flow relative to the wild type. Ethanol formation in the engineered strain ALK2 utilizes pyruvate:ferredoxin oxidoreductase with electrons transferred from ferredoxin to NAD(P). The homoethanologenic phenotype is stable for more than 150 generations in continuous culture. The growth rate of strain ALK2 is similar to the wild-type strain, with a reduction in cell yield proportional to the decreased ATP availability resulting from acetate kinase inactivation. Glucose and xylose are coutilized and utilization of mannose and arabinose commences before glucose and xylose are exhausted. Using strain ALK2 in simultaneous hydrolysis and fermentation experiments at 50°C allows a 2.5fold reduction in cellulase loading compared with using Saccharomyces cerevisiae at 37°C. The maximum ethanol titer produced by strain ALK2 is 37 g/liter 1.2.7.1 pyruvate synthase synthesis homologous protein production in Thermoacetobacter kivui by cloning and expression in Thermoacetobacter kivui. The encoded protein containing a genetically engineered His-tag is purified in only two steps to apparent homogeneity, and has the same properties as the native 1.2.7.7 3-methyl-2-oxobutanoate dehydrogenase (ferredoxin) synthesis engineering of Clostridium thermocellum to produce isobutanol. Both the native 2-oxoisovalerate-oxidoreductase KOR, EC 1.2.7.7, and the heterologous Lactococcus lactis 2-oxoisovalerate decarboxylase KIVD, EC 4.1.1.74, expressed are responsible for isobutanol production.The plasmid is integrated into the chromosome by single crossover. The resulting strain is stable without antibiotic selection pressure and produces 5.4g/l of isobutanol from cellulose in minimal medium at 50°C within 75 h, corresponding to 41% of theoretical yield 1.2.7.7 3-methyl-2-oxobutanoate dehydrogenase (ferredoxin) synthesis expression of a isobutanol synthesis pathway using ketoisovalerate ferredoxin oxidoreductase (Kor) from Clostridium thermocellum and bifunctional aldehyde/alcohol dehydrogenase (AdhE2) from Clostidium acetobutylicum. Only in recombinant Acetobacterium woodii strains, traces of isobutanol can be detected. Additional feeding of ketoisovalerate increases isobutanol production to 2.9 mM under heterotrophic conditions using isoform Kor3 and to 1.8 mM under autotrophic conditions using isoform Kor2. In Clostidium ljungdahlii, isobutanol can only be detected upon additional ketoisovalerate feeding under autotrophic conditions. Isoform Kor3 is the best suited gene cluster 1.2.98.1 formaldehyde dismutase synthesis formate production with immobilized enzyme 1.2.98.1 formaldehyde dismutase synthesis production of methanol and formic acid using a chemo-/biocatalytic oxidation cascade. Starting from synthetic biogas, methane is oxidized to formaldehyde over a mesoporous VOx/SBA-15 catalyst. Formaldehyde is disproportionated enzymatically towards methanol and formic acid in equimolar ratio by formaldehyde dismutase. The cascades generate methanol in much higher productivity compared to methane monooxygenase 1.3.1.8 acyl-CoA dehydrogenase (NADP+) synthesis oil-based extended fermentation of recombinant Streptomyces cinnamonensis, expressing the enzyme from Streptomyces collinus, is used to provide methylmalonyl-CoA precursors for monensin biosynthesis 1.3.1.19 cis-1,2-dihydrobenzene-1,2-diol dehydrogenase synthesis preparation of two recombinant strains each containing two enzymatic activities mutually expressed through regulated systems for production of functionalized epoxides in one-pot reactions. The gene coding for styrene monooxygenase from Pseudomonas fluorescens ST and the gene coding for naphthalene dihydrodiol dehydrogenase from Pseudomonas fluorescens N3 are expressed either in Pseudomonas putida PaW340 or in Escherichia coli. The combination of the enzymes allows to obtain the transformation of allylic primary alcohols into alpha,beta-epoxy acids 1.3.1.31 2-enoate reductase synthesis 2-methylsuccinic acid is a C5 branched-chain dicarboxylate that serves as an attractive synthon for the synthesis of polymers with extensive applications in coatings, cosmetic solvents and bioplastics. A promising platform for 2-methylsuccinic acid bioproduction is established. Over-expression of codon-optimized citramalate synthase variant CimA from Methanococcus jannaschii, endogenous isopropylmalate isomerase EcLeuCD and enoate reductase YqjM from Bacillus subtilis allows the production of 2-methylsuccinic acid in Escherichia coli. Incorporation of the enzyme (KpnER) into the 2-methylsuccinic acid biosynthetic pathway leads to 2-methylsuccinic acid production improvement to a titer of 0.96 g/l in aerobic condition 1.3.1.31 2-enoate reductase synthesis Bacillus sp. FM18civ1 can be useful for resolving a racemic mixture of carvone. Biotransformation of (4R)-(-)-carvone with Mentha pulegium leaves 1.3.1.31 2-enoate reductase synthesis hydrogenation activity of enoate reductases can potentially replace the chemical hydrogenation step in current synthetic protocols creating a completely bio-based pathway for the greener production of adipic acid or other biochemicals e.g. 3-phenylpropanoic acid from renewable feedstocks 1.3.1.31 2-enoate reductase synthesis hydrogenation activity of enoate reductases can potentially replace the chemical hydrogenation step in current synthetic protocols creating a completely bio-based pathway for the greener production of adipic acid or other biochemicals e.g. 3-phenylpropanoic acid from renewable feedstocks. High oxygen tolerance and thermostability make the enzyme useful for in vivo and in vitro applications to overcome the limitations of chemical catalysts 1.3.1.84 acrylyl-CoA reductase (NADPH) synthesis successful reconstitution a portion of the 3-hydroxypropionate/4-hydroxybutyrate cycle from Metallosphaera sedula in Escherichia coli to produce acrylic acid and propionic acid, involving the acryloyl-CoA reductase (ACR), overview 1.3.1.92 artemisinic aldehyde DELTA11(13)-reductase synthesis production of artemisin by incubating a mixture of artemisin precursors from engineered Saccharomyces cerevisiae with a cell-free extract of Artemisia annua. Use of cold-acclimated Artemisia annua cell-free extract gives rise to considerable artemisin content up to 0.65% 1.3.1.92 artemisinic aldehyde DELTA11(13)-reductase synthesis for enhanced artemisin production, Appropriate doses of Cd can increase the concentrations of artemisinic metabolites at a certain time point by upregulating the relative expression levels of key enzyme genes involved in artemisinin biosynthesis 1.3.1.97 botryococcene synthase synthesis an expression system for production of high levels of botryococcene in Nicotiana tabacum plants can be used involving the recombinant chimeric enzyme, trichome-specific expression of botryococcene metabolism, overview 1.3.1.103 2-haloacrylate reductase synthesis construction of a system for asymmetric reduction of 2-chloroacrylate to produce (S)-2-chloropropionate with recombinant Escherichia coli cells producing 2-haloacrylate reductase from Burkholderia sp. WS and an NADPH regeneration system. The system provided 37.4 g/l (S)-2-chloropropionate in more than 99.9%e.e. (S)-2-Chloropropionate is a synthetic intermediate for phenoxypropionic acid herbicides 1.3.1.103 2-haloacrylate reductase synthesis the enzyme is useful for the production of (S)-2-chloropropionate, which is used for the industrial production of aryloxyphenoxypropionic acid herbicides 1.3.1.103 2-haloacrylate reductase synthesis the enzyme is useful in the production of L-2-chloropropionate, a building block in the synthesis of aryloxyphenoxypropionate herbicides 1.3.1.109 butanoyl-CoA dehydrogenase complex (NAD+, ferredoxin) synthesis engineering of Clostridum sp. MT1962 by elimination of phosphotransacetylase and acetaldehyde dehydrogenase along with integration to chromosome synthetic thiolase, 3-hydroxybutyryl-CoA dehydrogenase, crotonase, butyryl-CoA dehydrogenase, butyraldehyde dehydrogenase, and NAD-dependent butanol dehydrogenase. Th resulting strain loses production of ethanol and acetate while initiated the production of 297 mM of n-butanol 1.3.3.5 bilirubin oxidase synthesis enzymatic oxidative polymerization of dihydroquercetin from Larix sibirica using bilirubin oxidase as a biocatalyst. As compared with the monomer, oligoDHQ demonstrates higher thermal stability and high antioxidant activity 1.3.3.5 bilirubin oxidase synthesis mediator-less, direct electro-catalytic reduction of oxygen to water is achieved on spectrographite electrodes modified by physical adsorption of bilirubin oxidases from Myrothecium verrucaria. Bilirubin oxidase (BOD) is the best catalyst for converting oxygen directly to water because of the very low overpotential necessary to catalyze the reaction 1.3.3.11 pyrroloquinoline-quinone synthase synthesis cofactor engineering of PQQ in Gluconobacter oxydans is beneficial for enhancing the production of quinoprotein-related products 1.3.5.2 dihydroorotate dehydrogenase (quinone) synthesis expression of full-length and N-terminally trucated enzyme lacking 29 amino acids in an Escherichia coli system 1.3.5.5 15-cis-phytoene desaturase synthesis the norflurazon-resistant phytoene desaturase gene is used for successful nuclear transformation and genetic engineering of the carotenoid biosynthesis pathway for accelerated astaxanthin biosynthesis (astaxanthin as a food supplement for humans) 1.3.5.5 15-cis-phytoene desaturase synthesis Dunalielle salina can be used as a cell factory for phytoene production 1.3.7.4 phytochromobilin:ferredoxin oxidoreductase synthesis production of full-length plant phytochrome assembled with phytochromobilin in Pichia pastoris by coexpressing apophytochromes and chromophore biosynthetic genes, heme oxygenase (HY1) and phytochromobilin oxidoreductase (HY2) from Arabidopsis thaliana. Mitochondria localization of the phytochromobilin biosynthetic genes increases the efficiency of holophytochrome biosynthesis 1.3.8.1 short-chain acyl-CoA dehydrogenase synthesis expression of acetyl-coenzyme A C-acetyltransferase, 3-hydroxybutyryl-CoA dehydrogenase, crotonase, phosphate butyryltransferase, and butyrate kinase and the butyryl-CoA dehydrogenase complex composed of the dehydrogenase and two electron-transferring flavoprotein subunits as a single plasmid-encoded operon in Escherichia coli to confer butyrate-forming capability 1.3.8.1 short-chain acyl-CoA dehydrogenase synthesis engineering of Clostridum sp. MT1962 by elimination of phosphotransacetylase and acetaldehyde dehydrogenase along with integration to chromosome synthetic thiolase, 3-hydroxybutyryl-CoA dehydrogenase, crotonase, butyryl-CoA dehydrogenase, butyraldehyde dehydrogenase, and NAD-dependent butanol dehydrogenase. Th resulting strain loses production of ethanol and acetate while initiated the production of 297 mM of n-butanol 1.3.99.4 3-oxosteroid 1-dehydrogenase synthesis engineered enzyme mutants, in which KsdDM is inactivated or augmented, are useful for production of phytosterols 4-androstene-3,17-dione or 1,4-androstadiene-3,17-dione, circumventing the difficulty of separating 4-androstene-3,17-dione from 1,4-androstadiene-3,17-dione, a key bottleneck to the microbial transformation of phytosterols in industry 1.3.99.4 3-oxosteroid 1-dehydrogenase synthesis kstDF is a promising enzyme in steroid DELTA1-dehydrogenation and steroid reduction that is propitious to construct genetically engineered steroid-transforming recombinants by heterologous overexpression 1.3.99.4 3-oxosteroid 1-dehydrogenase synthesis inactivation of KstD isoforms 1-3 leads to synthesis of about 6.02g/l of 9alpha-hydroxy-4-androstene-3,17-dione from 15 g/l of phytosterols. The product is mixed with 1.55 g/l of 4-androstene-3,17-dione as a major byproduct. Overexpression of the oxygenase component of 3-ketosteroid-9alpha-hydroxylase kshA, results in a yield of 9alpha-hydroxy-4-androstene-3,17-dione of 7.33 g/l 1.3.99.4 3-oxosteroid 1-dehydrogenase synthesis recombinant enzyme application potential of the former in the synthesis of prednisolone, method evaluation and optimization 1.3.99.17 quinoline 2-oxidoreductase synthesis production of uracil in supernatant of resting cells with a yield of more than 98% 1.3.99.27 1-hydroxycarotenoid 3,4-desaturase synthesis heterologous production of two unusual acyclic carotenoids, 1,1'-dihydroxy-3,4-didehydrolycopene and 1-hydroxy-3,4,3',4'-tetradehydrolycopene by combination of the crtC and crtD genes from Rhodobacter and Rubrivivax 1.3.99.28 phytoene desaturase (neurosporene-forming) synthesis swapping the native three-step CrtI for the four-step Pantoea agglomerans enzyme, EC 1.3.99.31, reroutes carotenoid biosynthesis and culminates in the production of 2,2'-diketo-spirilloxanthin under semi-aerobic conditions. Premature termination of this pathway by inactivating crtC or crtD produces strains with lycopene or rhodopin as major carotenoids. All of the spirilloxanthin series carotenoids are accepted by the assembly pathways for light-harvesting 2 complex and reaction centre-light-harvesting 1-PufX complexes 1.3.99.31 phytoene desaturase (lycopene-forming) synthesis swapping the native three-step CrtI in Rhodobacter sphaeroides, EC 1.3.99.28, for the four-step Pantoea agglomerans enzyme reroutes carotenoid biosynthesis and culminates in the production of 2,2'-diketo-spirilloxanthin under semi-aerobic conditions. Premature termination of this pathway by inactivating crtC or crtD produces strains with lycopene or rhodopin as major carotenoids. All of the spirilloxanthin series carotenoids are accepted by the assembly pathways for light-harvesting 2 complex and reaction centre-light-harvesting 1-PufX complexes 1.3.99.39 carotenoid phi-ring synthase synthesis expression of CrtU in the beta-carotene ketolase (crtO) knockout Rhodococcus erythropolis host produces higher purity chlorobactene than expression in the wild-type Rhodococcus host. Yeast extract medium supplemented with fructose gives the highest total yield of chlorobactene. In a 10-l fermentor, about 18 mg of chlorobactene is produced 1.4.1.1 alanine dehydrogenase synthesis batch synthesis of L-Ala at room temperature via reductive amination of pyruvate 1.4.1.1 alanine dehydrogenase synthesis continuous production of 3-fluoro-L-alanine with alanine dehydrogenase 1.4.1.1 alanine dehydrogenase synthesis alterating of cofactor specificity from NADH to NADPH via protein engineering for application as a regenerating enzyme in coupled reactions with NADPH-dependent alcohol dehydrogenases. Mutant D196A/L197R can be applied in a coupled oxidation/transamination reaction of the dicyclic dialcohol isosorbide to its diamines, catalyzed by Ralstonia sp. alcohol dehydrogenase and Paracoccus denitrificans omega-aminotransferase, allowing recycling of NADP+ and L-Ala. Recycling factors of 33 for NADP+ and 13 for L-Ala are observed 1.4.1.1 alanine dehydrogenase synthesis the enzyme can be used as heterogeneous biocatalyst for reductive aminations of 2-oxoacids 1.4.1.1 alanine dehydrogenase synthesis the enzyme from Amycolatopsis sulphurea has the ability to produce valuable molecules for various industrial purposes and represents a potential biocatalyst for biotechnological applications 1.4.1.2 glutamate dehydrogenase synthesis produktion of L-ornithine in Corynebacterium glutamicum SNK118, by deletion of genes argF, argR, and ncgl2228 to block the degradation of L-ornithine, and overexpression of NADP-dependent glyceraldehyde 3-phosphate dehydrogenases gene from Clostridium saccharobutylicum and glutamate dehydrogenase RocG. In fed-batch fermentation, L-ornithine of 88.26 g/l with productivity of 1.23 g/l/h (over 72 h) and yield of 0.414 g/g glucose are achieved by the final strain in a 10-l bioreactor 1.4.1.4 glutamate dehydrogenase (NADP+) synthesis in presence of polyethyleneimine , the enzyme almost maintains the full initial activity after 2 h under conditions where the untreated enzyme retains only 20% of the initial activity, and the effect of the enzyme concentration on enzyme stability almost disappears. This stabilization is maintained in the pH range 5–9, but it is lost at high ionic strength. This polyethyleneimine -GDH composite is much more stable than the unmodified enzyme in stirred systems 1.4.1.9 leucine dehydrogenase synthesis synthesis of L-selenomethionine from 2-oxo-4-methylselenobutanoate 1.4.1.9 leucine dehydrogenase synthesis conversion of ammonia or urea into essential amino acids, L-Leu, L-Val, and L-Ile, using artificial cells containing an immobilized multienzyme system that consists of EC 1.1.1.1, EC 1.4.1.9, EC 3.5.1.5 and dextran-NAD+ 1.4.1.9 leucine dehydrogenase synthesis production of L-Leu, L-Val and L-Ile by artificial cells containing a glucose dehydrogenase and leucine dehydrogenase 1.4.1.9 leucine dehydrogenase synthesis preparation of (S)-1-cyclopropyl-2-methoxyethanamine, a key chiral intermediate for the synthesis of a corticotropin releasing factor-1 (CRF-1) receptor antagonist, by a chemoenzymatic route using leucine dehydrogenase. Synthesis of (S)-1-cyclopropyl-2-methoxyethanamine starting from methylcyclopropyl ketone. Permanganate oxidation of the ketone gives cyclopropylglyoxylic acid, which is converted to (S)-cyclopropylglycine by reductive amination using leucine dehydrogenase from Thermoactinomyces intermedius, recombinantly expressed in Escherichia coli, with NADH cofactor recycling by formate dehydrogenase from Pichia pastoris 1.4.1.9 leucine dehydrogenase synthesis coexpression with Bacillus megaterium glucose dehydrogenase in Escherichia coli for the production of L-tert-leucine. A decagram preparation of L-tert-leucine is performed at a substrate concentration of 0.6 M in 1 l scale with 99% conversion after 5.5 h, resulting in 80.1% yield and > 99% enantiomeric excess 1.4.1.9 leucine dehydrogenase synthesis coexpression with NAD+-dependent FDH from Candida boidinii in Escherichia coli for synthesis of L-tert-leucine. In a continuous feeding process, at an overall substrate concentration up to 1.5 M, both conversion and enantiomeric excess of >99% and space-time yield of 786 g/l/d are achieved 1.4.1.9 leucine dehydrogenase synthesis formation of a bifunctional enzyme complex consisting of leucine dehydrogenase (LDH) and formate dehydrogenase from Candida boidinii via a miniscaffoldin for production of L-tert-leucine. Ninety-one grams of L-tert-leucine per liter with an enantiomeric purity of 99% e.e. can be obtained 1.4.1.9 leucine dehydrogenase synthesis high-throughput screening method for L-tert-leucine synthesis and directed evolution strategy to engineer LeuDH for improved efficiency of L-tert-leucine synthesis 1.4.1.9 leucine dehydrogenase synthesis production of L-2-aminobutanoate from L-threonine via overexpression of L-threonine deaminase from Escherichia coli, L-leucine dehydrogenase from Bacillus cereus, and formate dehydrogenase from Pseudomonas sp. in Escherichia coli with formate as a cosubstrate for NADH regeneration. 30 mol L-threonine are converted to 29.2 mol L-2-aminobutanoate with 97.3 % theoretical yield and with a productivity of 6.37 g/l/h at 50 l 1.4.1.9 leucine dehydrogenase synthesis enzyme cascade from threonine to synthesis of L-2-aminobutanoate. In this cascade, the threonine deaminase is used for threonine to 2-oxobutanoate, then LeuDH mutant and formate dehydrogenase are used for synthesis of L-2-aminobutanoate. Under optimized conditions, 1 M threonine is catalyzed by whole cells of Escherichia coli harboring the enzymes in 12 h in sodium phosphate buffer to the optically pure L-2-aminobutanoate with a yield of 99% and ee above 99% 1.4.1.9 leucine dehydrogenase synthesis enzyme coupled with recombinant formate dehydrogenase is used to catalyze trimethylpyruvic acid through reductive amination to generate enantiopure L-tert-leucine. Using a fed-batch feeding strategy, up to 0.8 M of trimethylpyruvate is transformed to L-tert-leucine, with an average conversion rate of 81% and L-tert-leucine concentration of 65.6 g/l 1.4.1.9 leucine dehydrogenase synthesis in Escherichia coli expressing IvlA, mutant K72A, formate dehydrogenase, under optimized conditions 150 g L-threonine is transformed to 121 g L-2-aminobutanoate in 5 l fermenter with 95% molar conversion rate, and a productivity of 5.04 g/l and h 1.4.1.9 leucine dehydrogenase synthesis production of L-tert-leucine by a fusion enzyme of leucine dehydrogenase and glucose dehydrogenase with a rigid peptide linker (GDH-R3-LeuDH). Compared with the free enzymes, both the environmental tolerance and thermal stability of GDH-R3-LeuDH is improved. The fusion structure accelerates the cofactor regeneration rate and maintains the enzyme activity. The space-time yield of L-tert-leucine synthesis by GDH-R3-LeuDH whole cells is up to 2136 g/l/day in a 200 ml scale system under the optimal conditions (pH 9.0, 30°C, 0.4 mM of NAD+ and 500 mM of substrate including trimethylpyruvic acid and glucose) 1.4.1.9 leucine dehydrogenase synthesis production of L-tert-leucine. A coupled reaction comprising LeuDH with glucose dehydrogenase of Bacillus amyloliquefaciens results in substrate inhibition at high trimethylpyruvate concentrations (0.5 M), which is overcome by batch-feeding of the substrate. The total turnover number and specific space-time conversion of 0.57 M substrate increases to 11400 and 22.8 mmol per h and l and g, respectively 1.4.1.9 leucine dehydrogenase synthesis production of L-valine in Escherichia coli on the base of the aminotransferase B-deficient strain V1 by introducing one chromosomal copy of the Bcd gene or the IlvE gene. The Bcd-possessing strain exhibits 2.2fold higher L-valine accumulation (up to 9.1 g/l) and 2.0fold higher yield (up to 35.3%) under microaerobic conditions than the IlvE-possessing strain 1.4.1.16 diaminopimelate dehydrogenase synthesis expression of DapDH in Escherichia coli significantly enhances carbon flux into the pentose phosphate pathway and L-lysine biosynthetic pathway, thus increasing the levels of NADPH and precursors for L-lysine biosynthesis. The coexistence of two DAP-pathways and sufficient ammonium availability are good for increasing the final titer of L-lysine with a high carbon yield and productivity in Escherichia coli. Fed-batch fermentation of the target strain results in 119.5 g/l of L-lysine with a carbon yield of 49.1% and productivity of 2.99 g per l and h 1.4.1.16 diaminopimelate dehydrogenase synthesis individual overexpression of ASPDH, aspartate-semialdehyde dehydrogenase from Tistrella mobilis, dihydrodipicolinate reductase from Escherichia coli, and diaminopimelate dehydrogenase from Pseudothermotoga thermarum in Corynebacterium glutamicum LC298, a basic lysine producer, increases the production of lysine by 30.7%, 32.4%, 17.4%, and 36.8%, respectively. The highest increase of lysine production (30.7%) is observed for a triple-mutant strain (27.7 g/L, 0.35 g/g glucose) expressing ASPDH, aspartate-semialdehyde dehydrogenase from Tistrella mobilis, dihydrodipicolinate reductase from Escherichia coli. A quadruple-mutant strain expressing all of the four NADH-utilizing enzymes allows high lysine production (24.1 g/l, 0.30 g/g glucose) almost independent of the oxidative pentose phosphate pathway 1.4.1.20 phenylalanine dehydrogenase synthesis formation of L-Phe from phenylpyruvate 1.4.1.20 phenylalanine dehydrogenase synthesis synthesis of allysine (S)-2-amino-5-(1,3-dioxolan-2-yl)-pentanoic acid 1.4.1.20 phenylalanine dehydrogenase synthesis formation of L-Phe from phenylpyruvate in presence of formate-dehydrogenase from Candida boidinii 1.4.1.20 phenylalanine dehydrogenase synthesis continous production of L-Phe in an enzyme-membrane-reactor 1.4.1.20 phenylalanine dehydrogenase synthesis expression of mutant R272M/E331Q/E196N in Escherichia coli via a fed-batch fermentation strategy. After 24 h of cultivation, the specific activity of crude extracts is 0.49 U/mg. The specific activity, recovery, yield and purification fold of PheDH are 3.21 U/mg, 104.19%, 84.53% and 6.55%, respectively 1.4.1.21 aspartate dehydrogenase synthesis potential application of AspDH for cost-effective and efficient L-Asp production via both fermentative and enzymatic systems. The ability to catalyze stereospecific reactions has also stimulated research interest in amino acid dehydrogenases as biocatalysts to produce synthons for pharmaceutical and food industries, e.g., enantiomerically pure non-natural amino acids as drug precursors 1.4.1.21 aspartate dehydrogenase synthesis individual overexpression of ASPDH, aspartate-semialdehyde dehydrogenase from Tistrella mobilis, dihydrodipicolinate reductase from Escherichia coli, and diaminopimelate dehydrogenase from Pseudothermotoga thermarum in Corynebacterium glutamicum LC298, a basic lysine producer, increases the production of lysine by 30.7%, 32.4%, 17.4%, and 36.8%, respectively. The highest increase of lysine production (30.7%) is observed for a triple-mutant strain (27.7 g/L, 0.35 g/g glucose) expressing ASPDH, aspartate-semialdehyde dehydrogenase from Tistrella mobilis, dihydrodipicolinate reductase from Escherichia coli. A quadruple-mutant strain expressing all of the four NADH-utilizing enzymes allows high lysine production (24.1 g/l, 0.30 g/g glucose) almost independent of the oxidative pentose phosphate pathway 1.4.1.27 glycine cleavage system synthesis recombinant expression of H-protein in Escherichia coli. When the cells are cultured in medium supplemented with 30 microM lipoate, about 10% of the protein expressed is the correctly lipoylated active form, 10% is an inactive aberrantly modified form, and the remaining 80% is the unlipoylated apoform 1.4.1.28 secondary-alkyl amine dehydrogenase [NAD(P)+] synthesis reductive amination of cyclohexanone (up to 100 mM) into cyclohexylamine is performed with an AmDH and formate dehydrogenase system with more than 99% conversion using Escherichia coli whole cell as well as purified enzymes. Chiral amine are produced from the corresponding ketone using inexpensive ammonium formate as sole sacrificial agent and enzymes omega-transaminase, AmDH, and formate dehydrogenase 1.4.1.28 secondary-alkyl amine dehydrogenase [NAD(P)+] synthesis in the asymmetric amination of a series of structurally different hydroxy ketones, high stereoselectivity (above >99%) is achieved the reduction of aliphatic and aromatic compounds. The aliphatic substrates 1-hydroxy-4-methylpentan-2-one and 1-hydroxyhexan-2-one are aminated with 99% conversions 1.4.1.28 secondary-alkyl amine dehydrogenase [NAD(P)+] synthesis reductive amination of cyclohexanone (up to 100 mM) into cyclohexylamine using transgenic Escherichia coli as whole-cell reactors as well as purified enzymes 1.4.3.2 L-amino-acid oxidase synthesis use of enzyme as a catalyst in supercritical CO2. enzyme activity increases after exposure to supercritical conditions by up to 15%. Enzyme is more stable in supercritical CO2 than under atmospherical pressure, and oxidation of 3,4-dihydroxyphenyl-L-Ala is best under supercritical conditions 1.4.3.2 L-amino-acid oxidase synthesis engineering Escherichia coli to produce phenylpyruvate. Knock-out of three aminotransferases increases the phenylpyuvate titer from 3.3 to 3.9 g/l and the substrate conversion ratio to 97.5%. The L-amino acid deaminase triple mutant D165K/F263M/L336M produces 10.0 g phenylpyruvate per l, with a substrate conversion ratio of 100%. An optimal fed-batch biotransformation process gives 21 g phenylpyruvate per l within 8 h 1.4.3.2 L-amino-acid oxidase synthesis expression of isoform LAOO4 in the yeast Pichia pastoris with a 9His-tag and comparison with the 6His-LAAO4 expressed in Escherichia coli. The expression of the enzyme with an ER-signal sequence in Pastoris pastoris results in a glycosylated, secreted protein. The enzymatic activity without activation is higher after expression in Pichia pastoris compared to Escherichia coli. Due to treatment with acidic pH, a striking increase of activity can be detected for both expression systems resulting in similar specific activities after acid activation 1.4.3.2 L-amino-acid oxidase synthesis purified aldehyde-tagged 6His-hcLAAO4 is covalently bound to a hexylamine resin via the Calpha-formylglycine residue. The immobilized enzyme can be reused repeatedly to generate phenylpyruvate from L-phenylalanine with a total turnover number of 17600 and is stable for over 40 days at 25 °C 1.4.3.2 L-amino-acid oxidase synthesis synthesis of phenylpyruvic acid from L-phenylalanine using biotransformations with specified amounts of purified enzyme or activated lysate prepared from the whole cells. 20 mM of substrate are converted after 4 h reaction. The formation of undesired byproducts such as phenylacetic acid is suppressed using a commercially available catalase enzyme 1.4.3.3 D-amino-acid oxidase synthesis oxidation of cephalosporin C in the two-step formation of 7-aminocephalosporanic acid 1.4.3.3 D-amino-acid oxidase synthesis production of alpha-ketoadipinyl-7-aminocephalosporanic acid 1.4.3.3 D-amino-acid oxidase synthesis intracellular production of HDAO under P(GAP), followed by affinity purification and immobilization on oxirane resin, may serve as an effective process for the manufacture of immobilized DAO for industrial application 1.4.3.3 D-amino-acid oxidase synthesis immobilization of Trigonopsis variabilis D-amino acid oxidase on solid support is the key to a reasonably stable performance of this enzyme in the industrial process for the conversion of cephalosporin C as well as in other biocatalytic applications 1.4.3.3 D-amino-acid oxidase synthesis pig kidney enzyme is used to make L-pipecolic acid from a racemic mixture via D-isomer oxidation 1.4.3.3 D-amino-acid oxidase synthesis the enzyme is used to develop biocatalysts for the production of 7-aminocephalosporanic acid from the natural antibiotic cephalosporin C, and to produce optically active L-methionine from D-amino acid with the yield of 100% in a cascade system of four enzymes, or to produce phenyl pyruvate from D-phenylalanine with the yield of 99% 1.4.3.3 D-amino-acid oxidase synthesis the enzyme is used to produce phenyl pyruvate from D-phenylalanine with the yield of 99% 1.4.3.3 D-amino-acid oxidase synthesis the enzyme might be useful as a biocatalyst for industrial applications and for therapeutic treatments, mechanism of this DAAO variant and on its cytotoxicity towards various mammalian cancer cell lines, overview 1.4.3.3 D-amino-acid oxidase synthesis faster substrate turnover, reduced glutaryl-7-aminocephalosporanic acid inhibition and improved thermostability of the F54Y mutant compared to the wild-type enzyme render it a useful candidate in industrial production of semi-synthetic cephems 1.4.3.3 D-amino-acid oxidase synthesis the soluble expression of is improved in Escherichia coli through N-terminal modification, but the cell biomass is decreased. When a variant carrying an additional glycine in position 3 is fused with three N-terminus histidine residues, the volumetric activity is increased by 3.1 times and the biomass is not significant change compared with the wild type. When the N-terminal disordered region of DAAO (HSQK) is replaced with HHHG, the variant enzyme activity reaches 80.7 U/ml in a 7.5 l fermenter in 24 h 1.4.3.3 D-amino-acid oxidase synthesis use of mutant Y228L/R283G for the deracemization of racemic(RS)-1-phenylethan-1-amine with NaBH4 to produce (1S)-1-phenylethan-1-amine with an enantiomeric excess of 99% 1.4.3.3 D-amino-acid oxidase synthesis use of surface layer (S-layer) protein of Lactobacillus brevis as a self-aggregating protein tag to enable cost-effective separation of human and yeast D-amino acid oxidases expressed in Escherichia coli cells. Human and yeast D-amino acid oxidases fused with S-layer proteins can be easily separated by aggregates at the interface and/or in a few conditions by precipitates to the bottom of the PEG-phosphate aqueous system 1.4.3.4 monoamine oxidase synthesis the enzyme is used to successfully identify the alkaloid (+/-)-crispine A as a target for chemo-enzymatic deracemisation yielding the biologically active (R)-enantiomer in 97% enantiomeric excess 1.4.3.4 monoamine oxidase synthesis monoamine oxidase (MAO-N) is a synthetic tool for the enantioselective synthesis of chiral amines 1.4.3.4 monoamine oxidase synthesis when deracemizing 1,2,3,4-tetrahydro-1-methylisoquinoline on a preparative scale, the MAO-N variant W230R/W430C/C214L allows access to the (S)-enantiomer. It also shows good activity in the preparation of (S)-1,2,3,4-tetrahydro-1-ethylisoquinoline and (S)-1,2,3,4-tetrahydro-1-isopropylisoquinoline on a preparative scale 1.4.3.5 pyridoxal 5'-phosphate synthase synthesis continuous production of pyridoxal 5'-phosphate with enzyme immobilized to iodo- and bromoacetyl polysaccharides 1.4.3.10 putrescine oxidase synthesis potential of putrescine oxidase for the bioproduction of N-heterocycles from cadaverine 1.4.3.11 L-glutamate oxidase synthesis development of glutaric acid production consortium system with 2-oxoglutarate regeneration by glutamate oxidase in Escherichia coli 1.4.3.11 L-glutamate oxidase synthesis engineering of L-glutamate oxidase has great potentials to enhance the industrial production of 2-oxoglutarate. A whole-cell biocatalyst for 2-oxoglutarate production is developed by co-expression of both S280T/H533L mutant and KatE catalase 1.4.3.11 L-glutamate oxidase synthesis to simplify technological processes and reduce production costs, cascade biocatalysis for 2-oxoglutarate production is constructed by simultaneously expressing L-glutamate oxidase (LGOX) from Streptomyces viridosporus and KatG from Escherichia coli W3110 in Escherichia coli BL21 (DE3). In vivo cascade biocatalysis is constructed and optimized by promoter engineering to finely control the coexpression of LGOX and KatG, thus resulting in a significant increase in 2-oxoglutarate concentration and its conversion rate with no catalase addition 1.4.3.12 cyclohexylamine oxidase synthesis the enzyme is a useful biocatalyst for the kinetic resolution and dereacemization of amines 1.4.3.12 cyclohexylamine oxidase synthesis chemoenzymatic deracemization is applied to prepare D-valine from racemic valine ethyl ester or L-valine ethyl ester in high yield (up to 95%) with excellent optical purity (more than 99% enantiomeric excess) by employing cyclohexylamine oxidase (CHAO) variant Y321I/M226T exhibiting catalytic efficiency that is 30times higher than that of the wild type enzyme 1.4.3.12 cyclohexylamine oxidase synthesis deracemization of racemic beta-amino alcohols is conducted by cyclohexylamine oxidase-catalyzed enantioselective deamination and transaminase-catalyzed enantioselective amination to afford (S)-beta-amino alcohols in excellent conversion (78-94%) and enantiomeric excess (above 99%) 1.4.3.12 cyclohexylamine oxidase synthesis stereoselective synthesis of various enantioenriched 1-benzyl-octahydroisoquinoline derivatives of potential pharmaceutical importance at a semipreparative scale 1.4.3.13 protein-lysine 6-oxidase synthesis required for the normal biosynthesis and maturation of collagen and elastin 1.4.3.16 L-aspartate oxidase synthesis improvement of quinolinate production by assembling quinolinate synthase (NadA) and L-aspartate oxidase (NadB) as an enzyme complex 1.4.7.1 glutamate synthase (ferredoxin) synthesis bioelectrosynthesis or bioelectrooxidation of glutamate with recombinant Fd-GltS from cyanobacteria. Bioelectrocatalytic oxidation of glutamate is oxygen independent. A sodium dithionite-methyl viologen (MV2+) system is used as the artificial reducing equivalents. Dithionite acts as the primary electron donor and reduces colorless MV2+ to the purple MV+-radical with a strong absorption at 604 nm. The system is capable of catalyzing either forward synthesis or backward oxidation by choosing different redox mediators 1.5.1.3 dihydrofolate reductase synthesis develoment of a method for bacterial expression of mouse translation factor eIF4E tagged with mutant dihydrofolate reductase. Recombinant eIF4E and DHFR-eIF4E both show to significantly enhance in vitro translation in dose dependent manner by 75% at 0.0005 mM. Increased concentrations of eIF4E and DHFR-eIF4E significantly inhibit translation in a dose dependent manner by a maximum at 0.0022 mM of 60% and 90%, respectively 1.5.1.3 dihydrofolate reductase synthesis the use of the dihydrofolate-reductase-targeted short hairpin RNA vector can enhance the IgG expression in the gene-amplified stable CHO cells and uphold the IgG expression in methotrexate-free cultures 1.5.1.3 dihydrofolate reductase synthesis transformation of murine dihydrofolate reductase cDNA by a series of nested PCRs to reproduce the amino acid coding sequence for bovine DHFR, which differs from the murine sequence by 19 amino acids. Expression of the bovine dihydrofolate reductase cDNA in bacterial cells produces a stable recombinant protein with high enzymatic activity and kinetic properties similar to those previously reported for the native protein 1.5.1.3 dihydrofolate reductase synthesis application of HEK-293 cells as an alternative host cell line for stable expression of therapeutic glycoproteins. DHFR-deficient cells are generated by disrupting both DHFR and DHFR1 in HEK-293E cells and an expression vector containing DHFR and monoclonal antibody gene is transfected into the disruption mutant. A stable high-producing recombinant HEK293 cell line can be established using DHFR/methotrexate-mediated gene amplification 1.5.1.3 dihydrofolate reductase synthesis folate production by the mutant L62F in two-stage fermentation process with temperature shift-up from 30°C to 40°C increases by three-fold compared with the parental strain 1.5.1.18 ephedrine dehydrogenase synthesis the wide substrate spectrum of the dehydrogenase, combined with its regio- and enantioselectivity, suggests a high potential for the industrial production of valuable chiral compounds 1.5.1.21 1-piperideine-2-carboxylate/1-pyrroline-2-carboxylate reductase (NADPH) synthesis enzymatic system for synthesis of L-pipecolic acid from L-lysine by commercial L-lysine alpha-oxidase and extract of Escherichia coli producing recombinant enzyme and glucose dehydrogenase. System provides 27g/l of L-pipecolic acid in laboratory scale, with 99.7% enantiomeric excess 1.5.1.24 N5-(carboxyethyl)ornithine synthase synthesis enzymatic biosyntheses of amino acids instead of chemical syntheses provide an attractive alternative to the former procedures since they are efficient and simple to perform. N,N-dialkylation does not occur and the enzyme-catalyzed reactions are both regio-and stereochemically specific 1.5.1.31 berberine reductase synthesis heterologous production of berberine and the optimization of the engineered biosynthetic pathway from rac-norlaudanosoline to (S)-canadine in yeast involving the recombinant berberine reductase 1.5.1.36 flavin reductase (NADH) synthesis generation of FMNH2 by enzyme encapsulated in poly(lactide-co-glycolide) nanoparticles. Enzymatic activity, stability, and reusability of the nanoparticles prepared using three different methods [(o/w), water in oil in water (w/o/w), and solid in oil in water (s/o/w)] are compared. The solid in oil in water method provides the optimal conditions for encapsulation of the reudctase, giving the highest enzyme activity, stability, and reusability. The solid in oil in water method improves enzyme activity about 11-and 9fold compared to water in oil in water and oil in water methods. Solid in oil in water nanoparticles can be reused 14 times with nearly 50% activity remaining 1.5.1.36 flavin reductase (NADH) synthesis in vivo production of ortho-hydroxylated flavonoids by recombinant Escherichia coli. When HpaC is linked with an S-Tag on the C terminus, the enzyme activity is significantly affected. The optimal culture conditions are a substrate concentration of 80 mg/l, an induction temperature of 28°C, an M9 medium, and a substrate delay time of 6 h after IPTG induction. The efficiency of eriodictyol conversion from recombinant strains fed naringin is up to 57.67. Highest conversion efficiencies for production of catechin and caffeate are 35.2 % and 32.93%, respectively 1.5.1.36 flavin reductase (NADH) synthesis overexpression of the HpaB and HpaC genes in Saccharomyces cerevisiae achieves hydroxytyrosol titers of 1.15 mg/l and 4.6 mg/l in a minimal medium in which either 1 mM tyrosine or 1 mM tyrosol are respectively added 1.5.1.40 8-hydroxy-5-deazaflavin:NADPH oxidoreductase synthesis Tfu-FNO variants can be used as NCB-Recycling systems for TsOYE-mediated conversions. TsOYE is known to efficiently hydrogenate ketoisophorone (2,6,6-trimethyl-2-cyclohexene-1,4-dione) to form the chiral product (R)-2,2,6- trimethylcyclohexane-1,4-dione with high enantiomeric excess values of 94-97%, when several NCBs as more efficient electron donors in comparison to NADPH are used. The Tfu-FNO variants G29Y, G29W, and P89H, which show high activity toward 1-aminoethylnicotinamide (AmNA+), 1-benzylnicotinamide (BNA+), and 1-benzyl-3-acetylpyridine (BAP+), are selected for use in conversion experiments as recycling catalysts for these NCBs. Potential use of these thermostable Tfu- FNO variants as NCB recycling systems at high temperatures 1.5.1.48 2-methyl-1-pyrroline reductase synthesis (R)-imine reductase is a biocatalyst for the asymmetric reduction of cyclic imines 1.5.1.54 methylenetetrahydrofolate reductase (NADH) synthesis improvement of the biosynthesis of 5-MTHF by Lactococcus lactis by systematic pathway engineering, including the overexpression of the key rate-limiting enzyme MTHFR, strengthening 5-MTHF biosynthesis and folate supply by coexpression of the drfA, metF, and folE genes, increasing NADPH supply by overexpression of G6PDH, and increasing the proportion of 5-MTHF in various folates by overexpressing 5-formyltetrahydrofolate cyclo-ligase. The 5-MTHF titer finally reaches 300 microg/l with key precursor addition 1.5.3.13 N1-acetylpolyamine oxidase synthesis application of simple guide molecules, being either covalently attached to polyamines or used as a supplement to the substrate mixtures, for controlling the enzyme's regioselectivity and stereospecificity 1.5.3.17 non-specific polyamine oxidase synthesis application of simple guide molecules, being either covalently attached to polyamines or used as a supplement to the substrate mixtures, for controlling the enzyme's regioselectivity and stereospecificity 1.5.3.25 fructosyl amine oxidase (glucosone-forming) synthesis addition of amadoriase I during peptide glycation reaction reduces the formation of Amadori products up to 80%. The effect is more evident for hydrophobic peptides. In protein-glucose systems, the effect is dependent on the molecular weight and steric hindrance being negligible for BSA and at a maximum for insulin 1.5.5.2 proline dehydrogenase synthesis ProDH is of interest for practical applications because the proline imino acid can serve as a building block for a wide range of peptides and antibiotics 1.6.1.4 NAD(P)+ transhydrogenase (ferredoxin) synthesis improvement of hydrogen production is achieved by overexpression of membrane-integral nicotinamide nucleotide transhydrogenase PntAB and deletion of soluble pyridine nucleotide transhydrogenase SthA. A 3.9fold increased hydrogen yield is observed 1.6.3.1 NAD(P)H oxidase (H2O2-forming) synthesis usage of the enzyme for regeneration of both NADP+ and NAD+ in alcohol dehydrogenase-catalyzed enantioselective oxidation of racemic 1-phenylethanol. NADP+ regeneration at 30°C by TkNOX coupled with (R)-specific ADH from Lactobacillus kefir results in successful acquisition of optically pure (S)-1-phenylethanol, or at 45-60°C with moderately thermostable (S)-specific ADH from Rhodococcus erythropolis in optically pure (R)-1-phenylethanol, giving the possibility to operate the enantioselective bioconversion accompanying NAD+ regeneration at high temperatures, advantage of the combination of thermostable enzymes 1.6.3.3 NADH oxidase (H2O2-forming) synthesis regeneration of NAD+ utilizing this enzyme made selective oxidation of mandelic acid or L-phenylalanine possible. This thermostable enzyme is expected to be applicable as a useful biocatalyst for NAD+ recycling 1.6.3.3 NADH oxidase (H2O2-forming) synthesis the enzyme is applicable as a biocatalyst for NAD+ recycling 1.6.3.4 NADH oxidase (H2O-forming) synthesis the enzyme may prove to be useful for NAD+ regeneration in the production rare L--sugars such as L-ribose, L-ribulose, and L-xylulose 1.6.99.1 NADPH dehydrogenase synthesis production of (6R)-levodione in Escherichia coli expressing the enzyme plus glucose dehydrogenase with a molar yield of 95% 1.7.1.3 nitrate reductase (NADPH) synthesis the purified native enzyme is successfully used for synthesis of silver nanoparticles in an NADPH-dependent manner using gelatin as a capping agent, analysis by X-ray diffraction, dynamic light scattering spectroscopy, and transmission and scanning electron microscopy, overview. The stable nonaggregating nanoparticles are spherical in shape with an average size of 50 nm and a zeta potential of -34.3. The synthesized nanoparticles show a strong growth inhibitory antimicrobial activity against all tested human pathogenic fungi and bacteria, overview 1.7.1.14 nitric oxide reductase [NAD(P)+, nitrous oxide-forming] synthesis coimmobilization of nitric oxide reductase and Bacillus megaterium glucose dehydrogenase for the continuous reduction of nitric oxide via cofactor recycling, using epoxide- and carboxyl-functionalized hyperporous microspheres. Enzyme activity maintenance of 158% for nitric oxide reductase and 104% for glucose dehydrogenase can be achieved 1.7.3.3 factor-independent urate hydroxylase synthesis high-yield expression of uricase in Escherichia coli and establishment of an efficient three-step protein purification protocol. The purity of the recombinant protein is more than 98% and the specific activity is 38.4 IU/mg 1.7.3.3 factor-independent urate hydroxylase synthesis at optimized growth parameters, the crude preparation shows uricase activity of 13.42 U/ml 1.7.3.3 factor-independent urate hydroxylase synthesis at optimized growth parameters, the crude preparation shows uricase activity of 17.7 U/ml 1.7.3.3 factor-independent urate hydroxylase synthesis expression of the Escherichia-coli-codon-optimized gene as a fusion with the N-terminus of Mxe GyrA intein and chitin-binding domain for simple purification. After purification, the cleavage of the fusion protein is induced by adding DTT. Pure and properly folded uricase is obtained 1.7.3.3 factor-independent urate hydroxylase synthesis optimization of enzyme production from a bacterium isolated in deep litter poultry soil. Up to 306 U/l extracellular enzyme can be obtained 1.7.3.3 factor-independent urate hydroxylase synthesis optimum uricase production is in basal medium containing sucrose as a sole carbon source, uric acid as a nitrogen source at pH 6. Presence of cysteine HCl, cystine and glutamic acid enhances uricase production. Up to 167 U/ml can be obtained 1.8.1.2 assimilatory sulfite reductase (NADPH) synthesis cell-free synthesis of gold nanoparticles using NADPH-dependent sulfitereductase. The nanoparticles are spherical with an average size of 10 nm and a zeta potential of -30. Gold nanoparticles show strong antifungal activity toward a wide range of human pathogenic fungi 1.8.4.1 glutathione-homocystine transhydrogenase synthesis maximum activity is obtained by nutritional optimization using GSH (0.4%) combined with homocystine (0.01%) and glucose (0.4%), NADH (30 mM) at medium initial pH 7.8. The yield of GHTHase is increased about 1.2 times upon starvation of the culture for two days, as compared to the non-sulfur starved culture. The GHTHase activity is increased by 13.7fold 1.8.4.11 peptide-methionine (S)-S-oxide reductase synthesis enzyme can be useful in the development and action of anti-cancer and anti-inflammation drugs 1.8.4.11 peptide-methionine (S)-S-oxide reductase synthesis reductive resolution for the synthesis of optically active sulfoxides. Whole-cell system expressing recombinant pmMsrA protein is designed for the preparation of chiral sulfoxides with R configuration through reductive resolution of racemic sulfoxides 1.8.4.11 peptide-methionine (S)-S-oxide reductase synthesis the enzyme can effectively accomplish the preparation of (R)-sulfoxides with approximately 50% yield and 94-99% enantiomeric excess through asymmetric reductive resolution of racemic sulfoxide. With the establishment of the enzyme regeneration system, the initial substrate concentration can be increased 40-100times. The (R)-sulfoxides are obtained with high enantioselectivity under the substrate concentration up to 200 mm (approximately 32 g/L), representing a quite high substrate concentration in biocatalytic preparation of chiral sulfoxides. This system shows fairly good activity and enantioselectivity towards a series of ortho- and para-substituted phenyl methyl sulfoxides under high substrate concentration 1.8.4.11 peptide-methionine (S)-S-oxide reductase synthesis the study supports that the asymmetric reductive resolution of rac-sulfoxides using MsrA would become an effective strategy for the green synthesis of optically pure sulfoxides 1.8.4.12 peptide-methionine (R)-S-oxide reductase synthesis enzyme can be useful in the development and action of anti-cancer and anti-inflammation drugs 1.8.4.12 peptide-methionine (R)-S-oxide reductase synthesis application of methionine sulfoxide reductase B for preparation of (S)-sulfoxides through kinetic resolution of racemic sulfoxides. The kinetic resolution of rac-sulfoxides is successfully accomplished at substrate concentrations up to 50 mM (7.8 g/l) when catalyzed by akMsrB whole-cell catalyst. The study supports that asymmetric reductive resolution of rac-sulfoxides catalysed by MsrB can become an effective strategy for green synthesis of optically pure sulfoxides 1.8.4.13 L-methionine (S)-S-oxide reductase synthesis enzyme can be useful in the development and action of anti-cancer and anti-inflammation drugs 1.8.7.2 ferredoxin:thioredoxin reductase synthesis dicistronic construct for the heterologous expression in Escherichia coli. The coding sequences for the two mature subunits have been inserted in tandem into the expression vector. The dicistronic construct is correctly translated yielding soluble, perfectly functional FTR. The recombinant enzyme is composed of both subunits, contains the correctly inserted FeS cluster and is indistinguishable from the enzyme isolated from leaves in its capacity to activate chloroplast fructose-1,6-bisphosphatase 1.8.99.2 adenylyl-sulfate reductase synthesis production of adenosine 5'-phosphosulfate labeled with either 14C or 35S 1.10.3.2 laccase synthesis addition of phenolic and aromatic monomers to growth medium to enhance enzyme production, ferulic acid plus vanillin are most efficient inducers increasing enzyme production up to 10 times 1.10.3.2 laccase synthesis Pycnoporus sp. SYBC-L1 is a potential candidate for laccase production 1.10.3.2 laccase synthesis owing to their broad substrate range laccases are considered to be versatile biocatalysts which are capable of oxidizing natural and non-natural industrial compounds, with water as sole by-product 1.10.3.2 laccase synthesis potential use of the laccase in lignin modification 1.10.3.2 laccase synthesis selective biotransformation of aromatic methyl group to aldehyde group in presence of diammonium salt of 2,2'-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid) as the mediator 1.10.3.2 laccase synthesis the enzyme will serve as a useful tool for enzymatic polymerization of diphenolic compounds such as caffeic acid and ferulic acid 1.10.3.2 laccase synthesis synthesis of bioactive 1,4-naphthoquinones. A high yield of naphthoquinones (74.93%) with 1,4-naphthoquinone (60.61%), and its derivative 2-hydroxy-1,4-naphthoquinone (14.32%) is obtained at the optimized reaction conditions 1.10.3.2 laccase synthesis synthesis of the C-N polydye at basic pHs 1.10.3.2 laccase synthesis the enzyme from Crinipellis sp. synthesizes oxaflavins for redox co-enzymes 1.10.3.2 laccase synthesis the enzyme from Pycnoporus cinnabarinus synthesizes 1. benzofuropyroles, which are potent pharmaceutical agents, 2. the pharmaceutical agent 6,7-dihydroxy-2,2-dimethyl-1,3,9-trioxa-fluoren-4-one 1.10.3.2 laccase synthesis the enzyme from Trametes hirsute synthesizes polyanniline 1.10.3.2 laccase synthesis the enzyme from Trametes versicolor synthesizes 1. polycatechol, a valuable polymer used as a chromatographic resin and in the formation of thin films for biosensors, 2. benzofuranones for medicinal chemistry, 3. poly allylamine with high antioxidant potential, 4. dyes used in hair dyeing, 5. benzoquinones used as intermediates in pharmaceuticals, 6. phenazine and phenoxazinone chromopheres for synthetic dyes 1.10.3.2 laccase synthesis the enzyme from Trametes villosa synthesizes benzofurans with antimicrobial and anti-inflammatory activities 1.10.3.2 laccase synthesis the enzyme from Ustilago maydis synthesizes polymers of quercitin and kampferol with improved antioxidant properties of the polymers compared to the monomers 1.10.3.11 ubiquinol oxidase (non-electrogenic) synthesis the AOX gene is an effective tool in metabolic engineering for efficient organic acids production from carbohydrates 1.11.1.1 NADH peroxidase synthesis Cu2+-ion-modified graphene oxide nanoparticles catalyze the H2O2 oxidation of NADH to the biologically active NAD+ cofactor. The catalytic NAD+ cofactor regeneration system may be coupled to NAD+-dependent enzymes 1.11.1.B2 chloride peroxidase (vanadium-containing) synthesis enzyme converts both aromatic and aliphatic alkenes into the corresponding halohydrins. Variation of pH value and addtion of ethanol as solvent leads to variation of product mixtures 1.11.1.6 catalase synthesis development of simple methods for production and purification of catalases, determination of adsorption capacity and effects upon binding on enzyme activity of different minerals, binding capacities and activities at different pH/pI, one of the most promising adsorbent is hydroxylapatite, overview 1.11.1.6 catalase synthesis covalent immobilization of catalase on florisil via glutaraldehyde. Optimal immobilization is at pH 6.0, 10°C, leading to a vmax of immobilized enzyme of 20 mM H2O2 per min and mg protein and a 50fold increase in Km value. the immobilized enzyme retains 40% of initial activtiy after 50 uses and is more stable than free enzyme 1.11.1.6 catalase synthesis high-density fermentation of recombinant Pichia pastoris in a 1-l-fermentor yields 684800 U/l of catalase, measured with permeabilized cells. Catalase shows a half-life of 4 days at 30°C and cells can be reused for synthesis up to 13 times. Permeabilized cells co-expressing catalase and D-amino acid oxidase convert D-phenylalanine into 99% phenylpyruvate within 100 min. In a batchwise conversion of cephalosporin C, about 90% 7-beta-(4-carboxybutanamido)-cephalosporanic acid is obtained at each cycle 1.11.1.6 catalase synthesis method for large-scale expression and purification of recombinant catalase in Pichia pastoris 1.11.1.6 catalase synthesis production of recombinant Bacillus subtilis catalase and purification from culture fluid. Purified enzyme has a specific activity of 34600 U/mg and is more resistant to acidic conditions than bovine liver catalase 1.11.1.6 catalase synthesis a total soluble catalase activity of 78,762 U/ml with the extracellular ratio of 92.5% is achieved by fed-batch fermentation in a 10 l fermentor 1.11.1.6 catalase synthesis glutathione-mediated superoxide generation in an aqueous solution is increased by presence of catalase. The catalase-exaggerated extracellular superoxide generation may give a harmful effect to living cells 1.11.1.6 catalase synthesis immobilization of enzyme onto an epoxy support at 25, 40, and 55°C. All preparations show higher stability than the free enzyme at alkaline pH 10.0. The 55°C immobilizate shows the highest thermal stability 1.11.1.6 catalase synthesis immobilized whole cells of the bacterium demonstrate the degradation of hydrogen peroxide (H2O2) in a packed bed reactor 1.11.1.6 catalase synthesis improvement of thermal stability, resistance to protease degradation, and resistance to ascorbate inhibition, while retaining enzyme structure and activity, by conjugation to poly(acrylic acid). 55-80% and 90-100% activity is retained for all samples synthesized at pH 5.0 and pH 7.0, respectively, Km or Vmax values do not differ significantly from those of the free enzyme. Conjugates synthesized at pH 7.0 are thermally stable up to 85-90°C, and retain 40-90% of their original activities after storing for 10 weeks at 8°C 1.11.1.7 peroxidase synthesis expression of 19 individual HRP isoenzymes recombinantly in the yeast Pichia pastoris and optimization of a 2-step purification strategy 1.11.1.7 peroxidase synthesis heterologous production in Pichia pastoris is greatlyenhanced by the addition of hemin with a titer of 0.41 U/ml peroxidase activity at the second day ofincubation 1.11.1.7 peroxidase synthesis preparation of starch-g-poly(butyl acrylate) copolymers using horseradish peroxidase, in the presence of hydrogen peroxide and acetylacetone. Poly(butyl acrylate) is successfully grafted onto starch by HRP-mediated graft copolymerization, improving the thermal stability of starch 1.11.1.7 peroxidase synthesis protein engineering to obtain an active and stable HRP variant with reduced surface glycosylation during production in Pichia pastoris. Mutant N13D/N57S/N255D/N268D produced in the Pichia pastoris benchmark strain shows considerable catalytic activity and thermal stability and is less glycosylated 1.11.1.7 peroxidase synthesis self-crosslinking of fibroin molecules and p-hydroxyphenylacetamide, as a model compound of tyrosine residues in fibroins, using the hydrogen peroxide-horseradish peroxidase system. Incubation leads to p-hydroxyphenylacetamide polymerization, and the molecular weight of fibroin proteins is also noticeably increased by treatment. The mechanical properties and thermal behavior for the modified fibroin membrane are improved. The obtained membrane exhibits good biocompatibility 1.11.1.8 iodide peroxidase synthesis expression and purification of recombinant protein from AD-293 cells unsing the aminoterminal cDNA sequence of luciferase protein from Gaussia princeps for secretion. A simplified purification procedure consists of a couple of anionexchange chromatography columns 1.11.1.8 iodide peroxidase synthesis baculovirus-mediated expression of a truncated recombinant TPO proteine. The signal peptide from the peptidyl-glycine alpha-amidating monooxygenase significantly promotes the secretion in High Five cells when fused to TPO. An optimized scale-up production procedure for TPOt in a 5-L wave-type bioreactor and subsequent purification achieving a protein purity of >95% give a protein with high sensitivity and specificity in reactions with positive or negative human serum samples via the double-antigen sandwich method 1.11.1.9 glutathione peroxidase synthesis construction of a smart artificial glutathione peroxidase triggered by directed self-assembly of supraamphiphiles composed of cyclodextrin-based telluronic acid. The nanostructures conveniently link the catalytic center of glutathione peroxidase and thermosensitive polymer to the host cyclodextrin molecules.By changing the temperature, the peroxidase activity can be switched on and off 1.11.1.9 glutathione peroxidase synthesis construction of recombinant proteins carrying both the activity of human glutathione peroxidase and Alvinella pompejana superoxide dismutase and expression in a single protein production system. The best construct displays excellent antioxidant capacity and synergistic effects of hte two activities 1.11.1.9 glutathione peroxidase synthesis preparation of a supramolecular artificial glutathione peroxidase based on a supramolecular graft copolymer and the catalytic center ADA-Te-OH. The artificial glutathione peroxidase was constructed by supramolecular host-guest self-assembly and displays noticeable temperature-dependent catalytic activity and typical saturation kinetics behavior. The change in the self-assembled structure during the temperature-dependent process plays a significant role in the temperature-dependent catalytic behavior 1.11.1.10 chloride peroxidase synthesis oxidation of hexen-1-ols with tert-butyl hydroperoxide 1.11.1.10 chloride peroxidase synthesis synthesis of semiconductor polymers 1.11.1.10 chloride peroxidase synthesis chloroperoxidase (CPO) has long been recognized as a powerful and versatile catalyst, potential of co-immobilized enzymes chloroperoxidase and glucose oxidase as MGO-GOx-CPO in environmental applications 1.11.1.10 chloride peroxidase synthesis Musa paradisiaca plant juice chloroperoxidase is a potential biocatalyst for organic epoxidation reactions 1.11.1.11 L-ascorbate peroxidase synthesis overexpression of thylakoidal isozyme, increased resistance to the herbicide Paraquat but not to photoinhibitory treatments ot iron or copper overload. Sodium nitroprusside partially inhibits enzyme activity 1.11.1.13 manganese peroxidase synthesis expression of active manganese peroxidase in an Escherichia coli cell-free protein synthesis system, and optimization of reaction conditions such as the concentrations of hemin, calcium ions, and disulfide bond isomerase. Cell-free synthesized manganese peroxidase purified using the hemagglutinin tag shows higher specific activity than the commercial wild-type enzyme 1.11.1.13 manganese peroxidase synthesis the enzymes laccase and manganese peroxidase from Klebsiella pneumoniae are employed for ethanol production from rice and wheat bran biomass which shows 39.29% improved production compared to control, evaluation 1.11.1.14 lignin peroxidase synthesis evaluation of the effect of enzyme dosage, incubation time, and H2O2 addition profile on lignin activation by quantifying the phenoxy radicals formed using electron paramagnetic resonance spectroscopy. At optimal conditions, i.e. dose of 15 /g and continuous addition of H2O2, the content of phenoxy radicals is doubled as compared with an untreated control 1.11.1.14 lignin peroxidase synthesis immobilization of enzyme by entrapping in xerogel matrix of trimethoxysilane and proplytetramethoxysilane to maximum immobilization efficiency of 88.6%. The free and immobilized enzymes have optimum pH values of 6 and 5 while optimum temperatures are 60°C and 80°C, respectively. Immobilization enhances the activity and thermo-stability potential significantly and immobilized enzyme remains stable over broad pH and temperature range 1.11.1.16 versatile peroxidase synthesis expression of enzyme under control of the alcohol dehydrogenase promoter of Aspergillus nidulans in this host. Culture temperature of 19°C enhances the level of peroxidase 5.8-fold and reduces the effective proteolytic activity twofold giving a peroxidase activity of 466 units per l. Application of the same conditions to expression in Aspergillus niger does not result in improved peroxidase activity, while the effective proteolytic activity is increased 1.11.1.16 versatile peroxidase synthesis production of the oxidase in strain BL21(DE3)pLysS, cultivated at 25°C in auto-induction medium and presence of heme 1.11.1.16 versatile peroxidase synthesis evaluation of the effect of enzyme dosage, incubation time, and H2O2 addition profile on lignin activation by quantifying the phenoxy radicals formed using electron paramagnetic resonance spectroscopy. At optimal conditions, i.e. dose of 15 /g and continuous addition of H2O2, the content of phenoxy radicals is doubled as compared with an untreated control 1.11.1.16 versatile peroxidase synthesis enzyme is produced by solid-state fermentation, using the agricultural residue banana peel as growth medium. An enzymatic activity of 10800 U/l i.e. 36 U/g of substrate is detected after 18 days, whereas only 1800 U/l are reached by conventional submerged fermentation with glucose-based medium 1.11.1.18 bromide peroxidase synthesis immobilization of enzyme on magnetic micrometre-sized particles in quantitative yields, with up to 40% retention of initial bromoperoxidase activity. The immobilized enzyme is stable with a half-life time of about 160 days. It serves as reusable catalyst for bromide oxidation with H2O2 in up to 14 consecutive experiments. Immobilized enzyme is used for methyl pyrrole-2-carboxylate conversion into derivatives of naturally occurring compounds e.g. from Agelas oroides with product selectivity of up to 75% 1.11.1.21 catalase-peroxidase synthesis biocatalytic method for stereoselective oxidation of beta-lactams, such as penicillin-G, penicillin-V and cephalosporin-G to their (R)-sulfoxides 1.12.1.2 hydrogen dehydrogenase synthesis coupling of enzyme reaction to carbonyl reductase from Candida parapsilosis leads to total turnover numbers up to 143666 with maximum activity for NAD+ reduction at 35°C and pH 8.0. Half-life of enzyme is 5.3 h under these conditions. Presence of ammonium and potassium ions increases the enzyme stability 1.12.1.2 hydrogen dehydrogenase synthesis immobilization of enzyme onto the anionic resin AmberliteTMFPA54 with immobilisation yield of 93.4% for adsorptive and 100% forcovalent attachment, corresponding activities of 48.9 and 39.3%, respectively, leading to stabilisation of enzyme. At elevated temperature and under agitation, stabilisation is further increased by modification of the covalently bound SH with methoxy-poly(ethylene) glycol(mPEG). In stationary aqueous solution, half-lives of up to 161 h at 25°C and 32 h at 35°C are obtained.In presence of the DMSO, DMF, 2-propanol and [EMIM][EtSO4] half-lives of 14-29 h can be achieved 1.12.1.2 hydrogen dehydrogenase synthesis system for cloning and expression of multiple genes in Escherichia coli BL21 demonstrate tby production and maturation of the NAD+reducing soluble [NiFe]-hydrogenase from Cupriavidus necator H16. The enzyme encoded in hoxFUYHI is successfully matured by co-expression of a dedicated set of auxiliary genes, comprising seven hyp genes (hypC1D1E1A2B2F2X) along with hoxW, which encodes a specific endopeptidase. Deletion of genes involved in enzyme maturation reduces maturation efficiency substantially. Further addition of hoxN1, encoding a high-affinity nickel permease, considerably increases maturation efficiency in Escherichia coli 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis biohydrogen production 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis biohydrogen production. Photochemical hydrogen production system using zinc porphyrin and hydrogenase in a micellar system of cetyltrimethylammonium bromide. Cetyltrimethylammonium bromide acts as a cationic surfactant to effectively separate the charges 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis biohydrogen production. The enzyme is immobilized between two layers of montmorillonite clay and poly(butylviologen) mixture. The amount of hydrogen produced relates closely to the applied potential, buffer pH and temperature 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis direct production and direct in situ regeneration of NADPH 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis biohydrogen production from sugars using a mixture of enzymes in an in vitro cell-free synthetic pathway. Development of this process at an industrial scale is limited by the availability of the H2-producing enzyme 1.12.1.3 hydrogen dehydrogenase (NADP+) synthesis the study provides a cost-efficient method to obtain hyperthermostable hydrogenases, which can be used in in vitro synthetic enzymatic biosystems for cofactor regeneration and hydrogen production. It has great potential in biocatalysis, bioelectrochemistry, and clean energy production 1.12.1.5 hydrogen dehydrogenase [NAD(P)+] synthesis the recombinant enzyme is efficient in vitro biohydrogen production 1.12.2.1 cytochrome-c3 hydrogenase synthesis adsorption of cytochrome c3 at a pyrolytic graphite electrode is observed in the room-temperature ionic liquids 1-butyl-3-methylimidazolium tetrafluoroborate, 1-ethyl-3-methylimidazolium tetrafluoroborate and 1-ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide. The electrochemical signal differs however from that obtained in aqueous buffer, and depended on the type of room-temperature ionic liquids. 1-Ethyl-3-methylimidazolium bis(trifluoromethanesulfonyl)imide, as a hydrophobic non-water-miscible room-temperature ionic liquids, stabilizes the native form of cytochrome c3 and allows an amount of electroactive protein 30fold higher than observed in aqueous buffer. Catalytic oxidation of H2 via [NiFe] hydrogenase mediated by cytochrome c33 fails however, possibly due to inhibition of the hydrogenase in presence of room-temperature ionic liquids 1.12.99.6 hydrogenase (acceptor) synthesis endogenous hydrogenases Hyd-1 and Hyd-2 can be utilized to engineer H2 in a microbial cell factory manner 1.12.99.6 hydrogenase (acceptor) synthesis hydrogenases Hyd-1 and Hyd-2 are engineered to enable the cells to efficiently utilize hydrogen gas as a source of reducing equivalents, which increased the yield of reduced fermentation products, e.g. succinate or lactate. By upregulating the expression of the hydrogenase, the spectrum of fermentation products shifts toward the reductive end compared to that of the wild type 1.13.11.1 catechol 1,2-dioxygenase synthesis production of cis,cis-muconic acid, which is a raw material for new resins, pharmaceuticals and agrochemicals 1.13.11.1 catechol 1,2-dioxygenase synthesis the enzyme is involved in synthesis of cis,cis-muconic acid in a greener and cleaner way. The recombinant Escherichia coli expressing high activity catechol 1,2-dioxygenase from Paracoccus sp. MKU1 holds promise as a potential candidate for yielding high concentrations of cis,cis-muconic acid at faster rates in low cost settings. Industrial importance of cis,cis-muconic acid as a precursor for the synthesis of several biopolymers 1.13.11.12 linoleate 13S-lipoxygenase synthesis application of the enzyme in the production of 13-hydroxyoctadecadienoic acid 1.13.11.20 cysteine dioxygenase synthesis the mechanism of cysteine dioxygenase-catalyzed C-F bond cleavage of 3,5-difluoro-1-tyrosine contains four elementary steps: H-abstraction, C-S bond formation, F-transfer, and C-F bond cleavage. C-F bond cleavage is the rate-determining step with an energy barrier of 18.8 kcal/mol 1.13.11.24 quercetin 2,3-dioxygenase synthesis optimization of enzyme production by variation of rutin concentration, nitrogen source and concentration, salt and metal salt concentration, yeast extract concentration and pH value. six-fold improvement of enzyme activity reaching a maximum activity of 0.000708 mM per min and ml of culture supernatant 1.13.11.27 4-hydroxyphenylpyruvate dioxygenase synthesis overexpression of the HppD gene in Escherichia coli to produce large amounts of pyomelanin for biotechnological purposes. The recombinant dioxygenase expression leads to the production of pyomelanin in Escherichia coli. When cells are grown at the mid-exponential phase in a mineral medium with added glucose 10 mM and casamino acid 0.2 % w/v, the administration of tyrosine 1 mM after 30 minutes of exposure to arabinose 1 % w/v allows to purify 213 mg/l of pyomelanin after 6 days of biotransformation 1.13.11.33 arachidonate 15-lipoxygenase synthesis production of 13-hydroxy-14,15-epoxy-eicosatrienoic acid (14,15-hepoxilin B3, 14,15-HXB3) and 13,14,15-trihydroxyeicosatrienoic acid (13,14,15-trioxilin B3, 13,14,15-TrXB3) from arachidonic acid in Escherichia coli expressing Archangium violaceum 15-LOX in presence and absence of Myxococcus xanthus epoxide hydrolase. Under the optimal conditions of 30 g cells/l, 200 mM ARA, 25°C, and initial pH 7.5, the cells convert 200 mM arachidonic acid into 192 mM 14,15-HXB3 and 100 mM 13,14,15-TrXB3 for 150 min, with conversion yields of 96% and 51% 1.13.11.34 arachidonate 5-lipoxygenase synthesis synthesis of 5-hydroperoxyeicosatetraenoic acid and 5(S),12(S)-dihydroxy-6,10-trans-8,14-cis-eicosatetraenoic acid 1.13.11.39 biphenyl-2,3-diol 1,2-dioxygenase synthesis a multistep conversion system composed of Pseudomonas sp. phenol hydroxylase (PHIND) and Burkholderia xenovorans 2,3-dihydroxy-biphenyl 1,2-dioxygenase (BphCLA-4) from is used to synthesize methylcatechols and semialdehydes from o- and m-cresol 1.13.11.40 arachidonate 8-lipoxygenase synthesis expression in Escherichia coli and purification method 1.13.11.52 indoleamine 2,3-dioxygenase synthesis production of 2,3-dihydro-1H-indole-2,3-diol 1.13.11.58 linoleate 9S-lipoxygenase synthesis the enzyme could be useful for in vitro synthesis of 9,10-ketol-octadecadienoic acid which is expected to be an ornamentally useful flower inducer when applied exogenously 1.13.11.58 linoleate 9S-lipoxygenase synthesis the yeast expressed Nb-9-LOX can be used to produce C9-aldehydes on a large scale in combination with a HPL gene with 9-hydroperoxide lyase function, or to effectively produce 9-hydroxy-10(E),12(Z)-octadecadienoic acid in a biocatalytic process in combination with cysteine as a mild reducing agent 1.13.11.60 linoleate 8R-lipoxygenase synthesis production of (7S,8S)-7,8-dihydroxy-(9Z,12Z)-octadecadienoic acid from linoleic acid by recombinant Escherichia coli. The optimal reaction conditions are pH 7.0, 18.6°C, 10.8% (v/v) dimethyl sulfoxide, 44.9 g/l cells, and 14.3 g/l linoleic acid, with agitation at 256 rpm. Under these conditions, recombinant cells produce 7,8-dihydroxy unsaturated fatty acids in the range of 7.0-9.8 g/l from 14.3 g/l linoleic acid, 14.3 g/l oleic acid, and plant oil hydrolysates such as waste oil and olive oil containing 14.3 g/l linoleic acid or oleic acid 1.13.11.62 linoleate 10R-lipoxygenase synthesis under the optimized conditions, the biotechnological production of (8E,10R,12Z,15Z)-10-hydroperoxyoctadeca-8,12-trienoate and (8E,10R,12Z)-10-hydroperoxyoctadeca-8,12-dienoate by enzyme PpoC from Aspergillus nidulans strain ATCC 10074 from linoleic acid, alpha-linolenic acid, and hempseed oil hydrolyzate as substrates is achieved, method optimization 1.13.11.63 beta-carotene 15,15'-dioxygenase synthesis production of retinal from beta-carotene using recombinant enzyme. The optimum pH, temperature, substrate and detergent concentrations, and enzyme amount for effective retinal production are 9.0, 37°C, 200 mg per ml beta-carotene, 5% w/v Tween 40, and 0.2 U per ml enzyme, respectively. Under optimum conditions, the recombinant enzyme produces 72 mg per ml retinal in a 15 h reaction time, with a conversion yield of 36% w/w 1.13.11.63 beta-carotene 15,15'-dioxygenase synthesis production of all-trans-retinal by recombinant enzyme. Toluene is an optimal solvent for the dissolution of beta-carotene, and the optimal solution for the conversion reaction contains 2.4% (w/v) Tween 20, 0.15 U enzyme/ml, and 350 mg beta-carotene/l. Under these conditions, the enzyme produces 181 mg retinal/l after 20 h 1.13.11.63 beta-carotene 15,15'-dioxygenase synthesis stepwise cleavage by BCO2, i.e. beta-carotene-9',10'-oxygenase, and BCO1 with beta-apo-10'-carotenol as an intermediate can provide a mechanism to tailor asymmetric carotenoids such as beta-cryptoxanthin for vitamin A production 1.13.11.71 carotenoid-9',10'-cleaving dioxygenase synthesis production of beta-apo-10'-carotenal by recombinant enzyme. The optimum pH, temperature, detergent type, and the optimum concentrations of detergent, substrate, and enzyme for beta-apo-10'-carotenal production are 8.0, 37°C, Tween 40, 2.4%, 300 mg beta-carotene/l, and 0.25 U/ml, respectively, giving 43 mg beta-apo-10'-carotenal/l after 21 h with a conversion of 14% 1.13.11.71 carotenoid-9',10'-cleaving dioxygenase synthesis enzyme ScBCO2 potentially can be used for the enzymatic biotransformation of beta-carotene to beta-apo-10'-carotenal in biotechnological applications 1.13.11.85 exo-cleaving rubber dioxygenase synthesis optimization of a transformation system via electroporation, and a conjugation system for expression of RoxA in Xanthomonas sp. About 6 mg purified RoxA are obtained from 1 l of cell-free culture fluid 1.13.11.87 endo-cleaving rubber dioxygenase synthesis production of functionalized oligo-isoprenoids by enzymatic cleavage of rubber. A method for the preparation of reactive oligo-isoprenoids that can be used to convert polyisoprene latex or rubber waste materials into value-added molecules, biofuels, polyurethanes or other polymers. Emulsions of polyisoprene (latex) are treated with RoxAXsp, RoxBXsp, LcpK30 or with combinations of the three proteins. The cleavage products are purified by solvent extraction and FPLC separation. All products have the same general structure with terminal functions (CHO-CH2- and -CH2-COCH3) but differ in the number of intact isoprene units in between 1.13.11.88 isoeugenol monooxygenase synthesis Escherichia coli cells expressing isoeugenol monooxygenase produce 28.3 g vanillin/l from 230 mM isoeugenol, with a molar conversion yield of 81% at 20°C after 6 h. No accumulation of undesired by-products, such as vanillic acid or acetaldehyde, is observed 1.13.11.88 isoeugenol monooxygenase synthesis the vanillin producing activity is induced by presence of isoeugenol. Under the optimized reaction conditions, Pseudomonas putida cells produce 16.1 g/l vanillin from 150 mM isoeugenol, with a molar conversion yield of 71% at 20 °C after a 24-h incubation in the presence of 10% (v/v) dimethyl sulfoxide 1.13.11.88 isoeugenol monooxygenase synthesis upon expression of mutant F281Q in Escherichia coli and employing sol-gel chitosan membrane for removing the produced vanillin from the biotransformation system, under the optimal conditions (pH 8.5, 30°C, 180 rpm, 0.5 g wet cells, 0.1 g chitosan membrane), vanillin concentrations reach 4.5 g/l after about 40 h 1.13.11.92 fatty acid alpha-dioxygenase synthesis biotechnological production of C12-C15 branched-chain fatty aldehydes. The enzyme efficiently transforms short-chained fatty acids. The transformation products (C12-C15 iso- and anteisoaldehydes) exhibit green-soapy, sweety odors with partial citrus-like, metallic, peppery, and savory-tallowy nuances 1.13.11.92 fatty acid alpha-dioxygenase synthesis synthesis of fatty aldehydes based on Escherichia coli cells expressing alpha-dioxygenase from Oryza sativa. In resting cells of recombinant Escherichia coli, conversion of different fatty acids to the respective fatty aldehydes shortened by one carbon atom is demonstrated. The usage of Triton X 100 improves the conversion rate up to 1 g aldehyde per liter per hour 1.13.12.2 lysine 2-monooxygenase synthesis installation of an Escherichia coli whole-cell biocatalytic system, environmentally friendly, for industrial production of 5-aminovalerate using recombinant enzymes 5-aminovaleramide amidohydrolase and L-lysine 2-monooxygenase, overview 1.13.12.2 lysine 2-monooxygenase synthesis L-lysine is a potential feedstock for the production of bio-based precursors for engineering plastics. Usage of the enzyme in a microbial process for high-level conversion of L-lysine into 5-aminovalerate that can be used as a monomer in nylon 6,5 synthesis, method overview 1.13.12.2 lysine 2-monooxygenase synthesis the enzyme can be used for production of 5-aminovalerate, a potential C5 platform chemical for synthesis of valerolactam, 5-hydroxyvalerate, glutarate, and 1,5-pentanediol. Escherichia coli is engineered for production of 5-aminovalerate from L-lysine by coupled reaction of recombinant DavB, L-lysine monooxygenase, and recombinant DavA, 5-aminovaleramidase. Because L-lysine is an industrial fermentation product, the two-enzyme coupled system presents a promising alternative for the production of 5-aminovalerate 1.13.12.2 lysine 2-monooxygenase synthesis 2-monooxygenase (DavB) and delta-aminovaleramidase (DavA) are co-expressed in Escherichia coli BL21(DE3) to produce nylon-5 monomer 5-aminovalerate from L-lysine. PP2911 (4-aminobutyrate transporter in Pseudomonas putida) and LysP (the lysine specific permease in Escherichia coli) are overexpressed to promote 5-aminovalerate production using whole cells of recombinant Escherichia coli. The constructed Escherichia coli strain overexpressing transport proteins exhibits good 5-aminovalerate production performance and might serve as a promising biocatalyst for 5-aminovalerate production from L-lysine. This strategy not only shows an efficient process for the production of nylon monomers but also might be used in production of other chemicals 1.13.12.19 2-oxoglutarate dioxygenase (ethene-forming) synthesis biological ethylene production can be achieved via expression of the ethylene-forming enzyme (EFE), found in some bacteria and fungi. It has the potential to provide a sustainable alternative to steam cracking. Ethylene is an important industrial compound for the production of a wide variety of plastics and chemicals 1.13.12.24 calcium-regulated photoprotein synthesis expression in Saccharomyces cerevisiae. The cells accumulate sufficient amounts of recombinant apoaequorin. The protein is active and not toxic to the cells 1.13.12.24 calcium-regulated photoprotein synthesis overexpression of apoaequorin in Escherichia coli, solubilization of inclusion bodies with urea, purification and refolding of His6-apoaequorin in a single chromatographic step by immobilized metal-ion affinity chromatography using Ni2+-nitrilotriacetic acid agarose.The purity is greater than 80%, the yield is 0.7–1mg apoaequorin from a 50 ml bacterial culture. The luminescence of the purified aequorin is a linear function of its concentration extending over six orders of magnitude 1.14.11.2 procollagen-proline 4-dioxygenase synthesis viral P4H (A085R) is coexpressed as a soluble and active monomer in Escherichia coli with full-length human collagen alpha1(III) chain (COL3A1). This coexpression system can be used for the production of hydroxylated human collagen alpha1(III) chain (COL3A1) in Escherichia coli. The method avoids the complexity of P4H assembly 1.14.11.9 flavanone 3-dioxygenase synthesis functional expression of plant-derived O-methyltransferase, flavanone 3-hydroxylase, and flavonol synthase in Corynebacterium glutamicum for production of pterostilbene, kaempferol, and quercetin 1.14.11.9 flavanone 3-dioxygenase synthesis expression of heterologous dioxygenase genes in (2S)-flavanone-producing Corynebacterium glutamicum strains enables the production of flavanonols and flavonols, e.g kaempferol and quercetin, starting from thephenylpropanoids p-coumaric acid and caffeic acid 1.14.11.11 hyoscyamine (6S)-dioxygenase synthesis overexpression of the enzyme (H6H) enhances the biosynthesis of anisodamine and scopolamine in hairy root cultures of Scopolia lurida, which should improve its commercial value. The tropane alkaloids anisodamine and and scopolamine are clinically used as anticholinergic agents 1.14.11.20 deacetoxyvindoline 4-hydroxylase synthesis maximal production of vindoline coincides with maximal activity of enzyme and of deacetylvindoline acetyl-CoA acetyl transferase and tryptophan decarboxylase 1.14.11.20 deacetoxyvindoline 4-hydroxylase synthesis artemisinic acid can be used as a promising elicitor, especially for the production of therapeutically important indole alkaloids 1.14.11.21 clavaminate synthase synthesis chemo-enzymatic synthesis of bicyclic gamma-lactams using recombinant purified CAS2 isozyme, reaction mechanism 1.14.11.21 clavaminate synthase synthesis clavulanic acid intermediates: deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid are heterologously produced in Streptomyces venezuelae recombinant using four sets of early genes from the clavulanic acid biosynthetic pathway 1.14.11.26 deacetoxycephalosporin-C hydroxylase synthesis expression in Escherichia coli, overexpression as an insoluble and inactive enzyme, elaboration of refolding scheme resulting in highly active and moderately stable enzyme 1.14.11.45 L-isoleucine 4-hydroxylase synthesis development of efficient method for the biotransformation of 4-hydroxyisoleucine by resting cells expressing L27I/E80D/G169H/S182D 151.9 mM of 4-hydroxyisoleucine/l (22.4 g/l) can be synthesized in 12 h while the substrates seldom exhibits additional consumption 1.14.11.45 L-isoleucine 4-hydroxylase synthesis double mutant I162T/T182N shows improvements in specific activity, protein expression level, and fermentation titer of 3.2-, 2.8-, and 9.4fold, respectively. L-Isoleucine (228 mM) is completely converted to (2S,3R,4S)-4-HIL with a space-time yield of up to 80.8 g/l and d. With a increase of the substrate loading to 1 M, a high conversion of 91% can also be achieved 1.14.11.45 L-isoleucine 4-hydroxylase synthesis dynamic regulation of IDO expression by modified Ile biosensors increases the 4-HIL titer from 24.7 mM to 28.9?74.4 mM and may yield more 4-HIL than the static strain overexpressing IDO by the strong PtacM promoter (69.7 mM). Synergistic modulation of 2-oxoglutarate supply and O2 supply improves the 4-HIL production significantly, and the highest titer achieved is 135.3 mM 1.14.11.45 L-isoleucine 4-hydroxylase synthesis improved synthesis of 4-HIL by ribosomal binding site engineering for gene expression in Corynebacterium glutamicum. To supply the cosubstrate 2-oxoglutarate at different levels, the OdhI gene is expressed using the ribosomal binding site sequences. The O2 supply is further enhanced in by overexpressing the Vgb gene. 4-HIL (up to 119.27 mM) is produced in the best strain. The synchronic supply of cosubstrates 2-oxoglutarate and O2 is critical for the high-yield production of 4-HIL 1.14.11.45 L-isoleucine 4-hydroxylase synthesis improved synthesis of 4-HIL in an optimized strain of Corynebacterium glutamicum by application of programming adaptive laboratory evolution. The programming evolutionary system contains a Lys biosensor LysG-PlysE and an evolutionary actuator composed of a mutagenesis gene and a fluorescent protein gene. After successive rounds of evolution, mutant strains with significantly increased 4-HIL production and growth performance are obtained. The maximum 4-HIL titer is 152.19 mM, 28.4% higher than the starting strain 1.14.11.45 L-isoleucine 4-hydroxylase synthesis in a genome-edited recombinant strain Escherichia coli BL21(DE3) DELTAsucABDeltaaceAK/pET-28a(+)-ido (2DELTA-ido), the bioconversion ratio of L-Ile to 4-HIL is enhanced by about 15% compared to Escherichia coli BL21(DE3)/pET-28a(+)-ido [BL21(DE3)-ido] 1.14.11.45 L-isoleucine 4-hydroxylase synthesis recombinant Escherichia coli expressing mutant N126H/T130K or wild-type synthesizes 66.50 mM and 26.09 mM 4-hydroxyisoleucine, respectively, in 24 h 1.14.11.53 mRNA N6-methyladenine demethylase synthesis visible-light-induced oxidation of m6ARNA is performed in live cells under atmospheric O2 by applying the metabolite flavin mononucleotide (FMN) as an efficient photosensitizer 1.14.11.55 ectoine hydroxylase synthesis efficient conversion of ectoine to hydroxyectoine in Halomonas elongata by heterologous expression of the ectoine hydroxylase gene, thpD, from Streptomyces anulatus under cotrol of the Hansenula elongata ectA promoter 1.14.11.55 ectoine hydroxylase synthesis expression of 5-hydroxyectoine biosynthesis enzymes EctA, EctB, EctC, and EctD in Hansenula polymorpha. 5-Hydroxyectoine synthesis in gram per liter scale (2.8 g/l culture supernatant, 365 mol/g dry cell weight) can be achieved, with almost 100% conversion of ectoine to 5-hydroxyectoine without necessity of high salinity 1.14.11.55 ectoine hydroxylase synthesis an Escherichia coli cell factory expressing the Pseudomonas stutzeri ectD gene from a synthetic promoter imports homoectoine via the ProU and ProP compatible solute transporters, hydroxylates it, and secretes the formed trans-5-hydroxyhomoectoine, independent from all currently known mechanosensitive channels, into the growth medium 1.14.11.55 ectoine hydroxylase synthesis effect of medium formulation (i.e., yeast extract (YE) medium and high yeast extract (HYE) medium) on hydroxyectoine production. Hydroxyectoine production is elevated when the high yeast extract medium is utilized. Hydroxyectoine production increases to 2.9 g/l when 50 mM of 2-oxoglutarate and 1 mM of iron are added to the high yeast extract medium 1.14.11.55 ectoine hydroxylase synthesis in an optimized medium containing 100 g/l NaCl in a 500-ml flask, the double mutant lacking EctD and ectoine hydrolaseDoeA synthesizes 3.13 g/l ectoine after 30 h cultivation. Mutants additionally lacking key Na+/H+ antiporter Mrp can synthesize around 7 g/l or 500 mg/(g DCW) in the medium containing lower concentration of NaCl. During a fed-batch fermentation process with 60 g/l NaCl stress, a maximum 10.5 g/l ectoine is accumulated by the Mrp-deficient strain,with a specific production of 765 mg/(g DCW) and a yield of 0.21 g/g monosodium glutamate 1.14.11.56 L-proline cis-4-hydroxylase synthesis coexpression of L-proline cis-4-hydroxylase and N-acetyltransferase Mpr1 from Saccharomyces cerevisiae converting cis-4-hydroxy-L-proline into N-acetyl cis-4-hydroxy-L-proline in Escherichia coli. M9 medium containing L-proline produces more N-acetyl cis-4-hydroxy-L-proline than LB medium containing L-proline. The addition of NaCl and L-ascorbate results in a 2fold increase in N-acetyl cis-4-hydroxy-L-proline production in the L-proline-containing M9 medium 1.14.12.10 benzoate 1,2-dioxygenase synthesis production of (R,S)-1,2-dihydroxycatechol, which is used as a polymerisation monomer 1.14.12.10 benzoate 1,2-dioxygenase synthesis Synthesis of useful chiral chemical products: biotransformation of benzoate to cis-diols 1.14.12.11 toluene dioxygenase synthesis screening of substituted arenes containing remote chiral centers as substrates, enantiomers are indiscriminately processed to diastereomeric pairs. Some of these new metabolites are useful as synthons for morphine synthesis 1.14.12.11 toluene dioxygenase synthesis a series of cis-dihydrodiol metabolites is obtained by bacterial biotransformation of the corresponding 1,4-disubstituted benzene substrates using Pseudomonas putida UV4, a source of toluene dioxygenase 1.14.12.11 toluene dioxygenase synthesis Pseudomonas putida KT2442 (pSPM01) harboring TDO genes can effectively biotransform a wide-range of aromatic substrates into their cis-diols 1.14.12.11 toluene dioxygenase synthesis Escherichia coli BW25113 DELTAgldA strain harboring pBAD18-TDO system is a suitable platform for the production of the valuable compound cis-1,2-dihydrocatechol at gram scale. The DHC scaffold finds extensive application in the production of a variety of fine chemicals and bioactive compounds 1.14.12.12 naphthalene 1,2-dioxygenase synthesis oxidation of naphthalene to produce optically pure (+)-cis-(1R,2S)-1,2-naphthalene dihydrodiol. Increase of productivity of the biotransformation by using resting cells. The biocatalyst is recycled for at least four runs in both suspended and immobilized form. Suspended resting cells retain their activity for at least four runs for 6 h recycle, but the stability is not retained for more than two runs for 12 h recycle. The stability of the 12 h recycle is improved by immobilization 1.14.12.18 biphenyl 2,3-dioxygenase synthesis cis-2',3'-dihydrodiol production on flavone B-ring by biphenyl dioxygenase from Pseudomonas pseudoalcaligenes KF707 expressed in Escherichia coli 1.14.12.26 chlorobenzene dioxygenase synthesis asymmetric dihydroxylation of non-natural cinnamonitrile to trans-3-[(5S,6R)-5,6-dihydroxycyclohexa-1,3-dienyl]-acrylonitrile by chlorobenzene dioxygenase in recombinant Escherichia coli JM101 (pTEZ30). Recombinant hosts with a strong expression system, such as Escherichia coli JM101 (pTEZ30), can be used to produce dioxygenases efficiently to performcis-dihydroxylations up to technical scales. The cells should be maintained in ametabolically active state duringthe biotransformation in order to increase and maintain the volumetric productivity for long-term reactions 1.14.12.26 chlorobenzene dioxygenase synthesis the enzyme is cloned under the strict control of the Palk promoter of Pseudomonas putida GPo1 and expressed in Escherichia coli JM101 cells carrying the resulting plasmid pTEZ30. Cells are used for regio- and stereoselective dihydroxylation of benzonitrile, ortho-, meta- and para-substituted benzonitriles, as well as cinnamonitrile and benzyl cyanide. cis-Dihydroxylations are at the 1,2 positions and the products have 42.9 to 97.1% enantiomeric excess 1.14.13.7 phenol 2-monooxygenase (NADPH) synthesis the multicomponent phenol hydroxylases can be used as biocatalysts for producing dyes, e.g. indigo, and hydroxyindoles such as 7-hydroxyindole from indole and its derivatives, overview, multicomponent phenol hydroxylases may serve as potential agents for organic syntheses as well as bioremediation 1.14.13.16 cyclopentanone monooxygenase synthesis the enzyme can be used for accessing tetrahydrofuran-based natural products by stereoselective Baeyer-Villiger biooxidation, in particular for the formation of chiral lactones, natural compound synthesis, completion of the formal total synthesis of (+)-showdomycin and establishing of the absolute configuration of biooxidation product as (1S,6S)-3,9-dioxabicyclo[4.2.1]non-7-en-4-one, overview 1.14.13.16 cyclopentanone monooxygenase synthesis the Escherichia coli overexpression systems of Baeyer–Villiger monooxygenases, cyclohexanone monooxygenase and cyclopentanone monooxygenase and their mutants derived from directed evolution are used as catalysts in oxidations of six 4-substituted cyclohexanones. Several substrates that give negative results with growing cells, afford excellent conversions in the transformations under non-growing conditions 1.14.13.16 cyclopentanone monooxygenase synthesis Baeyer-Villiger biooxidation of 4-methylcyclohexanone to 5-methyloxepane-2-one catalysed by recombinant Escherichia coli overexpressing cyclopentanone monooxygenase encapsulated in polyelectrolyte complex capsules. Viability of encapsulated cells decreases with increasing substrate concentration from 99 to 83%, while substrate conversions decrease from 100 to 6%. Storage stabilization of encapsulated cells is observed by increased substrate conversion form 68 to 96% 1.14.13.22 cyclohexanone monooxygenase synthesis production of optically pure sulfoxides by biotransformation in whole cell systems of several sulfides, dithianes and dithiolanes 1.14.13.22 cyclohexanone monooxygenase synthesis chiral catalysis for the laboratory-scale transformation of racemic and prochiral ketones to chiral lactones and organic sulfur compounds to optically active sulfoxides, as a whole cell preparation and as an isolated immobilized enzyme 1.14.13.22 cyclohexanone monooxygenase synthesis manufacturing of fine chemicals 1.14.13.22 cyclohexanone monooxygenase synthesis the enzyme in crude extract can be used for synthesis of enantiopure lactones, development of a simple and easy to handle method including a coupled NADPG regeneration system 1.14.13.22 cyclohexanone monooxygenase synthesis model enantioselective Baeyer-Villiger biooxidations of rac-bicyclo[3.2.0]hept-2-en-6-one to corresponding lactones (1R,5S)-3-oxabicyclo-[3.3.0]oct-6-en-3-one and (1S,5R)-2-oxabicyclo-[3.3.0]oct-6-en-3-one as important chiral synthons for the synthesis of bioactive compounds. Reactions are performed in the minireactor equipped with a column packed with encapsulated recombinant cells Escherichia coli overexpressing cyclohexanone monooxygenase. The cells are encapsulated in polyelectrolyte complex capsules formed by reaction of oppositely charged polymers. Encapsulated cells tested in inireactor exhibit high operational stability with 4 complete substrate conversions to products and 6 conversions above 80% within 14 repeated consecutive biooxidation tests. Encapsulated cells show high enzyme stability during 91 days of storage with substrate conversions above 80% up to 60 days of storage 1.14.13.22 cyclohexanone monooxygenase synthesis a fusion protein that can convert cyclohexanol to epsilon-caprolactone in vitro is created. The alcohol dehydrogenase(Mesotoga infera)/cyclohexanone monooxygenase(Thermocrispum municipale) fusion construct is expressed in Escherichia coli cells. By circumventing substrate and product inhibition, a more than 99% conversion of 200 mM cyclohexanol can be achieved in 24 h, with more than 13000 turnovers per fusion enzyme molecule 1.14.13.22 cyclohexanone monooxygenase synthesis cyclohexanone monooxygenase (CHMO) from Acinetobacter sp. NCIMB 9871 is a prototype Baeyer-Villiger monooxygenases (BVMO). The enzyme shows an impressive substrate scope with a high chemo-, regio- and/or enantioselectivity. BVMO reactions are often difficult, if not impossible to achieve by chemical approaches and this makes these enzymes highly desired candidates for industrial applications. The industrial use is hampered by several factors related to the lack of stability of these biocatalysts. An easy computational method is used for the prediction of stabilizing disulfide bonds in the cyclohexanone monooxygenase-scaffold. The most promising predicted disulfide pairs are created and biochemically characterized. The T415C single point variant is the most stable variant with a 30fold increased long-term stability (33% residual activity after 24 h incubation at 25°C) 1.14.13.22 cyclohexanone monooxygenase synthesis synthesis od lactones from cycloalkanes. A heterologous pathway comprising enzymes with compatible kinetics is designed in Pseudomonas taiwanensis VLB120 enabling in-vivo cascade for synthesizing lactones from cycloalkanes. The respective pathway includes cytochrome P450 monooxygenase (CHX), cyclohexanol dehydrogenase (CDH), and cyclohexanone monooxygenase (CHXON) from Acidovorax sp. CHX100. Resting cells of the recombinant host Pseudomonas taiwanensis VLB120 convert cyclohexane, cyclohexanol, and cyclohexanone to epsilon-caprolactone at 22, 80-100, and 170 U/g cell dry weight, respectively. Cyclohexane (5 mM) is completely converted with a selectivity of 65% for epsilon-caprolactone formation in 2 h without accumulation of intermediate products 1.14.13.22 cyclohexanone monooxygenase synthesis the enzyme catalyzes the Baeyer-Villiger oxidation of 2-butanone, yielding ethyl acetate and methyl propanoate as products. Methyl propanoate is of industrial interest as a precursor of acrylic plastic. Various residues near the substrate and NADP+ binding sites are subjected to saturation mutagenesis to enhance both the activity on 2-butanone and the regioselectivity toward methyl propanoate 1.14.13.22 cyclohexanone monooxygenase synthesis analysis of conversion kinetics for isolated enzyme, suspended whole cells, and biofilms, the latter two based on recombinant CHMO-containing P. taiwanensis VLB120. Biofilms show less favorable values for KS (9.3fold higher) and kcat (4.8fold lower) compared with corresponding KM and kcat values of isolated CHMO, but a favorable KI for cyclohexanone (5.3fold higher). Suspended cells show only 1.8fold higher KS, but 1.3- and 4.2fold higher kcat and KI values than isolated CHMO 1.14.13.22 cyclohexanone monooxygenase synthesis extracellular production of engineered CHMO by Pichia pastoris. The recombinant CHMO shows a higher flavin occupation rate than that produced by Escherichia coli, accompanied by a 3.2fold increase in catalytic efficiency. At a cell density of 150 g/l cell dry weight, a recombinant CHMO production rate of 1,700 U/l is achieved. By directly employing the pH adjusted supernatant as a biocatalyst, 10 g/l of pyrmetazole is almost completely transformed into the corresponding (S)-sulfoxide, with > 99% enantiomeric excess 1.14.13.22 cyclohexanone monooxygenase synthesis synthesis of lactones from cycloalkanes in Pseudomonas taiwanensis VLB120. The pathway includes cytochrome P450 monooxygenase, cyclohexanol dehydrogenase, and cyclohexanone monooxygenase from Acidovorax sp. CHX100. Recombinant P. taiwanensis converts cyclohexane, cyclohexanol, and cyclohexanone to epsilon-caprolactone at 22, 80-100, and 170 U/gCDW, respectively. Cyclohexane (5 mM) is completely converted with a selectivity of 65% for epsilon-caprolactone formation in 2 hr without accumulation of intermediate products. Analogous lactones can be obtained from cyclooctane and cyclodecane 1.14.13.22 cyclohexanone monooxygenase synthesis synthesis of trimethyl-epsilon-caprolactone using cyclohexanone monooxygenase immobilized on Mana-agarose and a highly active glucose dehydrogenase for cofactor regeneration. A biocatalyst yield of 37.3 g trimethyl-epsilon-caprolactone per g of CHMO and 474.2 g trimethyl-epsilon-caprolactone per g of glucose dehydrogenase are obtained 1.14.13.25 methane monooxygenase (soluble) synthesis cofactor-independent oxygenation reactions catalyzed by soluble methane monooxygenase at the surface of a modified gold electrode. The electrochemically driven enzyme shows the same catalytic activity and regulation by the protein component B (MMOB) as the natural NAD(P)H-driven reaction and may have the potential for development into economic NAD(P)H-independent oxygenation catalyst 1.14.13.25 methane monooxygenase (soluble) synthesis the enzyme is a base or scaffold for design of small molecule catalysts for use in large scale methanol synthesis 1.14.13.B28 monooxygenase CYP119A2 synthesis coexpression of 5-aminolevulinic acid synthase (ALAS) from Rhodobacter capsulatus improves the heterologous production of CYP119. Coexpression of ALAS increases the amount of heterologous CYP119 isolated and the ratio of its holo form. The ratio of holo-CYP119 resulting from the coexpression of ALAS in Escherichia coli is 99%, whereas that from cells expressing CYP119 exclusively is 66% 1.14.13.38 anhydrotetracycline 6-monooxygenase synthesis biosynthesis of tetracycline from anhydrotetracycline in Saccharomyces cerevisiae heterologously expressing the anhydrotetracycline hydroxylase OxyS, the dehydrotetracycline reductase CtcM, and the F420 reductase FNO from three bacterial hosts. This biosynthesis of tetracycline is enabled by OxyS performing just one hydroxylation step in S. cerevisiae 1.14.13.44 2-hydroxybiphenyl 3-monooxygenase synthesis production of 3-phenylcatechol, which is starting material for the synthesis of pharmaceutical compounds and artificial supramolecular systems 1.14.13.64 4-hydroxybenzoate 1-hydroxylase synthesis construction of a novel artificial pathway for arbutin biosynthesis in Escherichia colid. De novo biosynthesis of arbutin from simple carbon sources is established and a generalizable strategy for the biosynthesis of shikimate pathway derived chemicals is provided. Arbutin is a hydroquinone glucoside compound existing in various plants. It is widely used in pharmaceutical and cosmetic industries owing to its well-known skin-lightening property as well as anti-oxidant, anti-microbial, and anti-inflammatory activities. A 4-hydroxybenzoate 1-hydroxylase gene from Candida parapsilosis CBS604 and a glucosyltransferase (arbutin synthase) gene from Rauvolfia serpentina are introduced into Escherichia coli lead to the production of 54.71 mg/l of arbutin from glucose. Further redirection of carbon flux into arbutin biosynthesis pathway by enhancing shikimate pathway genes enables production of 3.29 g/l arbutin, which is a 60-fold increase compared with the initial strain. Final optimization of glucose concentration added in the culture medium is able to further improve the titer of arbutin to 4.19 g/l in shake flasks experiments, which is around 77-fold higher than that of initial strain 1.14.13.64 4-hydroxybenzoate 1-hydroxylase synthesis an artificial pathway is established in Escherichia coli for high-level production of arbutin from simple carbon sources in Escherichia coli for high-level production of arbutin from simple carbon sources. Introduction of the genes for 4-hydroxybenzoate 1-hydroxylase from Candida parapsilosis CBS604 and hydroquinone glucosyltransferase from Rauvolfia serpentina into Escherichia coli leads to the production of 54.71 mg/l of arbutin from glucose. Further redirection of carbon flux into arbutin biosynthesis pathway by enhancing shikimate pathway genes enables production of 3.29 g/l arbutin. Final optimization of glucose concentration added in the culture medium is able to further improve the titer of arbutin to 4.19 g/l in shake flasks experiments 1.14.13.69 alkene monooxygenase synthesis alkene MOs are of interest for their potential roles in industrial biocatalysis, most notably for the stereoselective synthesis of epoxides. Development of high-activity recombinant biocatalysts for alkene oxidation 1.14.13.81 magnesium-protoporphyrin IX monomethyl ester (oxidative) cyclase synthesis the Rhodobacter capsulatus kanamycin-resistant transposon bchE mutant strain DB575 is used for production of Mg-protoporphyrin IX monomethylester, method development, overview 1.14.13.84 4-hydroxyacetophenone monooxygenase synthesis the enzyme can be useful in the highly enantioselective in the synthesis of chiral phenyl and benzyl sulfoxides 1.14.13.84 4-hydroxyacetophenone monooxygenase synthesis usage of the enzyme as biocatalyst for synthesis of chiral 3-alkyl-3,4-dihydroisocoumarins, evaluation, overview 1.14.13.92 phenylacetone monooxygenase synthesis the enzyme has catalytic potential as a biocatalyst 1.14.13.92 phenylacetone monooxygenase synthesis the thermally stable phenylacetone monooxygenase and engineered mutants can be used as a practical catalysts for enantioselective Baeyer-Villiger oxidations of several ketones on a preparative scale under in vitro conditions, overview 1.14.13.92 phenylacetone monooxygenase synthesis the enzyme is useful for kinetic resolution of a set of racemic substituted 3-phenylbutan-2-ones and synthesis of enantiopure compounds, overview 1.14.13.92 phenylacetone monooxygenase synthesis usage of the enzyme for asymmetric sulfooxidation of thioanisole and of racemic 2-phenylpropionaldehyde in a Baeyer-Villiger oxidation reaction 1.14.13.92 phenylacetone monooxygenase synthesis phenylacetone monooxygenase (PAMO) is an exceptionally robust Baeyer-Villiger monooxygenase, which makes it ideal for potential industrial applications, usage as an active catalyst for the Baeyer-Villiger conversion of cyclohexanone to caprolactone, which is important as monomer in polymer science 1.14.13.92 phenylacetone monooxygenase synthesis phenylacetone monooxygenase (PAMO) is the most stable and thermo-tolerant member of the Baeyer-Villiger monooxygenase family, and the engineered enzyme is useful for the synthesis of industrially relevant compounds, in particular, in the biotransformation of long-chain aliphatic oils into potential biodiesels 1.14.13.92 phenylacetone monooxygenase synthesis phenylacetone monooxygenase (PAMO) catalyzes oxidation of ketones with molecular oxygen and NADPH with the formation of esters. PAMO demonstrates high catalytic constants in the oxidation of benzylacetone and other structurally similar aromatic ketones and can be used in the synthesis of flavoring substances, since the ester formed by benzylacetone oxidation has a pronounced fruity smell 1.14.13.92 phenylacetone monooxygenase synthesis the enzyme is an ideal candidate for the synthesis of industrially relevant ester or lactone compounds. But its limited substrate scope has largely limited its industrial applications. The engineered mutant quadruple enzyme variant P253F/G254A/R258M/L443F exhibits significantly improved activity towards 2-octanone 1.14.13.92 phenylacetone monooxygenase synthesis the H2O2-resistant enzyme variants are robust biocatalysts for synthetic applications 1.14.13.114 6-hydroxynicotinate 3-monooxygenase synthesis the isolated enzyme is used for the synthesis of 2,5-dihydroxypyridine, a precursor for the chemical synthesis of 5-aminolevulinic acid, which is applied as a plant growth hormone, a herbicide and in cancer therapy 1.14.13.131 dissimilatory dimethyl sulfide monooxygenase synthesis upon expression in Escherichia coli, DmoA and DmoB subunits are able to generate product, albeit at a lower turnover than the natively expressed enzyme. No static protein-protein interactions are observed under the conditions tested between the two subunits 1.14.13.154 erythromycin 12-hydroxylase synthesis systematically modulating the enzyme amounts of EryK and EryG by integrating additional eryK and eryG copies into the industrial strain Saccharopolyspora erythraea HL3168 E3 significantly enhances the process of biotransformation from erythromycin-D to erythromycin-A, nearly completely eliminates the by-products erythromycin-B and erythromycin-C, and efficiently improves erythromycin-A production and purity at the fermentation stage. In conjunction with other traditional and genetic ways to continuously evaluate the erythromycin-A production system 1.14.13.163 6-hydroxy-3-succinoylpyridine 3-monooxygenase synthesis development of an efficient process to transform HSP into 2,5-dihydroxypyridine (2,5-DHP) with heterologously expressed HSP hydroxylase and NADH-regenerating system, because 2,5-DHP, the product of the reaction catalyzed by HSP hydroxylase, is a valuable precursor for the chemical synthesis of 5-aminolevulinic acid, which is applied as a plant growth hormone, a herbicide and in cancer therapy 1.14.13.163 6-hydroxy-3-succinoylpyridine 3-monooxygenase synthesis enzyme HSPHZZ can be used for the enzymatic production of 2,5-dihydroxypyridine in biotechnology applications. 85.3 mg/l 2,5-dihydroxypyridine is produced in 40 min with a conversion of 74.9% at 30°C, pH 8.5, 1.0 mM substrate concentration, and 0.001 mM enzyme concentration 1.14.13.163 6-hydroxy-3-succinoylpyridine 3-monooxygenase synthesis production of 2,5-dihydroxypyridine from 6-hydroxy-3-succinoylpyridine in the presence of NADH and FAD using nicotine hydroxylase covalently immobilized on Immobead 150. At a protein loading of 15 mg/g, ImmHSPHZZ converts 93.6% of 6-hydroxy-3-succinoylpyridine to 2,5-dihydroxypyridine in 6 h. The optimal concentrations of ImmHSPHZZ and substrate are 30 mg/l and 0.75 mM, respectively. Under optimal conditions, 94.5 mg/l of 2,5-dihydroxypyridine is produced after 30 min with 85.4% conversion 1.14.13.172 salicylate 5-hydroxylase synthesis biosynthetic pathway for maleate production in engineered Escherichia coli. Expression of salicylate 5-hydroxylase, introduction of salicylate biosynthetic pathway and gentisate ring cleavage pathway allow the synthesis of maleate from glycerol. Further optimizations boost maleate titer to 2.4 g/l in shake flask experiments. Subsequent scale-up biosynthesis of maleate in a 3-l bioreactor under fed-batch culture conditions enables the production of 14.5 g/l of maleate 1.14.13.180 aklavinone 12-hydroxylase synthesis biosynthesis of carminic acid from glucose in engineered Escherichia coli. Expression of an optimized type II polyketide synthase machinery from Photorhabdus luminescens enables a high-level production of flavokermesic acid upon coexpression of the cyclases ZhuI and ZhuJ from Streptomyces sp. R1128. Mutants of aklavinone 12-hydroxylase (DnrF) from Streptomyces peucetius and C-glucosyltransferase (GtCGT) from Gentiana triflora with 2-5fold enhanced conversion efficiencies can perform hydroxylation and C-glucosylation of flavokermesic acid, respectively, resulting in 0.63 mg/l of carminic acid production from glucose 1.14.13.225 F-actin monooxygenase synthesis protocol for obtaining high levels of recombinant protein for the redox only portion of Mical. Cold adapted chaperonins alone do not work well for expressing the Mical redox domain, while the use and removal of solubility tags destabilizes. Low-temperature expression, chaperonins and no solubility tag lead to enhanced expression 1.14.13.234 5a,11a-dehydrotetracycline 5-monooxygenase synthesis biosynthesis of tetracycline from anhydrotetracycline in Saccharomyces cerevisiae heterologously expressing the anhydrotetracycline hydroxylase OxyS, the dehydrotetracycline reductase CtcM, and the F420 reductase FNO from three bacterial hosts. This biosynthesis of tetracycline is enabled by OxyS performing just one hydroxylation step in S. cerevisiae 1.14.13.236 toluene 4-monooxygenase synthesis by manipulation of vectors and gene inserts to eliminate adventitious catalytic turnover of NADH, up to 60fold increase in the volumetric yield of T4moH activity can be obtained from recombinant fermentations in Escherichia coli BL21(DE3) 1.14.13.236 toluene 4-monooxygenase synthesis conversion of benzene to phenol and naphthalene to 2-naphthol in a two-phase system using whole cells expressing wild-type toluene 4-monooxygenase and the alpha subunit variant TmoA I100A. With mutant I100A, the solubility of naphthalene is enhanced and the toxicity of the naphthols is prevented by the use of a water/dioctyl phthalate (80:20, vol%) system More than 99%2-naphthol is extracted to the dioctyl phthalate phase, dihydroxynaphthalene formation is prevented, 92% 2-naphthol is formed, and 12% naphthalene is converted. Using 50 vol% dioctyl phthalate, and wild-type T4MO, a 51% conversion of benzene is obtained and phenol is produced at a purity of 97% 1.14.13.236 toluene 4-monooxygenase synthesis development of a two-phase (dioctyl phthalate/aqueous medium) culture system to obtain maximal indirubin from indole. A 50% (v/v) dioctyl phthalate two-phase system using tryptophan medium containing 3 mM cysteine, 5 mM indole, and 1 mM isatin yields 102.4 mg/l of indirubin with no conversion of indole to indigo 1.14.13.236 toluene 4-monooxygenase synthesis improved synthesis of hydroxytyrosol in whole-cell biotransformation. An overall concentration of 133 mg/l hydroxytyrosol, corresponding to a volumetric productivity of 54 mg/l/h and a yield of 48% is achieved by a batch mode using 2 mM substrate. The use of beads conjugated with phenylboronic acid residues for adsorbing the product from the biotransformation bulk leads to 2fold purification resulting in 84% purity with 70% recovery yield 1.14.13.236 toluene 4-monooxygenase synthesis synthesis of hydroxytyrosol, a phenol present in olives, via double hydroxylation of 2-phenylethanol employing toluene monooxygenases as biocatalysts 1.14.13.236 toluene 4-monooxygenase synthesis synthesis of hydroxytyrosol, a phenol present in olives, via double hydroxylation of 2-phenylethanol employing toluene monooxygenases as biocatalysts. Mutant TmoA S395C shows a 15fold increase in 2-phenylethanol hydroxylation rate 1.14.13.236 toluene 4-monooxygenase synthesis transformation of 1 mM fluorobenzene by whole cells gives a 52% yield of 4-fluorocatechol as a single product. The yield is improved 1.6fold by adding 10 mM ascorbic acid to the biotransformations 1.14.13.244 phenol 2-monooxygenase (NADH) synthesis production of hydroxytyrosol by conversion of tyrosol using phenol hydroxylase 1.14.14.1 unspecific monooxygenase synthesis production of 6-beta-hydroxy-methyl-simvastatin, simvastatin and derivatives belong to the family of HMG-CoA reductase inhibitors, which are potent cholesterol-lowering therapeutic agents 1.14.14.1 unspecific monooxygenase synthesis the enzyme is intersting for regioselective production of compounds 1.14.14.1 unspecific monooxygenase synthesis P450-BM3 enzyme mutants are ueful for CH-activating oxidative hydroxylation of readily available 1-tetralone derivatives with high degrees of regio- and stereoselectivity, e.g. oxidative hydroxylation of indane and tetralin, an approach which is currently not possible using chiral synthetic CH-activating transition metal catalysts 1.14.14.1 unspecific monooxygenase synthesis coexpression of P450 reductase with Sam5 in Escherichia coli enhances hydroxylation by approximately from 34 to 50%, depending on the flavonoid used. Optimization of the production of bioactive flavonoids, 8-hydroxyluteolin and 3'-hydroxydaidzein, in this system gives 88 mg/l of 8-hydroxyluteolin and 75 mg/l of 3'-hydroxydaidzein 1.14.14.1 unspecific monooxygenase synthesis electrochemical conversion of p-xylene to 2,5-dimethylphenol by mutant R47S/Y51W/ I401M and comparison of mediators cobalt sepulchrate and pentamethylcyclopentadienyl rhodium 2,2'-bipyridin 1.14.14.1 unspecific monooxygenase synthesis BM3 is coimmobilized with glucose dehydrogenase (type IV, from B. megaterium), as fusion proteins with the polycationic binding module Zbasic2, on anionic sulfopropyl?activated carrier. Immobilization via Zbasic2 enables each enzyme to be loaded in controllable amount. Using lauric acid as a representative P450 substrate, complete hydroxylation at low catalyst loading (below 0.1 mol%) and efficient electron coupling (74%), inside of the catalyst particle, to the regeneration of NADPH from glucose (27 cycles) is achieved. The immobilized P450 BM3 shows a total turnover number of about 18000 per second 1.14.14.3 bacterial luciferase synthesis upon expression in Bacillus subtilis cells, luciferase is substantially more thermostable than in Escherichia coli. Thermal inactivation in Bacillus subtilis at 48.5#°C behaves as a first-order reaction. In Escherichia coli, the first order rate constant of the thermal inactivation exceeds that observed in B. subtilis cells 2.9 times. In dnaK-negative strains of Bacillus subtilis, both the rates of thermal inactivation and the efficiency of refolding are similar to that observed in wild-type strains 1.14.14.3 bacterial luciferase synthesis upon expression in Bacillus subtilis cells, luciferase is substantially more thermostable than in Escherichia coli. Thermal inactivation in the cells of Escherichia coli and Bacillus subtilis may be described by first and third order kinetics, respectively. In dnaK-negative strains of Bacillus subtilis, both the rates of thermal inactivation and the efficiency of refolding are similar to that observed in wild-type strains 1.14.14.9 4-hydroxyphenylacetate 3-monooxygenase synthesis enzyme completely transforms 4-substituted halophenols to 4-halocatechols at 2 mM within a 1-2 h period. An increase in 4-halophenol concentration to 4.8 mM results in a 2.5-20fold decrease in biotransformation efficiency depending on the substrate tested. Organic solvent extraction of the 4-halocatechol products followed by column chromatography results in the production of purified products with a final yield of between 33% and 38% 1.14.14.9 4-hydroxyphenylacetate 3-monooxygenase synthesis investigation of a flask-scale production of caffeic acid from p-coumaric acid as the model reaction for HpaBC-catalyzed synthesis of hydroxycinnamic acids. Since the initial concentrations of the substrate p-coumaric acid higher than 40 mM markedly inhibits its 4-hydroxyphenylacetate 3-hydroxylases-catalyzed oxidation, the reaction is carried out by repeatedly adding 20 mM of this substrate to the reaction mixture. Furthermore, by using the whole-cell catalyst in the presence of glycerol, the experimental setup achieves the high-yield production of caffeic acid, i.e., 56.6 mM (10.2 g/l) within 24 h. These catalytic activities of 4-hydroxyphenylacetate 3-hydroxylases will provide an easy and environment-friendly synthetic approach to hydroxycinnamic acids 1.14.14.9 4-hydroxyphenylacetate 3-monooxygenase synthesis enzyme shows relatively broad substrate specificity, which allows the conversion of a number of non-natural phenolic compounds, into the corresponding catechols. The reaction can be performed based on formate as the electron donor and the organometallic complex [Rh(bpy)-Cp*(H2O)]2+ (Cp*: 1,2,3,4,5-pentamethylcyclopentadiene, bpy: 2,2'-bipyridyl) as the catalyst for FAD reduction 1.14.14.9 4-hydroxyphenylacetate 3-monooxygenase synthesis in vivo production of ortho-hydroxylated flavonoids by recombinant Escherichia coli. When HpaC is linked with an S-Tag on the C terminus, the enzyme activity is significantly affected. The optimal culture conditions are a substrate concentration of 80 mg/l, an induction temperature of 28°C, an M9 medium, and a substrate delay time of 6 h after IPTG induction. The efficiency of eriodictyol conversion from recombinant strains fed naringin is up to 57.67. Highest conversion efficiencies for production of catechin and caffeate are 35.2 % and 32.93%, respectively 1.14.14.9 4-hydroxyphenylacetate 3-monooxygenase synthesis overexpression of the HpaB and HpaC genes in Saccharomyces cerevisiae achieves hydroxytyrosol titers of 1.15 mg/l and 4.6 mg/l in a minimal medium in which either 1 mM tyrosine or 1 mM tyrosol are respectively added 1.14.14.11 styrene monooxygenase synthesis development of a highly diastereo- and enantio-selective enzymatic synthesis of glycidol derivatives with contiguous stereogenic centers 1.14.14.11 styrene monooxygenase synthesis engineering of a stable whole-cell biocatalyst capable of (S)-2-phenyloxirane formation for continuous two-liquid-phase applications 1.14.14.11 styrene monooxygenase synthesis production of enantiopure styrene oxide by recombinant Escherichia coli. Two-liquid phase fed-batch process is established for the production of (S)-styrene oxide with hexadecane as an apolar carrier solvent and a nutrient feed consisting of glucose, magnesium sulfate, and yeast extract. Production of 11 g of enantiopure (S)-styrene oxide per liter liquid volume in 10 h 1.14.14.11 styrene monooxygenase synthesis production of optically pure (S)-styrene oxide, an important building block in organic synthesis. Recombinant Escherichia coli producing styrene monooxygenase catalyzes the formation of (S)-2-phenyloxirane from inexpensive styrene with an excellent enantiomeric excess of more than 99% at rates up to 180 U/g (dry weight) of cells 1.14.14.11 styrene monooxygenase synthesis synthesis of (S)-styrene oxide. Investigation of factors influencing biocatalytic efficiency of oxygenase-based whole-cell biocatalysts under process conditions 1.14.14.11 styrene monooxygenase synthesis synthesis of various chiral aryloxides, pilot-scale production of (S)-2-phenyloxirane 1.14.14.11 styrene monooxygenase synthesis the enzyme converts aryl ethenyl compounds to the corresponding epoxides in optically pure forms and with good yields 1.14.14.11 styrene monooxygenase synthesis construcution of fusion proteins that join the C-terminus of the epoxidase StyA to the N-terminus of FAD reductase StyB through a linker peptide and application in the synthesis of of a broad range of substituted indoles to indigoid chromophores. The fusion proteins are self-regulated and couple efficiently NADH oxidation to styrene epoxidation 1.14.14.11 styrene monooxygenase synthesis in an organic solvent-water biphasic reaction system, recombinant enzyme enantioselectively converts linear terminal alkenes to their corresponding (S)-epoxyalkanes using glucose and molecular oxygen. When 1-heptene and 6-chloro-1-hexene are used as substrates (400 mM) under optimized conditions, 88.3mM (S)-1,2-epoxyheptane and 246.5mM (S)-1,2-epoxy-6-chlorohexane, respectively, accumulate in the organic phase with good enantiomeric excess (84.2% and 95.5%, respectively) 1.14.14.11 styrene monooxygenase synthesis overexpression of StyA and StyB significantly enhances the indigo production, reaching 52.13 mg/L after 24 h. Overexpression of oxide isomerase gene styC does not increase indigo yield 1.14.14.17 squalene monooxygenase synthesis simultaneous overexpression of squalene epoxidase and 3-hydroxy-3-methylglutaryl coenzyme A enhances individual ganoderic acid production. The overexpressing strain produces maximum ganoderic acid-T, ganoderic acid-S, ganoderic acid-Mk, and ganoderic acid-Me contents of 90.4, 35.9, 6.2, and 61.8 microg/100 mg dry weight, respectively 1.14.14.20 phenol 2-monooxygenase (FADH2) synthesis phenol hydroxylase gene cloning from the thermophilic bacteria Geobacillus thermoglucosidasius is used to develop an effective method to convert tyrosol into the high-added-value compound hydroxytyrosol by hydroxylation 1.14.14.28 long-chain alkane monooxygenase synthesis the thermophilic soluble monomeric LadA is an ideal candidate for and biosynthesis of complex molecules 1.14.14.40 phenylalanine N-monooxygenase synthesis biosynthetic pathway for the production of phenylacetonitrile in Escherichia coli utilizing enzymes from the plant glucosinolate-biosynthetic and bacterial aldoxime-nitrile pathways, i.e. introducing the genes encoding cytochrome P450 (CYP) 79A2 and CYP reductase from Arabidopsis thaliana, yielding the E,Z-phenylacetaldoxime-producing transformant and further introducing the aldoxime dehydratase (Oxd) gene from Bacillus sp. strain OxB-1, yielding the phenylacetonitrile-producing transformant. Expression of active CYP79A2 and concentration of biomass are improved by the combination of the autoinduction method, coexpression of groE, encoding the heat shock protein GroEL/GroES, N-terminal truncation of CYP79A2, and optimization of the culture conditions, yielding a more than 60fold concentration of E,Z-phenylacetaldoxime up to 2.9 mM 1.14.14.60 ferruginol monooxygenase synthesis coexpression of mutant F112L or mutant V296L and cytochrome Cyp76AK6 in yeast leads to overall improvements of 24- and 14fold for pisiferic acid and salviol, respectively, compared to wild-type 1.14.14.82 flavonoid 3'-monooxygenase synthesis functional expression in Saccharomyces cerevisiae, to hydroxylate naringenin in whole recombinant cells. In a selective media, 200 mg/l of eriodictyol from naringenin can be produced 1.14.14.83 geraniol 8-hydroxylase synthesis artemisinic acid can be used as a promising elicitor, especially for the production of therapeutically important indole alkaloids 1.14.14.83 geraniol 8-hydroxylase synthesis introduction of genes encoding strictosidine synthase (STR) and geraniol 8-hydroxylase (G10H), into Ophiorrhiza pumila hairy roots. Overexpression of individual G10H significantly improves camptothecin prdouction. Cooverexpression of G10H and STR genes causes a 56% increase on the yields of camptothecin. Camptothecin extracted from different lines shows similar anti-tumor activity 1.14.14.86 ent-kaurene monooxygenase synthesis when KO2 is coexpressed in Escherichia coli engineered to produce ent-kaurene, turnover to entkauren-19-oic acid is observed. In addition, KO2 reacts with ent-isokaurene, and the closely related ent-trachylobane, converting the C4alpha methyl to a mixture of the alcohol and further oxidized carboxylic acid with each substrate 1.14.14.87 2-hydroxyisoflavanone synthase synthesis construction of an in-frame fusion of IFS and cytochrome P450 reductase from rice after removing the membrane binding domain of both enzymes. Expression of the fusion construct in Escherichia coli leads to conversion of naringenin into genistein. Upon optimization of conditions, 60 microM of genistein can be generated from 80 microM of naringenin 1.14.14.103 tabersonine 16-hydroxylase synthesis artemisinic acid can be used as a promising elicitor, especially for the production of therapeutically important indole alkaloids 1.14.14.108 2,5-diketocamphane 1,2-monooxygenase synthesis production of useful chiral synthons for chemoenzymatic synthesis 1.14.14.114 amorpha-4,11-diene 12-monooxygenase synthesis heterologous expression of Artemisia annua amorphadiene synthase and CYP71AV1 in tobacco lead to the accumulation of amorphadiene and artemisinic alcohol, but not artemisinic acid. Additional expression of artemisinic aldehyde DELTA11(13) double-bond reductase DBR2 with or without aldehyde dehydrogenase 1 leads to the additional accumulation dihydroartemisinic alcohol. Results and suggest that amorphane sesquiterpenoid aldehydes are formed, but conditions in the transgenic tobacco cells favour reduction to alcohols rather than oxidation to acids 1.14.14.114 amorpha-4,11-diene 12-monooxygenase synthesis selection of self-pollinated plants for artemisin biosynthesis. Selfpollinated F2 plants selected are grown under optimized growth conditions. The leaves on the main stems exhibit obvious morphological changes, from indented single leaves to odd, pinnately compound leaves. Leaves and flowers form glandular and T-shaped trichomes on their surfaces. The glandular trichome densities increases from the bottom to the top leaves. Leaves, flowers, and young seedlings of F2 plants produce artemisinin. In leaves, the levels of artemisinin increase from the bottom to the top of the plants, showing a positive correlation to the density increase of glandular trichomes. Progeny of selfpollinated plants expresses the amorpha-4, 11-diene synthase and cytochrome P450 monooxygenase 71 AV1genes, which are involved in artemisinin biosynthesis in leaves and flowers 1.14.14.114 amorpha-4,11-diene 12-monooxygenase synthesis Saccharomyces cerevisiae expressing the native Artemisia annua cytochrome P450 monooxygenase (CYP71AV1) and artemisinic aldehyde D11(13) reductase (DBR2) is used as a whole-cell biocatalyst to produce the immediate artemisinin precursor, dihydroartemisinic acid (DHAA) 1.14.14.131 bursehernin 5'-monooxygenase synthesis recombinant coexpression of the genes involved in the podophyllotoxin pathway permits to reconstitute the pathway to (-)-4'-demethylepipodophyllotoxin (the etoposide aglycone), a naturally occurring lignan that is the immediate precursor of etoposide and, unlike podophyllotoxin, a potent topoisomerase inhibitor, for production of the etoposide aglycone in tobacco and circumvent the need for cultivation of mayapple and semisynthetic epimerization and demethylation of podophyllotoxin 1.14.14.132 (-)-4'-demethyl-deoxypodophyllotoxin 4-hydroxylase synthesis recombinant coexpression of the genes involved in the podophyllotoxin pathway permits to reconstitute the pathway to (-)-4'-demethylepipodophyllotoxin (the etoposide aglycone), a naturally occurring lignan that is the immediate precursor of etoposide and, unlike podophyllotoxin, a potent topoisomerase inhibitor, for production of the etoposide aglycone in tobacco and circumvent the need for cultivation of mayapple and semisynthetic epimerization and demethylation of podophyllotoxin 1.14.14.150 costunolide synthase synthesis coexpression of CYP71BL9 with the germacrene A acid 8beta-hydroxylase CYP71BL1 lead to the formation of eupatolide which is found as cystein- and glutathion-adduct 1.14.14.155 3,6-diketocamphane 1,2-monooxygenase synthesis production of useful chiral synthons for chemoenzymatic synthesis 1.14.14.162 flavanone 2-hydroxylase synthesis two-step indirect glycosylation using combinations of flavanone 2-hydroxylase and C-glycosyltransferases, to convert 2-hydroxyflavanone intermediates into the 6C-glucoside flavones isovitexin and isoorientin, and the 8C-glucoside flavones vitexin and orientin. The ratio between 6C and 8C glycosylation depended on the C-glycosyltransferase used. The indirect route resuls in mixtures, similar to what has been reported for in vitro experiments 1.14.14.163 (S)-1-hydroxy-N-methylcanadine 13-hydroxylase synthesis de novo production of noscapine in Saccharomyces cerevisiae, through the reconstruction of a biosynthetic pathway comprising over 30 enzymes from plants, bacteria, mammals, and yeast itself, including 7 plant endoplasmic reticulum (ER)-localized enzymes. Optimization directed to tuning expression of pathway enzymes, host endogenous metabolic pathways, and fermentation conditions led to an over 18,000-fold improvement from initial noscapine titers to 2.2 mg/l 1.14.14.163 (S)-1-hydroxy-N-methylcanadine 13-hydroxylase synthesis reconstitution of the noscapine gene cluster in Saccharomyces cerevisiae to achieve the microbial production of noscapine and related pathway intermediates 1.14.14.164 fraxetin 5-hydroxylase synthesis the therapeutic compound 8-methoxypsoralen is hydroxylated by plants overexpressing CYP82C2 or CYP82C4, forming 5-hydroxy-8-methoxypsoralen 1.14.14.165 indole-3-carbonyl nitrile 4-hydroxylase synthesis the therapeutic compound 8-methoxypsoralen is hydroxylated by plants overexpressing CYP82C2 or CYP82C4, forming 5-hydroxy-8-methoxypsoralen 1.14.14.175 ferruginol synthase synthesis coexpression of mutant F112L or mutant V296L and cytochrome Cyp76AK6 in yeast leads to overall improvements of 24- and 14fold for pisiferic acid and salviol, respectively, compared to wild-type 1.14.14.175 ferruginol synthase synthesis establishment of miltiradiene production in Saccharomyces cerevisiae. Incorporation of CYP76AH1 and phyto-CYP reductase genes lead to heterologous production of ferruginol at 10.5 mg/l 1.14.14.176 taxadiene 5alpha-hydroxylase synthesis engineered biosynthetic pathway for taxane production in Saccharomyces cerevisiae via chromosomal integration of Taxus cuspidata CYP725A4 and CPR genes. When expressed in the Saccharomyces cerevisiae strain, a potential T5alphaol isomer is the predominant product with maximum titers of 24 and 31 mg/l at micro and 1 l bioreactor scale, respectively. Total oxygenated taxane titers are improved to 78 mg/l and the acetylated acetylated T5alpha product is detected with a final titer of 3.7 mg/l 1.14.14.176 taxadiene 5alpha-hydroxylase synthesis expression of taxadiene synthase, taxadiene-5alpha-hydroxylase, and cytochrome P450 reductase in Nicotiana benthamiana. Using a chloroplastic compartmentalized metabolic engineering strategy and enhancement of isoprenoid precursors, the engineered plants can produce taxadiene and taxadiene-5alpha-ol at 56.6 microg/g fresh weight and 1.3 microg /g fresh weight, respectively 1.14.15.1 camphor 5-monooxygenase synthesis camphor hydroxylation in reverse micelles depends on coexistence of enzyme with putidaredoxin, putidaredoxin reductase, and NADH, but not on H2O2 1.14.15.1 camphor 5-monooxygenase synthesis selective enzymatic oxidation of (+)-alpha-pinene to verbenol, verbenone, or myrtenol by enzyme mutants 1.14.15.1 camphor 5-monooxygenase synthesis the enzyme is useful in whole cell biocatalyst systems 1.14.15.1 camphor 5-monooxygenase synthesis fusion of putidaredoxin reductase PdR to the carboxy-terminus of camphor monooxygenase CYP101A1 (P450cam) via a linker peptide and reconstitution of camphor hydroxylase activity with free putidaredoxin enables the production of the almost fully heme-incorporated CYP-FdR fusion that is catalytically active in vivo and in vitro 1.14.15.3 alkane 1-monooxygenase synthesis bacterial alkane hydroxylases have tremendous potential as biocatalysts for the stereo- and regioselective transformation of a wide range of chemically inert unreactive alkanes into valuable reactive chemical precursors 1.14.15.3 alkane 1-monooxygenase synthesis electrocatalytic conversion from alkanes to alcohols mediated by AlkB using a rubredoxin AlkG immobilized screen-printed carbon electrode. As electrons can be transferred from the reduced AlkG to AlkB in a two-phase manner, electrocatalytic conversion from alkanes to alcohols can be applied. Use for conversion of gaseous propane and n-butane to 1-propanol and 1-butanol, respectively 1.14.15.6 cholesterol monooxygenase (side-chain-cleaving) synthesis construction of a fusion protein consisting of cytochrome P450scc (CYP11A1), adrenodoxin and adrenodoxin reductase including 2A peptide from Picornaviridae which is capable of self-cleavage. Introduction to Escherichia coli leads to a high level of expression but no cleavage. In yeast Saccharomyces cerevisiae, the discrete proteins P450scc-2A, adrenodoxin-2A and adrenodoxin reductase are expressed, with a significant proportion present in a fusion adrenodoxin-2A-adrenodoxin reductase. The enzyme system is catalytically active 1.14.15.8 steroid 15beta-monooxygenase synthesis a steroid 15beta-hydroxylating whole-cell solvent tolerant biocatalyst is constructed by expressing the Bacillus megaterium steroid hydroxylase CYP106A2 in the solvent tolerant Pseudomonas putida S12. Testosterone hydroxylation is improved by a factor 16 by co-expressing Fer, a putative Fe-S protein from Bacillus subtilis. The specificity for 15beta-hydroxylation is improved by mutating threonine residue 248 of CYP106A2 into valine. These insights provide the basis for an optimized whole-cell steroid-hydroxylating biocatalyst that can be applied with an organic solvent phase 1.14.15.8 steroid 15beta-monooxygenase synthesis efficient approach towards the preparative scale synthesis of hydroxylated steroid derivatives. Improve CYP106A2-catalyzed steroid hydroxylation towards higher productivity and quantitative product formation. Because substrate transport into the cell limits the whole-cell biotransformation, activity can be increased sixfold by using membrane-free crude cell as biocatalyst 1.14.15.8 steroid 15beta-monooxygenase synthesis an overexpressing Bacillus megaterium strain produces up to 115 mg/l/h 7beta-hydroxydehydroepiandrosterone in whole-cell conversions 1.14.15.22 vitamin D 1,25-hydroxylase synthesis expression of mutant R73V/R84A in Streptomyces lividans TK23 cells under the control of the tipA promoter leads to synthesis of 25-hydroxyvitamin D3 and large amounts of 1alpha,25-dihydroxyvitamin D3. In addition, polar metabolites 1alpha,25(R),26-trihydroxyvitamin D3 and 1alpha,25(R),26-trihydroxyvitamin D3 are observed at a ratio of 5:1 1.14.15.24 beta-carotene 3-hydroxylase synthesis synthesis of astaxanthin. Astaxanthin is a carotenoid of significant commercial value due to its superior antioxidant potential and wide applications in the aquaculture, food, cosmetic and pharmaceutical industries. The Brevundimonas sp. SD212 crtW and Pantoea ananatis crtZ genes are the best combination for astaxanthin production. After balancing the activities of beta-carotene ketolase and hydroxylase, an Escherichia coli ASTA-1 that carries neither a plasmid nor an antibiotic marker is constructed to produce astaxanthin as the predominant carotenoid (96.6%) with a specific content of 7.4 mg/g dry cell weight without an addition of inducer 1.14.15.24 beta-carotene 3-hydroxylase synthesis the combination of the foreign bacterial enzyme CrtW and the endogenous carotenoid biosynthesis enzymes for synthesizing antheraxanthin in tobacco plants isable to produce the novel carotenoid 4-ketoantherazanthin 1.14.15.36 sterol 14alpha-demethylase (ferredoxin) synthesis construction of an artificial self-sufficient cytochrome P450 monooxygenase by fusion of NADPH-P450 reductase FprD, CYP51, and iron-sulfur containing FprD. CYP51-FprD fusion enzymes F1 and F2 in recombinant Escherichia coli catalyze demethylation of lanosterol more efficiently, with kcat/Km values about 35fold higher compared to those of CYP51 and FprD alone 1.14.15.38 N,N-dimethyl phenylurea N-demethylase synthesis expression of PdmAB in Escherichia coli, Pseudomonas putida, and other sphingomonads results in a functional enzyme. Coexpression of a putative [3Fe-4S]-type ferredoxin from Sphingomonas sp. strain RW1 greatly enhances the catalytic activity of PdmAB in Escherichia coli 1.14.16.2 tyrosine 3-monooxygenase synthesis immobilization of tyrosinase on polyacrylamide-based support for production of L-dopa from L-tyrosine thereby modifying the enzyme activity to tyrosine hydroxylase 1.14.17.3 peptidylglycine monooxygenase synthesis usage of the recombinant PAM for insulin analogue amidation producing insulin glargine amide, an insulin derivative that shows a time/effect profile which is distinctly more flat and thus more advantageous than insulin glargine itself. The enzyme is used to modify glycine-extended A22(G)-B31(K)-B32(R) human insulin analogue (GKR). Hypoglycemic activity of amidated and non-amidated insulin is compared 1.14.18.2 CMP-N-acetylneuraminate monooxygenase synthesis enzyme CMAH can be used for large-scale production of N-glycolylneuraminic acid 1.14.18.4 phosphatidylcholine 12-monooxygenase synthesis CpFAH, thet D12 oleate hydroxylase of nonplant origin, is a good candidate for the transgenic production of hydroxyl fatty acids in oilseed crops 1.14.18.4 phosphatidylcholine 12-monooxygenase synthesis at the normal growth temperature of 30°C, Schizosaccharomyces pombe cells harboring FAH12 expression vector grow poorly when the FAH12 gene expression is induced. At 37°C, there is almost no growth inhibition. After preliminary growth at 37°C followed by a 5-day incubation at 20°C , the level of ricinoleic acid reaches 137.4 microg/ml of culture which corresponds to 52.6% of total fatty acids 1.14.18.4 phosphatidylcholine 12-monooxygenase synthesis coexpression of fatty acid hydroxylase FAH and diacylglycerol acyl transferase DGAT1 in Pichia pastoris produces higher lipid contents and ricinoleic acid levels than expression of FAH alone. Coexpression in a mutant haploid strain defective in the DELTA12 desaturase activity results in a higher level of ricinoleic acid than that in the diploid strain. The ricinoleic acid produced is mainly distributed in the neutral lipid fractions, particularly the free fatty acid form, but with little in the polar lipids 1.14.18.4 phosphatidylcholine 12-monooxygenase synthesis overexpression of multicopy suppressor Plg7, encoding phospholipase A2, in combinantion with oleate DELTA12-hydroxylase gene FAH12 enables Schizosaccharomyces pombe cells to secrete free ricinoleic acid into culture media. The FAH12 integrant in the absence of the overexpressed plg7 reaches 200 microg/ml of intracellular ricinoleic acid and only 69.3 microg/ml in culture media. The FAH12 integrant harboring the plg7 multicopy plasmid secretes ricinoleic acid in the media (184.5 microg/ml) without decreasing the amount in the cells 1.14.19.3 acyl-CoA 6-desaturase synthesis DELTA 6-fatty acid desaturase is used in the synthesis of polyunsaturated fatty acids from microorganisms to higher animals, including arachidonic acid and eicosapentaenoic acid 1.14.19.3 acyl-CoA 6-desaturase synthesis Oenothera biennis is grown commercially for gamma-linolenic acid production by the fatty acid DELTA6-desaturase enzymatic reaction 1.14.19.19 sphingolipid 10-desaturase synthesis the enzyme can be useful for production of fusaruside, a 10,11-unsaturated immunosuppressive fungal sphingolipid with medical potentials for treating liver injury and colitis, DELTA 10(E)-sphingolipid desaturase as a new regiospecific biocatalyst 1.14.19.49 tetracycline 7-halogenase synthesis a strain lacking CtcP activity accumulates tetracycline with a yield of 18.9 g/l. Overexpressionof ctcP in Streptomyces aureofaciens leads to CTC production to a final titer of 25.9g/l 1.14.19.58 tryptophan 5-halogenase synthesis use of enzyme in selective halogenation of organic compounds 1.14.19.58 tryptophan 5-halogenase synthesis regeneration of FADH2 using flavin dehydrogenase plus formiate dehydrogenase from Candida boidiniiat 0.28 U/ml leads to about 60% increase in yield 1.14.19.64 (S)-stylopine synthase synthesis a microbial system is established for producing a protoberberine-type alkaloid (stylopine) in Pichia cells 1.14.19.65 (S)-cheilanthifoline synthase synthesis a microbial system is established for producing a protoberberine-type alkaloid (stylopine) in Pichia cells 1.14.19.69 biflaviolin synthase synthesis enzyme can be used as biocatalyst for oxidative C-C coupling reactions and to facilitate the synthesis of diverse coupling products 1.14.20.1 deacetoxycephalosporin-C synthase synthesis enzyme produces the 7-aminodeacetoxycephalosporanic acid precursor utilized in industrial applications, optimization of enzyme activity for activity with the substrate penicillin G 1.14.20.1 deacetoxycephalosporin-C synthase synthesis expression in Escherichia coli, overexpression as an insoluble and inactive enzyme, elaboration of refolding scheme resulting in highliy active and moderately stable enzyme 1.14.20.1 deacetoxycephalosporin-C synthase synthesis bioconversion of penicillins to cephalosporins using deacetoxycephalosporin C synthase (DAOCS) is an alternative and environmentally friendly process for production of 7-aminodeacetoxycephalosporanic acid (7-ADCA), a key intermediate of many clinically useful semisynthetic cephalosporins 1.14.20.B3 cycloclavine dioxygenase (alternatively: multifunctional dioxygenase EasH) synthesis synthesis of alkaloid cycloclavine using a yeast-based expression platform at titers of more than 500 mg/l 1.14.20.5 flavone synthase I synthesis use of enzyme for construction of a system for producing unnatural flavonoids and stilbenes in Escherichia coli by expression of the respective genes on three plasmids. Incubation of the recombinant Escherichia coli with exogenously supplied carboxylic acids leads to production of 87 different polyketides, including 36 unnatural flavonoids and stilbenes 1.14.20.6 flavonol synthase synthesis production of active flavonol synthase from Camellia sinensis in Escherichia coli, with high specific activity 1.14.20.6 flavonol synthase synthesis functional expression of plant-derived O-methyltransferase, flavanone 3-hydroxylase, and flavonol synthase in Corynebacterium glutamicum for production of pterostilbene, kaempferol, and quercetin 1.14.20.7 2-oxoglutarate/L-arginine monooxygenase/decarboxylase (succinate-forming) synthesis biological ethylene production can be achieved via expression of the ethylene-forming enzyme (EFE), found in some bacteria and fungi. It has the potential to provide a sustainable alternative to steam cracking. Ethylene is an important industrial compound for the production of a wide variety of plastics and chemicals 1.14.20.8 (-)-deoxypodophyllotoxin synthase synthesis recombinant coexpression of the genes involved in the podophyllotoxin pathway permits to reconstitute the pathway to (-)-4'-demethylepipodophyllotoxin (the etoposide aglycone), a naturally occurring lignan that is the immediate precursor of etoposide and, unlike podophyllotoxin, a potent topoisomerase inhibitor, for production of the etoposide aglycone in tobacco and circumvent the need for cultivation of mayapple and semisynthetic epimerization and demethylation of podophyllotoxin 1.14.20.13 6beta-hydroxyhyoscyamine epoxidase synthesis Escherichia coli cells harboring mutant S14P/K97A in a 5-l-bioreactor produce scopolamine via a single enzyme-mediated two-step transformation from 500 mg/l hyoscyamine in 97% conversion 1.14.20.13 6beta-hydroxyhyoscyamine epoxidase synthesis overexpression in hairy roots of Atropa baetica leeds to an altered alkaloid profile in which hyoscyamine is entirely converted into scopolamine. Scopolamine accumulation increases up to 9fold amounting to 5.6 mg per g dry weight 1.14.99.14 progesterone 11alpha-monooxygenase synthesis membrane-associated enzymes catalyse several reactions of industrial interest, including steroid hydroxylation 1.14.99.14 progesterone 11alpha-monooxygenase synthesis immobilisation of membrane-bound multi-enzyme complexes for industrial use 1.14.99.14 progesterone 11alpha-monooxygenase synthesis because of its capacity to hydroxylate steroids, CYP106A2 is an interesting candidate for industrial production of steroids or their intermediates. CYP106A2 is known as a 15beta-hydroxylase, but also shows minor 11alpha-hydroxylase activity for progesterone. 11alpha-Hydroxyprogesterone is an important pharmaceutical compound with anti-androgenic and blood-pressure-regulating activity 1.14.99.15 4-methoxybenzoate monooxygenase (O-demethylating) synthesis CYP199A2 may be a valuable biocatalyst for the regioselective oxidation of various aromatic carboxylic acids 1.14.99.24 steroid 9alpha-monooxygenase synthesis enzyme occurs in many bacterial genera used in industrial processes 1.14.99.24 steroid 9alpha-monooxygenase synthesis steroid 9alpha-hydroxylase in company with delta-dehydrogenase is used in industrial processes 1.14.99.50 gamma-glutamyl hercynylcysteine S-oxide synthase synthesis the ergothioneine biosynthetic pathway (EgtA-EgtE catalysis) provides an opportunity for ergothioneine production through metabolic engineering 1.14.99.52 L-cysteinyl-L-histidinylsulfoxide synthase synthesis potential application of the oxidative coupling between hercynine and cysteine for industrial ergothioneine production 1.14.99.52 L-cysteinyl-L-histidinylsulfoxide synthase synthesis the ergothioneine biosynthetic pathway (EgtA-EgtE catalysis) provides an opportunity for ergothioneine production through metabolic engineering, but the kinetic properties and the presence of side-reaction render OvoA not suitable for use in ergothioneine production through metabolic engineering 1.14.99.53 lytic chitin monooxygenase synthesis construction of a cassette vector containing the XylS/Pm system that can easily be used for exchanging LPMO coding genes with or without signal sequences. The cassette reliably produces mature (translocated) LPMOs under controlled conditions. The signal sequence of LPMO10A from Serratia marcescens gives highest levels of recombinant protein production and translocation 1.14.99.53 lytic chitin monooxygenase synthesis efficient production in Escherichia coli is achieved using PelB as the most productive signal peptide for the extracellular production of CBP21 and Aeromonas veronii B565 chitinase Chi92 1.14.99.54 lytic cellulose monooxygenase (C1-hydroxylating) synthesis construction of a cassette vector containing the XylS/Pm system that can easily be used for exchanging LPMO coding genes with or without signal sequences. The cassette reliably produces mature (translocated) LPMOs under controlled conditions. The signal sequence of LPMO10A from Serratia marcescens gives highest levels of recombinant protein production and translocation 1.14.99.61 cyclooctat-9-en-7-ol 5-monooxygenase synthesis CotB3 can be employed to functionalize other diterpene olefins, such as (-)-casbene to yield sinularcasbane D 1.14.99.61 cyclooctat-9-en-7-ol 5-monooxygenase synthesis efficient heterologous reconstitution of the cyclooctatin biosynthesis cluster by introduction of a novel, non-native Streptomyces-derived redox system (reductase/ferredoxin system from Streptomyces afghaniensis). The complete, heterologous biosynthesis cascade is established in Escherichia coli. The microbial production system allows for a 43fold increased cyclooctatin yield compared to the native producer Streptomyces melanosporofaciens 1.14.99.62 cyclooctatin synthase synthesis efficient heterologous reconstitution of the cyclooctatin biosynthesis cluster by introduction of a novel, non-native Streptomyces-derived redox system (reductase/ferredoxin system from Streptomyces afghaniensis). The complete, heterologous biosynthesis cascade is established in Escherichia coli. The microbial production system allows for a 43fold increased cyclooctatin yield compared to the native producer Streptomyces melanosporofaciens 1.14.99.63 beta-carotene 4-ketolase synthesis construction of an astaxanthin biosynthesis pathway in Saccharomyces cerevisiae by introducing heterologous beta-carotene hydroxylase (CrtZ) and beta-carotene ketolase (CrtW) into an existing high beta-carotene producing strain. Astaxanthin yield of 3.1 mg/g dry cell weight are achieved. After change of promoter and hrough high cell density fed-batch fermentation using a carbon source restriction strategy, the production of astaxanthin in a 5-L bioreactor reaches to 81.0 mg/l 1.14.99.63 beta-carotene 4-ketolase synthesis synthesis of astaxanthin. Astaxanthin is a carotenoid of significant commercial value due to its superior antioxidant potential and wide applications in the aquaculture, food, cosmetic and pharmaceutical industries. The Brevundimonas sp. SD212 crtW and Pantoea ananatis crtZ genes are the best combination for astaxanthin production. After balancing the activities of beta-carotene ketolase and hydroxylase, an Escherichia coli ASTA-1 that carries neither a plasmid nor an antibiotic marker is constructed to produce astaxanthin as the predominant carotenoid (96.6%) with a specific content of 7.4/g dry cell weight without an addition of inducer 1.14.99.63 beta-carotene 4-ketolase synthesis synthesis of the 4-ketoantheraxanthin by Nicotiana tabacum by combination of the CrtW and CrtZ genes from Brevundimonas sp. SD21211 and the endogenous carotenoid biosynthesis enzymes 1.15.1.1 superoxide dismutase synthesis coexpression of enzyme and variants with yeast copper chaperone yCCS in Escherichia coli, in medium supplemented with copper and zinc, with high yield and activity 1.15.1.1 superoxide dismutase synthesis production of extracellular superoxide dismutase in Pichia pastoris. Secretion of enzyme into the culture medium to a concentration of 440 mg/l and an antioxidative activity of 760 U/mg of enzyme. Transformed yeast cells are more resistant to heat shock and H2O2 oxidative stress 1.17.1.4 xanthine dehydrogenase synthesis immobilized xanthine dehydrogenase for use in organic synthesis 1.17.1.4 xanthine dehydrogenase synthesis expression of enzyme in baculovirus-insect cell system, yields a mixture of native dimeric, demolydbo-dimeric and monomeric forms. All forms contain flavin, the monomeric forms lack molybdopterin and the iron-sulfur centers. Monomeric forms require only three electrons for complete reduction 1.17.1.4 xanthine dehydrogenase synthesis expression of genes xdhAB encoding the two subunits of enzyme, in Escherichia coli, produces active enzyme with moleybdenum content of 0.11-0.16 mol per alphabeta protomer and iron and FAD levels at stoichiometries similar to native enzyme. Coexpression of xdhAB genes with Pseudomonas aeruginosa xdhC gene increases level of molybdenum incorporated to a 1:1 stoichiometry and results in high levels of functional protein, up to 2284 units per mg and 8039 mg per l 1.17.1.4 xanthine dehydrogenase synthesis for maximal level of functional expression, co-expression of xdhC gene is required which increases level of molybdenum incorporation. Iron and FAD content of expressed enzymes are independent of xdhC expression 1.17.1.9 formate dehydrogenase synthesis production of NADH from NAD+ 1.17.1.9 formate dehydrogenase synthesis Formate dehydrogenase is a biocatalyst for converting CO2 to formic acid in ambient conditions. For improvement of the formic acid production efficiency with this system, the enhancement of electron relay processes among a photosensitizer, an electron carrier and formate dehydrogenase is required. By using viologen derivatives with carbamoyl group, 1,1'-dicarbamoylmethyl-4,4'-bipyridinium diiodide and 1-carbamoylmethyl-1'-methyl-4,4'-bipyridinium diiodide the effective formic acid production in the system of water-soluble zinc porphyrin and formate dehydrogenase is improved compared with that of methylviologen 1.17.1.9 formate dehydrogenase synthesis potential applications in NAD(H)-dependent industrial biocatalysis as well as in the production of renewable fuels and chemicals from carbon dioxide. Formate dehydrogenase from Myceliophthora thermophile possess a huge potential for CO2 reduction or NADH generation and under extreme alkaline conditions 1.17.1.10 formate dehydrogenase (NADP+) synthesis mutant FDH 1.17.1.10 formate dehydrogenase (NADP+) synthesis synthesis of chiral compounds, NADP+ regeneration system 1.17.3.2 xanthine oxidase synthesis the enzyme can be used for production of superoxide, from oxidation of an aldehyde, which in a co-oxidation system reacts with the aldehyde and converts beta-carotene to beta-ionone, method optimization, overview 1.17.7.4 4-hydroxy-3-methylbut-2-en-1-yl diphosphate reductase synthesis recombinant expression of enzyme can be greatly enhanced by coexpression of genes from the isc operon which are required for the assembly of iron-sulfur clusters 1.18.1.5 putidaredoxin-NAD+ reductase synthesis omega-hydroxylation of medium-chain n-alkanes and primary alcohols by CYP153 enzymes from Mycobacterium marinum and Polaromonas sp. strain JS666 in presence of putidaredoxin and putidaredoxin reductase. Without putidaredoxin and putidaredoxin reductase, CYP153A16 shows negligible P450 oxidation activity towards n-octane 1.18.1.5 putidaredoxin-NAD+ reductase synthesis stereoselective production of pravastatin from avastatin by Escherichia coli co-expressing putidaredoxin reductase and putidaredoxin from Pseudomonas putida with cytochrome P450 105A3 from Streptomyces carbophilus. Improved active P450 expression leads to 4.1fold increase in pravastatin yield 1.18.6.1 nitrogenase synthesis MoFe subunit mutants V70A and V70G will catalyze the reduction and coupling of CO to form methane, ethane, ethylene, propene, and propane. The rates and ratios of hydrocarbon production from CO can be adjusted by changing the flux of electrons through nitrogenase, by substitution of other amino acids located near the FeMo-cofactor, or by changing the partial pressure of CO. Increasing the partial pressure of CO shifts the product ratio in favor of the longer chain alkanes and alkenes 1.20.1.1 phosphonate dehydrogenase synthesis production of deuterium- or tritium-labeled substances, mutant enzymes could be applied as NADPH regeneration systems 1.21.1.1 iodotyrosine deiodinase synthesis high expression of a truncated derivative lacking the membrane domain, residues 2-33, at its N-terminal is observed in Sf9 cells, whereas expression in Pichia pastoris remains low despite codon optimization. The desired expression in Escherichia coli can be achieved after replacing the two conserved Cys residues of the deiodinase with Ala and fusing the resulting protein to thioredoxin. This construct provides abundant enzyme for crystallography and mutagenesis 1.21.3.1 isopenicillin-N synthase synthesis direct enzymic synthesis of antibiotics 1.21.3.3 reticuline oxidase synthesis substrate tuning by introducing a fluoro moiety at one potential reactive carbon center switches the reaction to the formation of exclusively one regioisomer with perfect enantioselectivity. The formation of 11-hydroxy-functionalized tetrahydroprotoberberines instead of the commonly formed 9-hydroxy-functionalized products from 1,2,3,4-tetrahydroisoquinolines can be successfully promoted 1.21.3.3 reticuline oxidase synthesis heterologous production of berberine and the optimization of the engineered biosynthetic pathway from rac-norlaudanosoline to (S)-canadine in yeast involving the recombinant berberine reductase 1.21.3.10 hercynylcysteine S-oxide synthase synthesis the ergothioneine biosynthetic pathway (EgtA-EgtE catalysis) provides an opportunity for ergothioneine production through metabolic engineering 1.21.3.10 hercynylcysteine S-oxide synthase synthesis expression of EGT1 from Neurospora crassa and EGT2 from Claviceps purpurea in Yarrowia lipolytica to obtain 158 mg/l of ergothioneine in small-scale cultivation. An additional copy of each gene improves the titer to 205 mg/l. A phosphate-limited fed-batch fermentation in 1 l bioreactors yields 1.63 g/l ergothioneine in 220 h 1.21.3.10 hercynylcysteine S-oxide synthase synthesis overexpression of the Egt-1 and -2 genes of Neurospora crassa in Aspergillus oryzae produces ergothioneine (231.0 mg/kg of media), with significant amounts of hercynine (32.7 mg/kg) accumulating in the culture 1.21.98.3 anaerobic magnesium-protoporphyrin IX monomethyl ester cyclase synthesis simplified method to prepare Mg-protoporphyrin monomethyl ester from freeze dried BchE mutant Rhodobacter capsulatus DB575 cells by extraction with acetone/H2O/25% NH3. Isolated Mg-protoporphyrin monomethyl ester can be identified by absorption and fluorescence spectroscopy, and its purity analyzed by HPLC. The extracted Mg-protoporphyrin monomethyl ester can be dried and redissolved in buffered DMSO to be used as substrate. Mg-protoporphyrin monomethyl ester is in addition a substrate to magnesium-protoporphyrin IX monomethyl ester (oxidative) cyclase, EC 1.13.14.81 2.1.1.4 acetylserotonin O-methyltransferase synthesis recombinant Escherichia coli expressing sheep serotonin N-acetyltransferase with rice caffeic acid O-methyltransferase produces 1.46 mg/L of melatonin in a culture medium, in response to 1 mM serotonin 2.1.1.6 catechol O-methyltransferase synthesis predictive artificial neural network models of the soluble COMT activity for production in a batch Escherichia coli culture process. Models predict a maximum COMT activity of 183.73 nmol/h, at 40°C, pH 6.5 and stirring rate of 351 rpm, and 132.90 nmol/h, at 35°C, pH 6.2 and stirring rate of 351 rpm, for semi-defined and complex medium, respectively. These results represent a 4fold increase in total activity by comparison to the standard operational conditions 2.1.1.53 putrescine N-methyltransferase synthesis overexpression of enzyme is not sufficient to boost scopolamine biosynthesis. Simultaneous overexpression of enzyme and hyoscyamine 6 beta-hydroxylase results in sígnificant enhancement of scopolamine accumulation 2.1.1.53 putrescine N-methyltransferase synthesis overexpression of enzyme leads to enhanced N-methylputrescine levels, but the subsequent alkaloid metabolites are not affected 2.1.1.56 mRNA (guanine-N7)-methyltransferase synthesis combination of chemical synthesis and enzymatic methylation in order to produce large amounts of RNA oligonucleotides carrying a cap-0 or cap-1. Short RNAs are synthesized on solid support by the phosphoramidite 2'-O-pivaloyloxymethyl chemistry. The cap structure is then coupled by the addition of GDP after phosphorylation of the terminal 5'-OH and activation by imidazole. GpppN-RNAs or GpppN2'-Om-RNAs are purified before the N7-methyl group is added using the human (guanine-N7)-methyl transferase to yield 7mGpppN-RNAs (cap-0) or 7mGpppN29-Om-RNAs (cap-1) 2.1.1.68 caffeate O-methyltransferase synthesis engineered COMT enzymes can be useful for metabolic engineering of both lignin and benzaldehyde-derived flavors and fragances 2.1.1.B74 apigenin 7-O-methyltransferase synthesis expression together with basil flavonoid 6-hydroxylase encoded by CYP82D33 and further ethyltransferases in Saccharomyces cerevisiae to produce 8- and/or 6-substituted, methoxylated flavones from their natural precursor apigenin. The yeast cells produce substantial amounts of 6-hydroxylated, methylated derivatives of naringenin and luteolin while the corresponding derivatives of flavonol kaempferol are only detected in trace amounts. The substrate specificity of both the hydroxylases and the flavonoid O-methyltransferases is limiting yields obtained with alternative substrates 2.1.1.B75 flavonoid 7-O-methyltransferase synthesis the recombinant SaOMT is used for 7-O-methylation of naringenin in recombinantly engineered production of 7-O-methyl aromadendrin, which is important in medicinal applications, via sakuranetin from 4-coumaric acid in Escherichia coli, overview 2.1.1.B75 flavonoid 7-O-methyltransferase synthesis OMT2 exhibits more diverse regiospecificity and catalyzes mono-, di-, and tri-methylation of stilbene, flavanone, and flavone when it is expressed in Streptomyces venezuelae compared to expression in Escherichia coli. For the efficient production of multi-methylated phenylpropanoids, a cocultivation system uses engineered Escherichia coli strains producing pterostilbene, naringenin, and apigenin, respectively, along with a OMT2-expressing Streptomyces venezuelae mutant 2.1.1.B76 flavone/flavonol 7-O-methyltransferase synthesis using Escherichia coli transformants harboring both PFNS-1 and POMT-7, naringenin could be converted into genkwanin 2.1.1.B76 flavone/flavonol 7-O-methyltransferase synthesis mass production of rhamnetin by recombinant Escherichia coli for medical use 2.1.1.88 8-hydroxyquercetin 8-O-methyltransferase synthesis O-methyltransferases (OMTs) from microorganism sources have broad scope to methylate diverse classes of plant natural products to synthesize methyl derivatives. Enzyme SpOMT2884 from Streptomyces peucetius cloned into Escherichia coli shows the possibility of generating diverse O-methyl flavonoids 7,8-dihydroxyflavone (7,8-DHF), luteolin, quercetin, rutin, naringenin, daidzein, and formononetin 2.1.1.95 tocopherol C-methyltransferase synthesis introducing the enzyme into soybean can be used to increase the vitamin E composition in seeds 2.1.1.111 anthranilate N-methyltransferase synthesis expression of anthranilate N-methyltransferase, anthraniloyl-coenzyme A (CoA):methanol acyltransferase from Vitis labrusca, and anthranilate coenzyme A ligase from Pseudomonas aeruginosa in Escherichia coli mutants mutants (metJ, trpD, tyrR), which provide more anthranilate and/or S-adenosyl methionine, to synthesize N-methylanthranilate, methyl anthranilate, and methyl N-methylanthranilate. Approximately, 185.3 microM N-methylanthranilate and 95.2 microM methyl N-methylanthranilate are synthesized 2.1.1.111 anthranilate N-methyltransferase synthesis production of 4-hydroxycoumarin, 2,4-dihydroxyquinoline, and 4-hydroxy-1-methyl-2(1H)-quinolone in Escherichia coli using genes for the synthesis of salicylic acid from chorismate to supply the substrate for 4-hydroxycoumarin and the gene encoding N-methyltransferase for the synthesis of N-methylanthranilate from anthranilate. 255.4 mg/l 4-hydroxycoumarin, 753.7 mg/l dihydroxyquinoline, and 17.5 mg/l 4-hydroxy-1-methyl-2(1H)-quinolone are synthesized 2.1.1.111 anthranilate N-methyltransferase synthesis three anthranilate derivatives, N-methylanthranilate, methyl anthranilate, and methyl N-methylanthranilate are synthesized using metabolically engineered stains of Escherichia coli. NMT encoding N-methyltransferase from Ruta graveolens, AMAT encoding anthraniloyl-coenzyme A (CoA):methanol acyltransferase from Vitis labrusca, and pqsA encoding anthranilate coenzyme A ligase from Pseudomonas aeruginosa are cloned and Escherichia coli strains harboring these genes were used to synthesize the three desired compounds. Escherichia coli mutants (metJ, trpD, tyrR mutants), which provide more anthranilate and/or S-adenosyl methionine, are used to increase the production of the synthesized compounds. 0.1853 mM N-methylanthranilate and 0.0952 mM methyl N-methylanthranilate are synthesized 2.1.1.115 (RS)-1-benzyl-1,2,3,4-tetrahydroisoquinoline N-methyltransferase synthesis the enzyme immobilized on CH-Sepharose or CPG-10 glass beads is a useful tool for the preparative synthesis of isotopically labelled N-methylated benzylisoquinoline alkaloids 2.1.1.128 (RS)-norcoclaurine 6-O-methyltransferase synthesis production of the economically important analgesic morphine and the antimicrobial agent berberine 2.1.1.130 precorrin-2 C20-methyltransferase synthesis preparative multi-enzyme synthesis of precorrin-3A 2.1.1.142 cycloartenol 24-C-methyltransferase synthesis rational drug target design based on structure-function-relation 2.1.1.160 caffeine synthase synthesis production of caffeine in metabolically engineered Escherichia coli by the expression of caffeine synthase TCS1. Caffeine accumulation is increased using higher-level expression of the target enzymes, and enhancement of xanthine and S-adenosyl-L-methionine biosynthesis. The final strain produces up to 21.46 mg/l caffeine from 20 g/l of glucose in shake flask culture, yielding caffeine up to 2.96 mg/g glucose 2.1.1.165 methyl halide transferase synthesis producing methyl halides from non-food agricultural resources by using a symbiotic co-culture of an engineered yeast and the cellulolytic bacterium Actinotalea fermentans, methyl halide production from unprocessed switchgrass (Panicum virgatum), corn stover, sugar cane bagasse, and poplar (Populus sp.). Methyl halides are used as agricultural fumigants and are precursor molecules that can be catalytically converted to chemicals and fuels 2.1.1.195 cobalt-precorrin-5B (C1)-methyltransferase synthesis coexpression of the cobA gene from Propionibacterium freudenreichii and the cbiA, -C, -D, -E, -T, -F, -G, -H, -J, -K, -L, and -P genes from Salmonella enterica serovar typhimurium in Escherichia coli result in the production of cobyrinic acid a,c-diamide. A cbiD deletion mutant of this strain produces 1-desmethylcobyrinic acid a,c-diamide, indicating that CbiD is involved in C-1 methylation in the anaerobic pathway to cobalamin 2.1.1.196 cobalt-precorrin-6B (C15)-methyltransferase [decarboxylating] synthesis coexpression of the cobA gene from Propionibacterium freudenreichii and the cbiA, -C, -D, -E, -T, -F, -G, -H, -J, -K, -L, and -P genes from Salmonella enterica serovar typhimurium in Escherichia coli result in the production of cobyrinic acid a,c-diamide 2.1.1.196 cobalt-precorrin-6B (C15)-methyltransferase [decarboxylating] synthesis investigation on the use of the oxidized form factor 3 of the trimethylated intermediate precorrin 3 as a substrate for the enzymes of the anaerobic pathway to vitamin B12. Production of factor 3 octamethylester by expression of enzymes enzymes CbiH, CbiG, CbiF, and CbiT, in Escherichia coli and addition of factor 3 octapiperidinium salt 2.1.1.231 flavonoid 4'-O-methyltransferase synthesis coexpression of Oryza sativa ROMT-9, which methylates specifically at the 3'-hydroxyl group of quercetin and SOMT-2, which methylates at the 4'-hydroxyl group, in Escherichia coli. Reaction products of quercetin with the transformant show two methylation products that correspond to the 3'-methylated and the 3',4'-dimethylated quercetin. More than 90% of quercetin is converted into the 3',4'-dimethylated quercetin after 24 h incubation 2.1.1.232 naringenin 7-O-methyltransferase synthesis the recombinant SaOMT is used for 7-O-methylation of naringenin in recombinantly engineered production of 7-O-methyl aromadendrin, which is important in medicinal applications, via sakuranetin from 4-coumaric acid in Escherichia coli, overview 2.1.1.232 naringenin 7-O-methyltransferase synthesis production of sakuranetin (7-O-methylnaringenin) and ponciretin (4-O-methylnaringenin) in Escherichia coli by reconstruction of the naringenin biosynthesis pathway. Engineering the shikimic acid pathway, and overexpression of several genes for the biosynthesis of ponciretin and sakuranetin such as tyrosine ammonia lyase, 4-coumaroyl CoA ligase, chalcone synthase, and O-methyltransferase and deletion of isocitrate dehydrogenase finally gives ponciretin and sakuranetin from glucose in Escherichia coli at the concentration of 42.5 mg/l and 40.1 mg/l, respectively 2.1.1.236 dTDP-3-amino-3,6-dideoxy-alpha-D-galactopyranose N,N-dimethyltransferase synthesis synthesis of TDP-D-ravidosamine, necessary for future in vitro glycosylation assays. TDP-D-ravidosamine is the anticipated sugar donor substrate of RavGT (the glycosyltransferase that links D-ravidosamine to the polyketide derived backbone defuco-gilvocarcin V). Defuco-gilvocarcin V exhibits superior anticancer/antibacterial activities 2.1.1.240 trans-resveratrol di-O-methyltransferase synthesis the enzyme from Sorghum bicolor, SbROMT3syn, can be used as an enzyme to produce pinostilbene by methylating resveratrol in microorganisms 2.1.1.240 trans-resveratrol di-O-methyltransferase synthesis the enzyme is useful for conversion of 4-coumaric acid to pterostilbene via co-expression of 4CL::STS and ROMT in recombinant Escherichia coli and Saccharomyces cerevisiae, overview 2.1.1.240 trans-resveratrol di-O-methyltransferase synthesis the Vitis riparia enzyme VrROMTsyn cannot be used as an enzyme to produce pinostilbene by methylating resveratrol in microorganisms, since it has no or very poor enzyme activity toward resveratrol as a substrate 2.1.1.240 trans-resveratrol di-O-methyltransferase synthesis introduction of enzyme into a resveratrol-producing Corynebacterium glutamicum strain allows synthesis of 42 mg/l (0.16 mM) of the di-O-methylated pterostilbene from p-coumaric acid. A fusion of O-methyltransferase with the maltose-binding protein of Escherichia coli lacking its signal peptide increases the solubility of the O-methyltransferase. Expression of heterologous dioxygenase genes in (2S)-flavanone-producing Corynebacterium glutamicum strains enables the production of flavanonols and flavonols starting from p-coumaric acid and caffeic acid. For the flavonols kaempferol and quercetin, maximum product titers of 23 mg/l (0.08 mM) and 10 mg/l (0.03 mM) can be achieved 2.1.1.240 trans-resveratrol di-O-methyltransferase synthesis the enzyme is used for production of large amounts of easily recoverable extracellular resveratrol in a bacterial system, method, overview. Constitutive expression of either Vitis vinifera resveratrol O-methyltransferase (VvROMT) or human cytochrome P450 hydroxylase 1B1 (HsCYP1B1) lead to pterostilbene or piceatannol, respectively, after the engineered cell cultures are treated with elicitors. i.e. methylated cyclodextrins and methyl jasmonate. Functionality of both gene products is first assessed in planta by Nicotiana benthamiana agroinfiltration assays, in which tobacco cells transiently express stilbene synthase and VvROMT or HsCYP1B1 2.1.1.246 [methyl-Co(III) methanol-specific corrinoid protein]:coenzyme M methyltransferase synthesis in vitro methanol production from methyl coenzyme M using the Methanosarcina barkeri MtaABC protein complex 2.1.1.280 selenocysteine Se-methyltransferase synthesis the ability to transfer the selenocysteine methyltransferase gene into a higher biomass, rapidly growing, and edible plant can lead to a cost effective method of producing Se-methyl-selenocysteine gamma-glutamylmethylselenocysteine enriched plant material 2.1.1.280 selenocysteine Se-methyltransferase synthesis expression of AbSMT in transgenic tomato enables production of MeSeCys from selenate or selenite and biofortification of a food crop with the anticancer compound MeSeCys is a possibility 2.1.1.281 phenylpyruvate C3-methyltransferase synthesis the nonenzymatic stereoselective formation of C-C bonds by methylation is challenging and usually requires multistep syntheses. In contrast, the asymmetric introduction of a methyl group in natural product biosynthesis is performed in a single step and catalyzed by methyltransferases (MTs), which usually require S-adenosylmethionine (SAM) as the methyl donor. Enzyme SgvM might be a valuable tool for asymmetric biocatalytic C-alkylation reactions. It catalyze the transfer of the electrophilic methyl group of SAM to the C3 position 2.1.1.283 emodin O-methyltransferase synthesis O-methyltransferases (OMTs) from microorganism sources have broad scope to methylate diverse classes of plant natural products to synthesize methyl derivatives. Enzyme SpOMT2884 from Streptomyces peucetius cloned into Escherichia coli shows the possibility of generating diverse O-methyl flavonoids 7,8-dihydroxyflavone (7,8-DHF), luteolin, quercetin, rutin, naringenin, daidzein, and formononetin 2.1.1.292 carminomycin 4-O-methyltransferase synthesis production of a new hybrid anthracycline antibiotic. Expression in Streptomyces violaceus pMK100 (epelmycin producer). The transformant produces the hybrid anthracycline antibiotic 7-O-L-hodosaminyl-4-O-methyl-epsilon-rhodomycinone (4-O-methylepelmycin D) together with host epelmycins when cultured in antibiotic production medium in the presence of thiostrepton. Attempts on production of hybrid 4-O-methylaclarubicin and 4-O-methyl-l-deoxyobelmycin by the transformants of aclarubicin and 1-deoxyobelmycin producers are unsuccessful 2.1.1.356 [histone H3]-lysine27 N-trimethyltransferase synthesis expression of NSDs via tagging with a human influenza hemagglutinin tag greatly improves the quality of the recombinant NSDs, resulting in more than 95% pure, stable, and active NSD-hemagglutinins, with an increase in production yield up to 22.4fold and up to 6.25 mg/l from LB Escherichia coli culture, and without further purification 2.1.1.357 [histone H3]-lysine36 N-dimethyltransferase synthesis expression of NSDs via tagging with a human influenza hemagglutinin tag greatly improves the quality of the recombinant NSDs, resulting in more than 95% pure, stable, and active NSD-hemagglutinins, with an increase in production yield up to 22.4fold and up to 6.25 mg/l from LB Eschewrichia coli culture, and without further purification 2.1.1.363 pre-sodorifen synthase synthesis introduction of the Serratia plymuthica WS3236 sodorifen biosynthetic gene cluster into Escherichia coli leads to a significant increase in both sodorifen product yield and purity compared to the native producer 2.1.1.371 [histone H3]-lysine27 N-dimethyltransferase synthesis expression of NSDs via tagging with a human influenza hemagglutinin tag greatly improves the quality of the recombinant NSDs, resulting in more than 95% pure, stable, and active NSD-hemagglutinins, with an increase in production yield up to 22.4fold and up to 6.25 mg/l from LB Escherichia coli culture, and without further purification 2.1.1.382 methoxylated aromatic compound—corrinoid protein Co-methyltransferase synthesis the demethylation by VdmB transforms a variety of aryl methyl ethers with two functional methoxy moieties either in 1,2-position or in 1,3-position. Biocatalytic reactions enable the regioselective monodemethylation of substituted 3,4-dimethoxy phenol as well as the monodemethylation of 1,3,5-trimethoxybenzene. The VdmB is also applied for the regioselective demethylation of natural compounds such as papaverine and rac-yatein. Best results are obtained in presence of orcinol as final methyl acceptor 2.1.2.1 glycine hydroxymethyltransferase synthesis improved method for preparation of optically pure beta-hydroxy-alpha-amino acids, catalyzed by serine hydroxymethyl transferase with threonine aldolase activity. Usage of substrates beta-phenylserine, beta-(nitrophenyl) serine and beta-(methylsulfonylphenyl) serine with immobilized recombinant enzyme for SHMT activity, optimal at pH 7.5 and 45°C. The immobilized cells are continuously used 10 times, yielding an average conversion rate of 60.4% 2.1.2.13 UDP-4-amino-4-deoxy-L-arabinose formyltransferase synthesis engineering of Escherichia coli to ynthesize the plant-specific flavonoid O-pentosides quercetin 3-O-xyloside and quercetin 3-O-arabinoside. For UDP-xylose biosynthesis, genes UXS (UDP-xylose synthase) from Arabidopsis thaliana and ugd (UDP-glucose dehydrogenase) from E.scherichia coli, are overexpressed. The gene encoding ArnA, which competes with UXS for UDP-glucuronic acid, is deleted. For UDP-arabinose biosynthesis, UXE (UDP-xylose epimerase) i overexpressed. UDP-dependent glycosyltransferases are engineered to ensure specificity for UDP-xylose and UDP-arabinose. The srains thus obtained synthesize approximately 160 mg/liter of quercetin 3-O-xyloside and quercetin 3-O-arabinoside 2.2.1.1 transketolase synthesis desing of an enzyme microreaktor by reversible immobilization of His6-tagged enzyme a 200-microm ID fused silica capillary for quantitative kinetic analysis. Transketolase kinetic parameters in the mircoreactor are comparable with those measured in free solution. Quantitative elution of the immobilized transketolase and the regeneration and reuse of the derivatized capillary over five cycles are possible 2.2.1.1 transketolase synthesis mathematical model for the modes of operation in the reaction of beta-hydroxypyruvate and glycolaldehyde as an alternative to a batch process. The performance of the system strongly depends on the solubility of beta-hydroxypyruvate. The best option for the base case scenario is to use an initial beta-hydroxypyruvate concentration at its solubility level with a slight excess of glycolaldehyde and an initial catalyst concentration of 0.5 g/l for 120 min 2.2.1.2 transaldolase synthesis overexpression of enzyme in Escherichia coli harboring poly-beta-hydroxybutyrate operon phbCAB leads to increase in poly-beta-hydroxybutyrate from 28.2% to 52.3% 2.2.1.2 transaldolase synthesis improvement of xylose-to-ethanol bioconversion by mutation Q263R, mutantion leads to 5fold increase in activity and increases ethanol production by 36% and 100% as measured by volumetric production rate and specific production rate, respectively 2.2.1.2 transaldolase synthesis mutation F178Y/R181E is based on mutant F178Y, which is able to use dihydroxyacetone as donor in aldol reactions. Mutant F178Y/R181E exhibits an at least fivefold increase in affinity towards glyceraldehyde and can use D- and L-glyceraldehyde as acceptor substrates, resulting in preparative synthesis of D-fructose, D-xylulose and L-sorbose when dihydroxyacetone is used as donor. Mutant enzyme does not show transaldolase activity 2.2.1.5 2-hydroxy-3-oxoadipate synthase synthesis application of the E1 component of the Escherichia coli 2-oxoglutarate dehydrogenase multienzyme complex in the synthesis of chiral ompounds with multiple functional groups in good yield and high enantiomeric excess, by varying both the donor substrate (different 2-oxo acids) and the acceptor substrate (glyoxylate, ethyl glyoxylate and methyl glyoxal). The enzyme can accept 2-oxovalerate and 2-oxoisovalerate in addition to its natural substrate 2-oxoglutarate, and the tested acceptors are also acceptable in the carboligation reaction 2.2.1.6 acetolactate synthase synthesis co-expression of acetolactate synthase and omega-transaminase in Escherichia coli as a whole-cell biocatalyst for production of (S)-alpha-benzylamine. Product (S)-alpha-benzylamine can be moved into the extraction solution via an organic solvent 2.2.1.6 acetolactate synthase synthesis construction of a mutant with a deleted C-terminal domain in the regulatory subunit IlvN. The constructed enzyme shows altered kinetic properties, i.e., an about twofold-lower Km for the substrate pyruvate and an about fourfold-lower Vmax, a slightly increased Km for the substrate alpha-ketobutyrate with an about twofold-lower Vmax, and insensitivity against the inhibitors L-valine, L-isoleucine, and L-leucine. Introduction of the mutant into the L-lysine producers Corynebacterium glutamicum DM1729 and DM1933 increases L-lysine formation by 43% and 36%, respectively. Complete inactivation of the AHAS in Corynebacterium glutamicum DM1729 and DM1933 by deletion of the ilvB gene, encoding the catalytic subunit of AHAS, leads to L-valine, L-isoleucine, and L-leucine auxotrophy and to further-improved L-lysine production. In batch fermentations, the mutant produces about 85% more L-lysine and shows an 85%-higher substrate-specific product yield 2.2.1.6 acetolactate synthase synthesis transformation of a H+-ATPase defective strain with a C-terminal truncation of acetohydroxyacid synthase gene ilvBN results in increased valine production from 21.7 mM for wild-type to 46.7 mM and increase in the valine intermediate acetoin. Inserting acetohydroxyacid isomeroreductase gene into the ilvBN plasmid further increases valine producion 2.2.1.6 acetolactate synthase synthesis Bacillus subtilis acetolactate synthase can act as key biocatalyst in the formation of isobutanol which is deemed to be a next-generation biofuel and a renewable platform chemical. The enzyme AlsS catalyzes the conversion of 2-ketoisovalerate into isobutyraldehyde, the immediate precursor of isobutanol 2.2.1.6 acetolactate synthase synthesis the enzyme in Pyrococcus furiosus is a potential platform for the biological production of acetoin at temperatures in the 70-80°C range. Acetoin, or 3-hydroxybutanone, is an important four-carbon compound that serves as a building block for valuable bio-based chemical compounds and is a common flavor additive and preservative in the food industry 2.2.1.6 acetolactate synthase synthesis acetoin (i.e. 3-hydroxybutanone), is a major product at temperatures below 80 °C. Acetolactate synthase ALS, which is involved in branched-chain amino acid biosynthesis, is responsible and deletion of the Als gene abolishes acetoin production. Deletion of Als in a strain of Pyrococcus furiosus heterologously expressing an alcohol dehydrogenase gene from Thermoanaerobacter sp. X514 for ethanol production significantly improves the yield of ethanol 2.2.1.6 acetolactate synthase synthesis construction of isobutanol production systems by overexpression of effective 2-oxoacid decarboxylase KivD and combinatorial overexpression of valine biosynthetic enzymes in Saccharomyces cerevisiae D452-2. Isobutanol production by the engineered strain is assessed in micro-aerobic batch fermentations using glucose as a sole carbon source, leading to production of 93 mg/l isobutanol, which corresponds to a fourfold improvement as compared with the control strain. Isobutanol production is further enhanced to 151 mg/l by additional overexpression of acetolactate synthase Ilv2p, acetohydroxyacid reductoisomerase Ilv5p, and dihydroxyacid dehydratase Ilv3p in the cytosol 2.2.1.6 acetolactate synthase synthesis engineering of the wild type of Corynebacterium glutamicum for the growth-decoupled production of 2-ketoisovalerate from glucose by deletion of the aceE gene encoding the E1p subunit of the pyruvate dehydrogenase complex, deletion of the transaminase B gene ilvE, and additional overexpression of the ilvBNCD genes, encoding the L-valine biosynthetic enzymes acetohydroxyacid synthase (AHAS), acetohydroxyacid isomeroreductase, and dihydroxyacid dehydratase. 2-Ketoisovalerate production is further improved by deletion of the pyruvate:quinone oxidoreductase gene pqo. In fed-batch fermentations at high cell densities, the newly constructed strains produce up to 188 mM (21.8 g/liter) 2-ketoisovalerate and showd a product yield of about 0.47 mol per mol (0.3 g/g) of glucose and a volumetric productivity of about 4.6 mM (0.53 g/liter) 2-ketoisovalerate per h in the overall production phase 2.2.1.6 acetolactate synthase synthesis overexpression of the als gene leads to high levels of 2,3-butanediol 2.2.1.7 1-deoxy-D-xylulose-5-phosphate synthase synthesis Agrobacterium tumefaciens strain KCCM 10413, overexpressing the enzyme might be useful in industrial production of ubiquinone-10, i.e. UbiQ(10) 2.3.1.4 glucosamine-phosphate N-acetyltransferase synthesis overexpression in Bacillus subtilis for synthesis of N-acetylglucosamine. The expression level of GNA1 is enhanced at the translational level via fusion of an epitope tag to the 5'-terminus of GNA1 gene and ribosome binding site sequence engineering. Enhanced expression of glutamine-fructose-6-phosphate aminotransferaseGlmS is achieved at the transcriptional and translational levels. Under the control of engineered GNA1 and GlmS, the GlcNAc titer and yield in the shake flask increase to 18.5 g/l and 0.37 g GlcNAc/g glucose 2.3.1.4 glucosamine-phosphate N-acetyltransferase synthesis use of enzyme for synthesis of N-acetylglucosamine in Bacillus subtilis. GNA1 is evolved through error-prone PCR under pyruvate stress to enhance its catalytic activity. Urease from Bacillus paralicheniformis is expressed intracellularly to neutralize the intracellular pH. The activity of mutant CeGNA1 increases by 11.5% at pH 6.5-7.5, with the catalytic efficiency increasing by 27.5% to 1.25 per s and microM. Modulated expression of urease increases the intracellular pH from 6.0 to 6.8. The final engineered produce 25.6 g/l N-acetylglucosamine with a yield of 0.43 g N-acetylglucosamine/g glucose in a shake flask fermentation and produces 82.5 g/l N-acetylglucosamine with a yield of 0.39 g GlcNAc/g glucose by fed-batch fermentation 2.3.1.8 phosphate acetyltransferase synthesis a lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta) deletion strain is evolved for 2,000 h, resulting in a stable strain with 40:1 ethanol selectivity and a 4.2-fold increase in ethanol yield over the wild-type strain. In a coculture of organic acid-deficient engineered strains of both Clostridium thermocellum and Thermoanaerobacterium saccharolyticum, fermentation of 92 g/liter Avicel results in 38 g/liter ethanol, with acetic and lactic acids below detection limits, in 146 h. engineering is based on a phosphoribosyl transferase (Hpt) deletion strain, which produces acetate, lactate, and ethanol in a ratio of 1.7:1.5:1.0, similar to the 2.1:1.9:1.0 ratio produced by the wild type. The Hpt/Ldh double mutant strain does not produce significant levels of lactate and has a 1.4:1.0 ratio of acetate to ethanol. Similarly, the Hpt/Pta double mutant strain does not produce acetate and has a 1.9:1.0 ratio of lactate to ethanol. The Hpt/Ldh/Pta triple mutant strain achieves ethanol selectivity of 40:1 relative to organic acids 2.3.1.8 phosphate acetyltransferase synthesis overexpression of phosphotransacetylase in the yeast Rhodosporidium toruloides for enhanced cell growth and lipid production. Compared with the parental strain, the engineered strain shows significant improvement in glucose consumption, cell growth and lipid accumulation when cultivated under nitrogen limited conditions in an Erlenmeyer flask as well as a stirred tank bioreactor. The phosphotransacetylase enzyme has little direct effects on the enzymes of fatty acid biosynthetic pathway, but might facilitate the fatty acid precursor acetyl-CoA supply 2.3.1.9 acetyl-CoA C-acetyltransferase synthesis the enzyme has biotechnological potential in haloarchaea for production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with controllable 3-oxopentano content, from unrelated cheap carbon sources 2.3.1.9 acetyl-CoA C-acetyltransferase synthesis engineering of Escherichia coli for direct and modulated biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer. An Escherichia coli strain with an activated sleeping beauty mutase (Sbm) operon is used to generate propionyl-CoA as a precursor. Two acetyl-CoA moieties or acetyl-CoA and propionyl-CoA are condensed to form acetoacetyl-CoA and 3-ketovaleryl-CoA, respectively, by functional expression of beta-ketothiolases from Cupriavidus necator (i.e. PhaA and BktB). The resulting thioester intermediates are channeled into the polyhydroxyalkanoate biosynthetic pathway through functional expression of acetoacetyl-CoA reductase (PhaB) for thioester reduction and PHA synthase (PhaC) for subsequent polymerization. High-level poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production ranging from 3 to 19 mol%, can be achieved 2.3.1.9 acetyl-CoA C-acetyltransferase synthesis upon overexpression of acetyl-CoA acetyltransferase (genes AtoAD) in an engineered Escherichia coli strain, carrying polyhydroxyalkanoate synthetic genes bktB, phaB, and phaC and with propionate as a cosubstrate, the 3-hydroxyvalerate faction in Polyhydroxyalkanoate increases up to 7.3fold compared to a strain without genes AtoAD expressed in trans (67.9 mol%). Overexpression of AtoAD decreases the amount of acetyl-CoA and increases the propionyl-CoA/acetyl-CoA ratio, ultimately resulting in an increased 3-hydroxyvalerate fraction in polyhydroxyalkanoate. Synthesis of poly(3-hydroxybutanoate-co-3-hydroxyvalerate) containing 57.9 mol% of 3-hydroxyvalerate is achieved by fed-batch fermentation with propionate 2.3.1.15 glycerol-3-phosphate 1-O-acyltransferase synthesis potential of genetic manipulation of GPAT to increase the glycerolipid level in Lobosphaera incisa and other microalgae 2.3.1.16 acetyl-CoA C-acyltransferase synthesis engineering of Escherichia coli for direct and modulated biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer. An Escherichia coli strain with an activated sleeping beauty mutase (Sbm) operon is used to generate propionyl-CoA as a precursor. Two acetyl-CoA moieties or acetyl-CoA and propionyl-CoA are condensed to form acetoacetyl-CoA and 3-ketovaleryl-CoA, respectively, by functional expression of beta-ketothiolases from Cupriavidus necator (i.e. PhaA and BktB). The resulting thioester intermediates are channeled into the polyhydroxyalkanoate biosynthetic pathway through functional expression of acetoacetyl-CoA reductase (PhaB) for thioester reduction and PHA synthase (PhaC) for subsequent polymerization. High-level poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production ranging from 3 to 19 mol%, can be achieved 2.3.1.19 phosphate butyryltransferase synthesis the phosphotransbutyrylase promoter is used to produce butanol from butyryl-CoA with alcohol/aldehyde dehydrogenase 2.3.1.21 carnitine O-palmitoyltransferase synthesis comparison of COS7 cell and yeast expression of isoform CPT1b. The mitochondrial fraction prepared from yeast cells expressing CPT1b shows 25% higher CPT1 activity than that obtained from COS7 cells. The expression level of CPT1b in the former is 3.8 times lower than that in the latter; and thus the specific activity of CPT1b expressed in yeast cells is estimated to be approximately five times higher than that expressed in COS7 cells 2.3.1.B25 octaketide synthase synthesis the enzyme can be used in the production of carminic acid, a C-glucosylated octaketide anthraquinone and the main constituent of the natural dye carmine (E120). Carmine possesses unique coloring, stability, and solubility properties 2.3.1.B25 octaketide synthase synthesis the enzyme OKS can be used in the heterologous production of the widely used natural food colorant carminic acid in Aspergillus nidulans, method, overview 2.3.1.28 chloramphenicol O-acetyltransferase synthesis engineering of a thermostable chloramphenicol acetyltransferase for enhanced isobutyl acetate production at elevated temperatures. Mutant F97W significantly increases conversion of isobutanol to isobutyl acetate. Using F97W, direct conversion of cellulose into isobutyl acetate is possible by an engineered Clostridium thermocellum strain at elevated temperatures 2.3.1.29 glycine C-acetyltransferase synthesis enzyme is applied in synthesis of 2,5-dimethylpyrazine. Bacillus subtilis can synthesize 2,5-dimethylpyrazine by using L-threonine as the substrate via aminoacetone. The reaction from L-threonine to L-2-amino-acetoacetate is catalyzed by L-threonine-3-dehydrogenase. L-2-Amino-acetoacetate can spontaneously decarboxylate to form aminoacetone. The reaction from aminoacetone to 2,5-dimethylpyrazine is a pH-dependent nonenzymatic reaction. Inactivation of 2-amino-3-ketobutyrate CoA ligase improves 2,5-dimethylpyrazine production 2.3.1.37 5-aminolevulinate synthase synthesis optimization of recombinant aminolevulinate synthase production using factorial design. Initial succinate, glucose, and IPTG concentration are key factors affecting enzyme activity. Fermentation yields up to 5.2 g/l 5-aminolevulinate 2.3.1.37 5-aminolevulinate synthase synthesis production of 5-aminolevulinic acid by a genetic engineering. As glycine and succinate are inexpensive, the enzymatic production of 5-aminolevulinic acid from glycine and succinate might be an attractive process 2.3.1.37 5-aminolevulinate synthase synthesis by expression in Escherichia coli Rosetta and combining D-xylose as a inhibitor for 5-aminolevulinate dehydratase with D-glucose in fed-batch culture and based on the optimal culture system, the yield of 5-aminolevulinate achieved is 7.3 g/l or 56 mM under the appropriate conditions 2.3.1.37 5-aminolevulinate synthase synthesis in Escherichia coli expressing 5-aminolevulinate synthase, increasing D-glucose concentration in culture enhances final cell density and 5-aminolevulinate yield and simultaneously decreases the activities of 5-aminolevulinate synthase and 5-aminolevulinate dehydratase. The inhibitory effect of D-glucose on 5-aminolevulinate synthase activity is relieved with the metabolism of D-glucose. A final extracellular 5-aminolevulinate concentration of 3.1 g/l can be reached by feeding with D-glucose 2.3.1.37 5-aminolevulinate synthase synthesis production of 5-aminolevulinate by recombinant Escherichia coli. Terrific broth medium results in significantly higher cell growth and 5-aminolevulinate production than Luria-Bertani medium. 5-Aminolevulinate production is significantly enhanced by the addition of succinate together with glycine in the medium. Maximal 5-aminolevulinate production of 2.5 g/l is observed upon the addition of D-glucose as an 5-aminolevulinate dehydratase inhibitor in the late-log culture phase and maintenance of a pH value of 6.5 2.3.1.39 [acyl-carrier-protein] S-malonyltransferase synthesis an effective metabolic engineering strategy for industrial polyunsaturated fatty acid production is provided 2.3.1.43 phosphatidylcholine-sterol O-acyltransferase synthesis baculovirus-mediated expression of LCAT in mammalian cells as a high-mannose glycoform suitable for deglycosylation by Endo H and its purification to homogeneity and characterization. Treatment of the protein with Endo H results in a recombinant protein product that retains its native form and is suitable for structural determination by X-ray crystallography 2.3.1.75 long-chain-alcohol O-fatty-acyltransferase synthesis expression of a fusion protein between Marinobacter hydrocarbonoclasticus wax synthase and Marinobacter adhaerens fatty acyl-CoA reductase in Nicotiana benthaminana. Compared to wild-type controls, transgenic plants show both in leaves and stems a significant increase in the total level of wax esters, being eight-fold at the whole plant level. The profiles of fatty acid methyl ester and fatty alcohol in wax esters are related, and C16 and C18 molecules constitute predominant forms. Strong transformants display certain developmental aberrations, such as stunted growth and chlorotic leaves and stems. These negative effects are associated with an accumulation of fatty alcohols 2.3.1.75 long-chain-alcohol O-fatty-acyltransferase synthesis synthesis of fatty acid ethyl esters in Saccharomyces cerevisiae by gene integration into chromosomal delta sequences using repetitive transformation, which results in 1-6 copies of the integration construct per genome. The corresponding fatty acid ethyl ester production increases up to 34 mg/L, an about sixfold increase compared to the equivalent plasmid-based producer. The cassette in the yeast genome is stably maintained in nonselective medium after deletion of RAD52. Additional overexprerssion of genes encoding endogenous acyl-CoA binding protein and a bacterial NADP+-dependent glyceraldehyde-3-phosphate dehydrogenase (gapN) leads to another 40% increase in fatty acid ethyl ester production 2.3.1.75 long-chain-alcohol O-fatty-acyltransferase synthesis the enzyme can be used for oleyl oleate production in plant seed oil 2.3.1.81 aminoglycoside 3-N-acetyltransferase synthesis development of a methodology that utilizes aminoglycoside acetyltransferase and unnatural acyl-CoA analogues for the chemoenzymatic generation of regioselectively N-acylatedaminoglycoside analogues. Aminoglycosides are broad-spectrum antibiotics commonly used for the treatment of serious bacterial infections 2.3.1.84 alcohol O-acetyltransferase synthesis engineering of Escherichia coli to produce biodiesel, which depends on the expression of Saccharomyces cerevisiae alcohol acetyltransferase ATF1, fatty acyl-ACP reductase from Synechococcus elongates, carboxylic acid reductase from Mycobacterium marinum, fatty acyl-CoA reductase from Marinobacter aquaeolei, aldehyde reductase from Escherichia coli, fatty acyl-CoA synthetase from Escherichia coli, and TesA, a truncated fatty acyl-ACP thioesterase from Escherichia coli 2.3.1.84 alcohol O-acetyltransferase synthesis overexpression of ATF1 in Chinese liquor yeast through precise and seamless insertion of PGK1 promoter. In the liquid fermentation of corn hydrolysate, both mRNA level and alcohol acetyltransferase activity of ATF1 in mutant are pronounced higher than the parental strain. Productivity of ethyl acetate increases from 25.04 to 78.76 mg/l 2.3.1.84 alcohol O-acetyltransferase synthesis production of acetate esters by a mutant alpha5 strain overexpressing ATF1 with aminotransferase BAT2 deletion is 1353.4 mg/l, which is 43.16fold higher than that by the original strain. Isoform ATF1 overexpression with BAT2 deletion can further increase the ratio of ester to higher alcohol to a greater extent than isoform ATF2 overexpression with BAT2 deletion 2.3.1.84 alcohol O-acetyltransferase synthesis production of acetate esters by a mutant alpha5 strain overexpressing ATF1 with aminotransferase BAT2 deletion is 73.4 mg/l, which is 2.34fold higher than that by the original strain. Isoform ATF1 overexpression with BAT2 deletion can further increase the ratio of ester to higher alcohol to a greater extent than isoform ATF2 overexpression with BAT2 deletion 2.3.1.86 fatty-acyl-CoA synthase system synthesis design of a synthetic route consisting of two engineered FAS modules, module 1 optimized to produce octanoyl-CoA, and module 2 to nonreductively elongate this intermediate, yielding 6-heptyl-4-hydroxypyran-2-one 2.3.1.86 fatty-acyl-CoA synthase system synthesis expression of heterologous cytochrome P450 enzyme in combination with their cognate reductase for omega-hydroxylation of octanoic acid in a yeast strain, whose fatty acid synthase is engineered for octanoic acid production, results in de novo biosynthesis of 8-hydroxyoctanoic acid up to 3 mg/l. Cytochromes P450 activities are limiting 8-hydroxyoctanoic acid synthesis. The hydroxylation of both externally added and intracellularly produced octanoic acid is strongly dependent on the carbon source used, with ethanol being preferred 2.3.1.86 fatty-acyl-CoA synthase system synthesis short-chain acyl-CoA producing yeast Fas1 mutant R1834K/Fas2 fatty acid synthase variant is expressed together with carboxylic acid reductase from Mycobacterium marinum and phosphopantetheinyl transferase Sfp from Bacillus subtilis in a Saccharomyces cerevisiae DELTAfas1 DELTAfas2 DELTAfaa2 mutant strain. The synthesized octanoyl-CoA is endogenously converted to 1-octanol up to a titer of 26.0 mg/l in a 72-h fermentation. When octanoic acid is supplied externally to the yeast cells, it can be efficiently converted to 1-octanol. Additional overexpression of aldehyde reductase Ahr from Escherichia coli nearly completely prevents accumulation of octanoic acid and increases 1-octanol titers up to 49.5 mg/l. 1-octanol acts inhibitive before secretion 2.3.1.87 aralkylamine N-acetyltransferase synthesis recombinant Escherichia coli that expresses sheep SNAT with rice caffeic acid O-methyltransferase produces up to 1.46 mg/l meatonin in culture medium in response to 1 mM serotonin 2.3.1.94 6-deoxyerythronolide-B synthase synthesis the deoxyerythronolide B synthase genes are functionally expressed in Bacillus subtilis when the native eryAI–III operon is separated into three individual expression cassettes with optimized ribosomal binding sites. A synthesis of 6-deoxyerythronolide B can be detected by using the acetoininducible acoA promoter and a fed-batch simulating EnBase-cultivation strategy. Bacillus subtilis is capable of the secretion of 6-deoxyerythronolide B into the medium. The deletion of the prpBD operon of Bacillus subtilis, responsible for propionyl-CoA utilization, results in a significant increase of the 6-deoxyerythronolide B product yield when exogenous propionate is provided 2.3.1.95 trihydroxystilbene synthase synthesis synthesis of resveratrol from 4-coumaric acid can be achieved after codon-optimization for expression in Escherichia coli and coexpression with cinnamate/4-coumarate:coenzyme A ligase. Additional expression of resvertrol O-methyltransferase results in production of pinostilbene and pterostilbene 2.3.1.95 trihydroxystilbene synthase synthesis synthesis of resveratrol from 4-coumaric acid upon coexpression with cinnamate/4-coumarate:coenzyme A ligase in Escherichia coli 2.3.1.95 trihydroxystilbene synthase synthesis expression of stilbene synthase from Vitis vinifera and/or the transcription factor MYB12 from Arabidopsis thaliana in tobacco hairy root cultures. The transgenic cultures are able to biosynthesize stilbenes, achieving a production of 40 microg/L of trans-resveratrol, which is partially metabolized into trans-piceatannol and trans-pterostilbene (up to 2.2 microg/l and 86.4 microg/l, respectively), as well as its glucoside piceid (up to 339.7 microg/l) 2.3.1.95 trihydroxystilbene synthase synthesis in Silybum marinum cell cultures stably transformed with Vitis vinifera stilbene synthase 3, the epression of the transgene leads to extracellular trans-resveratrol accumulation at the level of milligrams per litre under elicitation. Resveratrol synthesis occurs at the expense of coniferyl alcohol 2.3.1.97 glycylpeptide N-tetradecanoyltransferase synthesis recombinant expression of N-myristoylated proteins in Escherichia coli can be achieved by co-expressing N-myristoyltransferase and supplementing the growth medium with myristic acid. Undesired incorporation of the 12-carbon fatty acid lauric acid can also occur, leading to heterogeneous samples. A strategy for obtaining lauryl-free samples of myristoylated proteins in both rich and minimal media 2.3.1.140 rosmarinate synthase synthesis expression of rosmarinic acid synthase from Coleus blumei, 4-coumarate: CoA ligase from Arabidopsis thaliana, 4-hydroxyphenyllactate 3-hydroxylase from Escherichia coli and D-lactate dehydrogenase from Lactobacillus pentosus in an L-tyrosine over-producing Escherichia coli strain leads to a yield of rosmarinic acid of 130 mg/l. In addition, intermediate caffeoyl-phenyllactate is produced at about 55 mg/l 2.3.1.140 rosmarinate synthase synthesis production of rosmarinic acid analogues in Escherichia coli by expression of 4-coumarate: CoA ligase from Arabidopsis thaliana and rosmarinic acid synthase from Coleus blumei. Incubation of the recombinant strain with exogenously supplied phenyllactic acid and analogues, and coumaric acid and analogues as donor substrates leads to production of 18 compounds, including unnatural analogues 2.3.1.144 anthranilate N-benzoyltransferase synthesis by exploiting the substrate flexibility of both 4-coumaroyl:CoA ligase 4CL5 from Arabidopsis thaliana and HCBT, rapid biosynthesis of more than 160 cinnamoyl, dihydrocinnamoyl, and benzoyl anthranilates in yeast can be achieved upon feeding with both natural and non-natural cinnamates, dihydrocinnamates, benzoates, and anthranilates 2.3.1.144 anthranilate N-benzoyltransferase synthesis coexpression of HCBT and 4-coumarate:CoA 4CL1 ligase from Nicotiana tabacum in the Escherichia coli anthranilate-accumulating strain W3110 trpD9923 allows the production of Avn D [N-(4'-hydroxycinnamoyl)-anthranilic acid] and Avn F [N-(3',4'-dihydroxycinnamoyl)-anthranilic acid] upon feeding with p-coumarate and caffeate, respectively. Additional expression of a tyrosine ammonia-lyase from Rhodotorula glutinis (TAL) leads to the conversion of endogenous tyrosine into p-coumarate and results in the production of Avn D [N-(4'-hydroxycinnamoyl)-anthranilic acid] from glucose. A 135fold improvement in Avn D titer is achieved by boosting tyrosine production. Expression of either the p-coumarate 3-hydroxylase Sam5 from Saccharothrix espanensis or the hydroxylase complex HpaBC from Escherichia coli results in the endogenous production of caffeate and biosynthesis of Avn F [N-(3',4'-dihydroxycinnamoyl)-anthranilic acid] 2.3.1.150 Salutaridinol 7-O-acetyltransferase synthesis potential application in biotechnological production of morphine alkaloids 2.3.1.150 Salutaridinol 7-O-acetyltransferase synthesis coexpression of Papaver somniferum enzymes salutaridine synthase, salutaridine reductase and salutaridinol in Saccharomyces cerevisiae allows for productive spontaneous rearrangement of salutaridinol-7-O-acetate and synthesis of thebaine from (R)-reticuline. A 7-gene pathway for the production of codeine and morphine from (R)-reticuline can be established. Yeast cell feeding assays using (R)-reticuline, salutaridine or codeine as substrates show that all enzymes are functionally coexpressed and that activity of salutaridine reductase and codeine-O-demethylase likely limit flux to morphine synthesis 2.3.1.156 phloroisovalerophenone synthase synthesis total enzymatic synthesis of acylphloroglucinol glucosides is achieved by co-incubation of recombinant dual functional chalcone/valerophenone synthase and UGT71K3 proteins with essential coenzyme A esters and UDP-glucose. Proteins UGT71K3a/b catalyze the glucosylation of diverse hydroxycoumarins, naphthols and flavonoids, enzymatically synthesized acylphloroglucinol aglycones and pelargonidin 2.3.1.158 phospholipid:diacylglycerol acyltransferase synthesis the enzyme is useful for production of hydroxylated fatty acids and furtheron polyesters, biodiesel, and lubricants in transgenic plants expressing PDAT thereby avoiding the toxicity of castor bean seeds 2.3.1.164 isopenicillin-N N-acyltransferase synthesis strategy to improve penicillin biosynthesis by Penicillium chrysogenum by reducing reactive and toxic metabolic byproducts, 2-oxoaldehydes. This is achieved by overexpressing the genes encoding glyoxalase I and II, which results in a 10% increase in penicillin titers relative to the control strain. The protein levels of isopenicillin N synthase and isopenicillin N acyltransferase are increased in the glyoxalase transformants, whereas their transcript levels remains unaltered 2.3.1.165 6-methylsalicylic-acid synthase synthesis biotechnological de novo production of m-cresol from sugar in complex yeast extract-peptone medium with the yeast Saccharomyces cerevisiae. A heterologous pathway based on the decarboxylation of the polyketide 6-methylsalicylic acid is introduced into a CEN.PK yeast strain. Overexpression of codon-optimized 6-methylsalicylic acid synthase from Penicillium patulum together with activating phosphopantetheinyl transferase npgA from Aspergillus nidulans results in up to 367 mg/l 6-methylsalicylic acid production. Additional genomic integration of the genes have a strongly promoting effect and 6-methylsalicylic acid titers reach more than 2 g/l. Simultaneous expression of 6-methylsalicylic acid decarboxylase patG from Aspergillus clavatus leads to the complete conversion of 6-methylsalicylic acid and production of up to 589 mg/L m-cresol 2.3.1.165 6-methylsalicylic-acid synthase synthesis Corynebacterium glutamicum is used as a host for polyketide production. 6-Methylsalicylic acid synthase ChlB1 from Streptomyces antibioticus is introduced into this bacterium (ChlB1Sa). Challenges related to protein folding can be overcome by translation fusion of ChlB1Sa to the C-terminus of the maltose-binding protein MalE from Escherichia coli. ChlB1Sa is also active in the absence of a heterologous 4'-phosphopantetheinyl transferase, which leads to the discovery that the endogenous 4'-phosphopantetheinyl transferases PptACg of Corynebacterium glutamicum can also activate ChlB1Sa. 6-Methylsalicylic acid is an important building block in the biosynthesis of several antibiotics, including chlorothricin, maduropeptin, pactamycin, and polyketomycin 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis bioengineered Escherichia coli cells synthesize nonnatural 10-deacetyl-10-acylbaccatin III derivatives as potential precursors of second-generation Taxol derivatives using the enzyme in concert with acyl-CoA transferases, determination of optimal alkanoic acid concentration for in vivo production of baccatin III analogues, overview 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis synthesis of baccatin III in Escherichia coli producing endogenous acetyl-CoA and overexpressing recombinant 10-deacetylbaccatin III 10-O-acetyltransferase. Use of enzyme to couple unnatural acyl-CoA analogues various amino and/or hydroxyl acceptors 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis transgenic Flammulina velutipes expressing the DBAT gene is able to produce baccatin III, an advanced precursor of paclitaxel 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis key enzyme in the biosynthesis of the anticancer drug paclitaxel, which catalyses the formation of baccatin III from 10-deacetylbaccatin III 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis mutant enzyme G38R/F301V with a catalytic efficiency approximately six times higher than that of the wild-type is combined with a beta-xylosidase to obtain an in vitro one-pot conversion of 7-beta-xylosyl-10-deacetyltaxol to Taxol yielding 0.64 mg/mlx02taxol in 50 ml at 15 h. This approach represents a promising environmentally friendly alternative for Taxol production from an abundant analogue 2.3.1.167 10-deacetylbaccatin III 10-O-acetyltransferase synthesis paclitaxel is a type of broad-spectrum anticancer drug in short supply. The price of acetyl-CoA, which is the acetyl group donor for the enzymatic synthesis of the intermediate, baccatin III, is the bottleneck of the mass production of paclitaxel. The study reports that N-acetyl-D-glucosamineas an acetyl group donor can substantially reduce the cost of production 2.3.1.168 dihydrolipoyllysine-residue (2-methylpropanoyl)transferase synthesis functional expression of the branched chain alpha-keto acid dehydrogenase complex in Escherichia coli by independent and selective optimization of individual subunit genes of the complex. Codon optimization significantly improves the expression of complex component proteins BkdH and LpdA1 but expression of dehydrogenase E1 alpha subunit BkdF and dehydrogenase E1 beta subunit BkdG depends on coexpression of the BkdH gene. The optimized branched chain alpha-keto acid dehydrogenase complex supplies sufficient short branched-chain acyl-CoA to synthesize phlorisovalerophenone 2.3.1.175 deacetylcephalosporin-C acetyltransferase synthesis production of acetylcephalosporin C by gene expression in Penicillium chrysogenum. Recombinant strains secrete significant amounts of deacetylcephalosporin C, but acetylcephalosporin C is not detected in culture broth. Even when accumulating intracellularly, acetylcephalosporin C is not found extracellularly 2.3.1.180 beta-ketoacyl-[acyl-carrier-protein] synthase III synthesis the recombinant enzyme is useful for production of fatty acids by directed biosynthesis profiting of the flexible substrate specificity for acyl-CoAs of the enzyme, overview 2.3.1.190 acetoin dehydrogenase system synthesis engineering a budC acoABCD double mutant, which has lost 2,3-butanediol dehydrogenase activity and the acetoin dehydrogenase system for R-acetoin production. Optimized conditions include aerobic batch culture and mildly acidic conditions. 62.3 g per l R-acetoin can be produced by budC and acoABCD double mutants in 57 h culture, with an optical purity of 98.0%, and a substrate conversion ratio of 28.7% 2.3.1.194 acetoacetyl-CoA synthase synthesis nphT7 can be used to significantly increase the concentration of acetoacetyl-CoA in cells, eventually leading to the production of useful terpenoids and poly-3-hydroxybutyrate 2.3.1.194 acetoacetyl-CoA synthase synthesis usage of the enzyme for recombinant high-level production of poly(3-hydroxybutyrate), P(3HB), in Escherichia coli and Corynebacterium glutamicum. P(3HB) is the natural aliphatic polyester that can be processed into a wide variety of consumer products, including plastics, films and fibers 2.3.1.201 UDP-2-acetamido-3-amino-2,3-dideoxy-glucuronate N-acetyltransferase synthesis the enzymes WbpA, WbpB, WbpE, WbpD and WbpI which act stepwise manner starting from UDP-GlcNAc can be combined in vitro to generate UDP-ManNAc(3NAc)A in a single reaction vessel 2.3.1.206 3,5,7-trioxododecanoyl-CoA synthase synthesis a synthetic pathway for the production of olivetolic acid in Escherichia coli is developed. Through combining OLA synthase and OLA cyclase expression with the required modules of a beta-oxidation reversal for hexanoyl-CoAgeneration, we demonstrate the in vivo synthesis of olivetolic acid from a single carbon source. The integration of additional auxiliary enzymes to increase hexanoyl-CoA and malonyl-CoA, along with evaluation of varying fermentation conditions enabled the synthesis of 80 mg/l olivetolic acid. Olivetolic acid has various pharmacological activities 2.3.1.211 bisdemethoxycurcumin synthase synthesis an engineered Escherichia coli strain expressing phenylalanine ammonia-lyase, 4-coumarate:CoA ligase 4CL1 from Arabidopsis thaliana and curcuminoid synthase from Oryza sativa leads to the production of dicinnamoylmethane at a high level of 0.36 g/l. Supplement of 2-fluoro-phenylalanine yields fluorinated dicinnamoylmethane derivatives, 6,6'-difluorodicinnamoylmethane and 6-fluoro-dicinnamoylmethane, of which the latter is a new curcuminoid 2.3.1.211 bisdemethoxycurcumin synthase synthesis an artificial pathway for curcuminoid production in Escherichia coli is constructed. Overexpression of curcuminoid synthase from Oryza sativa in Escherichia coli results in the production of the major curcuminoid, bisdemethoxycurcumin from p-coumaric acid. It is demonstrated that enhancement of the intracellular malonyl-CoA pool is essential for increasing the final production titer of bisdemethoxycurcumin. Expression of a recombinant pathway that allows the conversion of malonate to malonyl-CoA encoded by genes matB and matC results in a 25fold improvement of final bisdemethoxycurcumin titer 2.3.1.211 bisdemethoxycurcumin synthase synthesis engineering of a Pseudomonas putida strain to produce bisdemethoxycurcumin 2.3.1.211 bisdemethoxycurcumin synthase synthesis production of curcuminoids in engineered Escherichia coli. PAL (phenylalanine ammonia lyase) or TAL (tyrosine ammonia lyase), along with Os4CL (p-coumaroyl-CoA ligase) and CUS (curcumin synthase) genes, are introduced into Escherichia coli, and each strain produces dicinnamoylmethane or bisdemethoxycurcumin, respectively. In order to increase the production of curcuminoids in Escherichia coli, the shikimic acid biosynthesis pathway, which increases the substrates for curcuminoid biosynthesis, is engineered. Using the engineered strains, the production of bisdemethoxycurcumin increases from 0.32 to 4.63 mg/l, and that of dicinnamoylmethane from 1.24 to 6.95 mg/l 2.3.1.211 bisdemethoxycurcumin synthase synthesis production of curcuminoids in engineered Escherichia coli. Two curcuminoids (dicinnamoylmethane and bisdemethoxycurcumin) are synthesized from glucose in Escherichia coli. PAL (phenylalanine ammonia lyase) or TAL (tyrosine ammonia lyase), along with Os4CL (p-coumaroyl-CoA ligase) and CUS genes, are introduced into Escherichia coli, and each strain produces dicinnamoylmethane or bisdemethoxycurcumin, respectively. In order to increase the production of curcuminoids in Escherichia coli, the shikimic acid biosynthesis pathway, which increases the substrates for curcuminoid biosynthesis, is engineered. Using the engineered strains, the production of bisdemethoxycurcumin increases from 0.32 to 4.63 mg/l, and that of dicinnamoylmethane from 1.24 to 6.95 mg/l 2.3.1.212 benzalacetone synthase synthesis possibility of peparation of tetramic acids from various amino acids through the enzyme 2.3.1.212 benzalacetone synthase synthesis a de novo pathway for the production of raspberry ketone is assembled using four heterologous genes, encoding phenylalanine/tyrosine ammonia lyase, cinnamate-4-hydroxlase, coumarate-CoA ligase and benzalacetone synthase, in an industrial strain of Saccharomyces cerevisiae. It is possible to produce sensorially-relevant quantities of raspberry ketone in the industrial heterologous host 2.3.1.212 benzalacetone synthase synthesis a heterologous pathway to produce raspberry ketone from p-coumaric acid, including 4-coumarate:CoA ligase (4CL), benzalacetone synthase (BAS), and raspberry ketone/zingerone synthase (RZS1) from plants, is assembled in Escherichia coli. Raspberry ketone is an important ingredient in the flavor and fragrance industries. Engineered strain CZ-8 which cooverexpresses At4CL1, RpBAS, and RiRZS1 achieves levels of 90.97 mg/l of raspberry ketone 2.3.1.217 curcumin synthase synthesis construction of a fusion protein diketide-CoA synthase::curcumin synthase. Comparing to CURS, the fusion protein shows similar substrate specificities and catalytic potentials to catalyze the formation of various curcuminoids, with increased yields of curcuminoids 2.3.1.217 curcumin synthase synthesis production of curcuminoids using an engineered artificial pathway in Escherichia coli. Expression of Arabidopsis thaliana 4-coumaroyl-CoA ligase and Curcuma longa diketide-CoA synthase (DCS) and curcumin synthase (CURS1) leads to synthesis of 70 mg/l of curcumin from ferulic acid. Bisdemethoxycurcumin and demethoxycurcumin are produced, but in lower concentrations, by feeding 4-coumaric acid or a mixture of 4-coumaric acid and ferulic acid, respectively. To produce caffeic acid, tyrosine ammonia lyase from Rhodotorula glutinis and 4-coumarate 3-hydroxylase from Saccharothrix espanaensis are used. Caffeoyl-CoA 3-O-methyltransferase from Medicago sativa converts caffeoyl-CoA to feruloyl-CoA. Using caffeic acid, 4-coumaric acid or tyrosine as a substrate, 3.9, 0.3, and 0.2 mg/l of curcumin are produced, respectively 2.3.1.217 curcumin synthase synthesis a curcuminoid producing unnatural fusion protein diketide-CoA synthase:curcumin synthase is constructed. The fusion protein may contribute to further understand the biosynthesis of curcuminoids in ginger but also be advantage to further manipulate the biosynthesis of curcuminoid analogs, particularly including tetrahydrobisdemethoxycurcumin (THBDC) and various dihydrocurcuminoid derivatives in microorganisms 2.3.1.217 curcumin synthase synthesis biosynthetic pathway of p-coumaric acid, caffeic acid and curcumin in Escherichia coli can be triggered by using heat shock promoters, suggesting its potential for the development of new industrial bioprocesses or even new bacterial therapies. p-Coumaric acid is successfully produced from tyrosine and caffeic acid produced either from tyrosine or p-coumaric acid using tyrosine ammonia lyase (TAL) from Rhodotorula glutinis, 4-coumarate 3-hydroxylase (C3H) from Saccharothrix espanaensis or cytochrome P450 CYP199A2 from Rhodopseudomonas palustris. The highest p-coumaric acid production obtained is 2.5 mM. Caffeic acid production reaches 0.370 mM. 0.017 mM cumin is produced using 4-coumarate-CoA ligase (4CL1) from Arabidopsis thaliana, diketide-CoA synthase (DCS) and curcumin synthase 1 (CURS1) from Curcuma longa 2.3.1.217 curcumin synthase synthesis design, construction and optimization of a heterologous pathway to produce curcuminoids in Escherichia coli. This pathway involves six enzymes, tyrosine ammonia lyase (TAL), 4-coumarate 3-hydroxylase (C3H), caffeic acid O-methyltransferase (COMT), 4-coumarate-CoA ligase (4CL), diketide-CoA synthase (DCS), and curcumin synthase (CURS1). Curcumin production is enhanced and reachs 43.2 mM, corresponding to an improvement of 160% comparing to mono-culture system 2.3.1.217 curcumin synthase synthesis production of curcuminoids in engineered Escherichia coli. Two curcuminoids (dicinnamoylmethane and bisdemethoxycurcumin) are synthesized from glucose in Escherichia coli. PAL (phenylalanine ammonia lyase) or TAL (tyrosine ammonia lyase), along with Os4CL (p-coumaroyl-CoA ligase) and CUS (curcumin synthase) genes, are introduced into Escherichia coli, and each strain produces dicinnamoylmethane or bisdemethoxycurcumin, respectively. In order to increase the production of curcuminoids in Escherichia coli, the shikimic acid biosynthesis pathway, which increases the substrates for curcuminoid biosynthesis, is engineered. Using the engineered strains, the production of bisdemethoxycurcumin increases from 0.32 to 4.63 mg/l, and that of dicinnamoylmethane from 1.24 to 6.95 mg/l 2.3.1.218 phenylpropanoylacetyl-CoA synthase synthesis a curcuminoid producing unnatural fusion protein diketide-CoA synthase:curcumin synthase is constructed. The fusion protein may contribute to further understand the biosynthesis of curcuminoids in ginger but also be advantage to further manipulate the biosynthesis of curcuminoid analogs, particularly including tetrahydrobisdemethoxycurcumin (THBDC) and various dihydrocurcuminoid derivatives in microorganisms 2.3.1.218 phenylpropanoylacetyl-CoA synthase synthesis design, construction and optimization of a heterologous pathway to produce curcuminoids in Escherichia coli. This pathway involves six enzymes, tyrosine ammonia lyase (TAL), 4-coumarate 3-hydroxylase (C3H), caffeic acid O-methyltransferase (COMT), 4-coumarate-CoA ligase (4CL), diketide-CoA synthase (DCS), and curcumin synthase (CURS1). Curcumin production is enhanced and reachs 43.2 mM, corresponding to an improvement of 160% comparing to mono-culture system 2.3.1.219 demethoxycurcumin synthase synthesis a curcuminoid producing unnatural fusion protein diketide-CoA synthase:curcumin synthase is constructed. The fusion protein may contribute to further understand the biosynthesis of curcuminoids in ginger but also be advantage to further manipulate the biosynthesis of curcuminoid analogs, particularly including tetrahydrobisdemethoxycurcumin (THBDC) and various dihydrocurcuminoid derivatives in microorganisms 2.3.1.226 carboxymethylproline synthase synthesis carboxymethylproline synthases is a biocatalysts for preparing functionalised N-heterocycles in a diastereoselective fashion. The products can be converted into the respective bicyclic beta-lactams of potential application in the semisynthesis of stable beta-lactam antibiotics 2.3.1.226 carboxymethylproline synthase synthesis coupling of methylmalonyl-CoA epimerase, crotonyl-CoA carboxylase reductase and carboxymethylproline synthase mutants in a three-enzyme one-pot sequential synthesis of functionalised C-5 carboxyalkylprolines starting from crotonyl-CoA and carbon dioxide 2.3.1.226 carboxymethylproline synthase synthesis biocatalytic production of bicyclic beta-lactams with three contiguous chiral centres using engineered crotonases. Structurally guided substitutions of wild type carboxymethylproline synthases enable tuning of their C-N and C-C bond forming capacity to produce 5-carboxymethylproline derivatives substituted at C-4 and C-6, from amino acid aldehyde and malonyl-CoA derivatives. Use of tandem enzyme incubations comprising an engineered carboxymethylproline synthase and an alkylmalonyl-CoA forming enzyme (i.e. malonyl-CoA synthetase or crotonyl-CoA carboxylase reductase) can improve stereocontrol and expand the product range. Some of the prepared 4,6-disubstituted-5-carboxymethylproline derivatives are converted to bicyclic beta-lactams by carbapenam synthetase catalysis 2.3.1.229 4-coumaroyl-homoserine lactone synthase synthesis development of an artificial biosynthetic process for phenylacetyl-homoserine lactone analogs, including cinnamoyl-homoserine lactone, 4-coumaroyl-homoserine lactone, caffeoyl-homoserine lactone, and feruloyl-homoserine lactone, using coexpression of the codon-optimized synthase RpaI and 4-coumaroyl-CoA ligase 4CL2nt, EC 6.2.1.12, in Escherichia coli. De novo production of p-coumaroyl-homoserine lactone in Escherichia coli can be achieved by expression of the rpaI gene in addition to 4-coumaroyl-CoA biosynthetic genes. The yields for 4-coumaroyl-homoserine lactone reach 93 and 142 mg/l in the S-adenosyl-L-methionine and L-methionine feeding culture, respectively 2.3.1.229 4-coumaroyl-homoserine lactone synthase synthesis production of bacterial quorum sensing antagonists, caffeoyl- and Feruloyl-HSL, by an artificial biosynthetic pathway. An Escherichia coli system containing artificial biosynthetic pathways that yield phenylacetyl-homoserine lactones from simple carbon sources. The artificial biosynthetic pathways contain the LuxI-type synthase gene (rpaI) in addition to caffeoyl-CoA and feruloyl-CoA biosynthetic genes, respectively. The yields for caffeoyl-phenylacetyl-homoserine lactone and feruloyl-phenylacetyl-homoserine lactone are 97.1 and 65.2 mg/l, respectively, by tyrosine-overproducing Escherichia coli with a L-methionine feeding strategy 2.3.1.232 methanol O-anthraniloyltransferase synthesis three anthranilate derivatives, N-methylanthranilate, methyl anthranilate, and methyl N-methylanthranilate are synthesized using metabolically engineered stains of Escherichia coli. NMT encoding N-methyltransferase from Ruta graveolens, AMAT encoding anthraniloyl-coenzyme A (CoA):methanol acyltransferase from Vitis labrusca, and pqsA encoding anthranilate coenzyme A ligase from Pseudomonas aeruginosa are cloned and Eschetrichia coli strains harboring these genes were used to synthesize the three desired compounds. Escherichia coli mutants (metJ, trpD, tyrR mutants), which provide more anthranilate and/or S-adenosyl methionine, are used to increase the production of the synthesized compounds. 0.1853 mM N-methylanthranilate and 0.0952 mM methyl N-methylanthranilate are synthesized 2.3.1.233 1,3,6,8-tetrahydroxynaphthalene synthase synthesis heterologous expression of soceCHS1, bdsA, and bdsB from Sorangium cellulosum in Streptomyces coelicolor causes secretion of 1,8-dihydroxynaphthalene, a key precursor of dihydroxynaphthalene melanin, from Sorangium cellulosum 2.3.1.238 monacolin J acid methylbutanoate transferase synthesis the enzyme can be a useful biocatalyst for the synthesis of simvastatin and other statin analogs. It is an attractive enzyme for engineered biosynthesis of pharmaceutically important cholesterol-lowering drugs 2.3.1.239 10-deoxymethynolide synthase synthesis substrate engineering approaches to control the catalytic cycle of a full polykeitde synthase module harboring multiple domains. Using alternatively activated native hexaketide substrates, product formation may be controled with greater than 10:1 selectivity for either full module catalysis, leading to a 14-membered macrolactone, or direct cyclization to a 12-membered ring 2.3.1.239 10-deoxymethynolide synthase synthesis versatile method for generating and identifying functional chimeric PKS enzymes for synthesizing custom macrolactones and macrolides. PKS genes from the pikromycin and erythromycin pathways are hybridized in Saccharomyces cerevisiae to generate hybrid libraries. Streptomyces venezuelae strains that expressed active chimeric enzymes with new functionality are capable of producing engineered macrolactones 2.3.1.240 narbonolide synthase synthesis substrate engineering approaches to control the catalytic cycle of a full polykeitde synthase module harboring multiple domains. Using alternatively activated native hexaketide substrates, product formation may be controled with greater than 10:1 selectivity for either full module catalysis, leading to a 14-membered macrolactone, or direct cyclization to a 12-membered ring 2.3.1.240 narbonolide synthase synthesis versatile method for generating and identifying functional chimeric PKS enzymes for synthesizing custom macrolactones and macrolides. PKS genes from the pikromycin and erythromycin pathways are hybridized in Saccharomyces cerevisiae to generate hybrid libraries. Streptomyces venezuelae strains that expressed active chimeric enzymes with new functionality are capable of producing engineered macrolactones 2.3.1.244 2-methylbutanoate polyketide synthase synthesis using a multivector expression system in Saccharomyces cerevisiae, two natural statins from two fungal species, i.e., lovastatin from Aspergillus terreus and FR901512 from Xylaria grammica are reconstituted, but also new statin structures by mixing their genes 2.3.1.251 lipid IVA palmitoyltransferase synthesis deacylated lipid A, deacylated and palmitoylated lipid A, and palmitoylated lipid A species are generated in Escherichia coli cells heterologously expressing salmonellae lipid A 3-O-deacylase PagL and/or PagP, and then purified by sequential thin-layer chromatography. The purified lipid A species show m/z values that correspond to single lipid A species on mass spectrometry analysis. The modified lipid A species show reduced ability to induce cellular signaling through Toll-like receptor 4 2.3.1.253 phloroglucinol synthase synthesis the enzyme can be used for production of important industrial chemicals from acetate in in vitro systems 2.3.1.264 beta-lysine N6-acetyltransferase synthesis expression of the N-acetyltransferase YodP and L-lysine 2,3-aminomutase kamA genes from an isopropyl beta-D-1-thiogalactopyranoside-inducible promoter on a plasmid in Bacillus subtilis, leads to synthesis of 0.28 micromol/mg protein of Nepsilon-acetyl-beta-lysine 2.3.1.268 ethanol O-acetyltransferase synthesis Eat1-catalyzed ethyl acetate production occurs in yeast mitochondria 2.3.1.282 phenolphthiocerol/phthiocerol/phthiodiolone dimycocerosyl transferase synthesis use of PapA5 and a mutant mycocerosic acid synthase for synthesis of nonmethylated variants of mycocerosate esters 2.3.1.285 (13S,14R)-1,13-dihydroxy-N-methylcanadine 13-O-acetyltransferase synthesis de novo production of noscapine in Saccharomyces cerevisiae, through the reconstruction of a biosynthetic pathway comprising over 30 enzymes from plants, bacteria, mammals, and yeast itself. Optimization of conditions leads to an over 18,000fold improvement from initial noscapine titers to about2.2 mg/l. By feeding modified tyrosine derivatives to the optimized noscapine-producing strain, microbial production of halogenated benzylisoquinoline alkaloids is possible 2.3.1.300 branched-chain beta-ketoacyl-[acyl-carrier-protein] synthase synthesis replacement of the acetyl-CoA-specific Escherichia coli FabH with branched-chain-acyl-CoA-specific Bacillus subtilis FabH1 increases the synthesis of branched-chain fatty acids, resulting in a significant enhancement in branched-chain fatty acids titer compared to a strain containing both acetyl-CoA- and branched-chain-acyl-CoA-specific FabHs. The titer of branched-chain fatty acids reaches 19.4 mg/L. The composition of branched-chain fatty acids can be tuned by engineering the upstream pathway to control the supply of different branched-chain acyl-CoAs, leading to the production either even-chain-iso-, odd-chain-iso-, or odd-chain-anteiso-branched-chain fatty acids separately 2.3.1.300 branched-chain beta-ketoacyl-[acyl-carrier-protein] synthase synthesis replacement of the acetyl-CoA-specific Escherichia coli FabH with branched-chain-acyl-CoA-specific Bacillus subtilis FabH2 increases the synthesis of branched-chain fatty acids, resulting in a significant enhancement in branched-chain fatty acids titer compared to a strain containing both acetyl-CoA- and branched-chain-acyl-CoA-specific FabHs. The titer of branched-chain fatty acids reaches 17.5 mg/L. The composition of branched-chain fatty acids can be tuned by engineering the upstream pathway to control the supply of different branched-chain acyl-CoAs, leading to the production either even-chain-iso-, odd-chain-iso-, or odd-chain-anteiso-branched-chain fatty acids separately 2.3.1.300 branched-chain beta-ketoacyl-[acyl-carrier-protein] synthase synthesis replacement of the acetyl-CoA-specific Escherichia coli FabH with branched-chain-acyl-CoA-specific Staphylococcus aureus FabH increases the synthesis of branched-chain fatty acids, resulting in a significant enhancement in branched-chain fatty acids titer compared to a strain containing both acetyl-CoA- and branched-chain-acyl-CoA-specific FabHs. The titer of branched-chain fatty acids reaches 40.3 mg/L. The composition of branched-chain fatty acids can be tuned by engineering the upstream pathway to control the supply of different branched-chain acyl-CoAs, leading to the production either even-chain-iso-, odd-chain-iso-, or odd-chain-anteiso-branched-chain fatty acids separately 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis usage of the enzyme for recombinant high-level production of poly(3-hydroxybutyrate), P(3HB), in Escherichia coli and Corynebacterium glutamicum. P(3HB) is the natural aliphatic polyester that can be processed into a wide variety of consumer products, including plastics, films and fibers 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis a Cupriavidus necator polyhydroxyalkanoate-negative transformant harboring the polyhydroxyalkanoate synthase gene from bacterium SC8 is able to accumulate 55 weight% of polyhydroxyalkanoate of the cell dry weight when crude palm kernel oil is used as the carbon source. It produces 14 weight% of polyhydroxyalkanoate when 10 g per l of fructose is used 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis engineering of an in vivo polylactic acid biosynthesis system in Escherichia coli to synthesize 2-hydroxybutyrate-containing polyhydroxyalkanoate. Propionyl-CoA is used as precursor for 2-ketobutyrate that is converted into 2-hydroxybutyrate-CoA by the sequential actions of Lactococcus lactis (D)-2-hydroxybutyrate dehydrogenase (PanE) and PctCp and then 2-hydroxybutyrate-CoA is polymerized by an engineered PhaC1 from Pseudomonas sp. (mutant E130D/S325T/S477G/Q481K). The recombinant Escherichia coli expressing the system can be grown to 0.66 g/l and successfully produces P(70 mol% 3-hydroxybutyrate-co-18 mol% 3-hydroxybutyrate-co-12 mol% lactate) up to the polyhydroxyalkanoate content of 66 weight% from 20 g/l of glucose, 2 g/l of 3-hydroxybutyrate and 1 g/l of sodium propionate 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis enzyme shows both polyhydroxyalkanoate polymerization and alcoholysis activities, unsing various alcohols other than ethanol for alcoholysis.. Through the use of bifunctional compounds for alcoholysis, the polyhydroxyalkanoate carboxy terminus can be modified with thiol, alkynyl, hydroxy, and benzyl groups, resulting in the functionalization of the polyhydroxyalkanoate carboxy terminus 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis expansion of the substrate specificity and enhancment of biosynthesis of polyhydroxyalkanoate by site-specific mutagenesis. Mutated PhaC1s are coexpressed with beta-ketothiolase and acetoacetyl-CoA reductase to supply sufficient short-chain length (R)-3-hydroxyacyl-CoA as a substrate. Mutation L484V remarkably enhances the monomer ratio of (R)-3-hydroxybutyrate in a polyhydroxyalkanoate accumulation experiment. Val is the most favorable amino acid for incorporating (R)-3-hydroxybutyrate unit synthesis. A single mutation at Q481M, S482G and A547V obviously increases polyhydroxyalkanoate yields. Q481M and S482G enhance the (R)-3-hydroxyhexanoate monomer composition in the polyhydroxyalkanoate accumulation 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis heterologous expression in Escherichia coli with simultaneous overexpression of chaperone proteins. Compared to expression of synthase alone (14.55 mg per liter), coexpression with chaperones results in the production of larger total quantities of enzyme, including a larger proportion in the soluble fraction. The largest increase is seen when the GroEL/GroES system is coexpressed, resulting in approximately 6fold greater enzyme yields (82.37 mg per liter) than in the absence of coexpressed chaperones. The specific activity of the purified enzyme is unaffected by coexpression with chaperones 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis mutant A510S is able to synthesize a lactate-3-hydroxybutanoate copolymer containing 7 mol% lactate and with a averge molecular weight of 320000 Da. The polymer contains a high ratio of an LA-LA-LA triad sequence 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis biodegradable bio-based polyhydroxyalkanoate (PHA) is gaining much attention as a promising biomaterial that can replace some conventional petroleum-based plastics especially the single use plastics 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis biodegradable bio-based polyhydroxyalkanoate (PHA) is gaining much attention as a promising biomaterial that can replace some conventional petroleum-based plastics especially the single use plastics. The most studied class I PhaC from Cupriavidus necator (PhaCCn) is often used as a study model to increase its ability to incorporate medium-chain length (MCL) monomers into PHA 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis biodegradable bio-based polyhydroxyalkanoate (PHA) is gaining much attention as a promising biomaterial that could replace some conventional petroleum-based plastics especially the single use plastics. Class II PhaC1 from Pseudomonas sp. 61-3 (PhaC1Ps) is useful in terms of its broad substrate specificity, and engineering of PhaC1Ps successfully increases its short-chain-length (SCL) incorporation into PHA 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis poly(3-hydroxyalkanoates) (PHAs) such as poly(3-hydroxybutyrate) (PHB) are suitable biobased and biodegradable candidates to replace petroleum-based nondegradable plastics. Enzyme PhaEC can be used in its biocatalytic production, PHB production in clostridia, method development and evaluation, overview. The successful transfer and expression of phaJ and phaEC in autotrophic gas-fermenting clostridia leading to PHB formation now opens the possibility to establish an economically viable route to biodegradable plastics from waste and greenhouse gases, as the gas fermentation technology has meanwhile matured and is already performed at industrial scale. A novel metabolic pathway leading to 3-hydroxybutyrate has also been engineered that will allow similarly better economic production of this platform chemical 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis replacement of conventional plastics by bioplastics. Polyhydroxyalkanoates (PHA) are bio-polyesters accumulated in cells by a wide range of bacteria. Polyhydroxyalkanoates production from synthetic waste using Pseudomonas palleronii polyhydroxyalkanoate synthase enzyme activity 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis replacement of conventional plastics by bioplastics. Polyhydroxyalkanoates (PHA) are bio-polyesters accumulated in cells by a wide range of bacteria. Polyhydroxyalkanoates production from synthetic waste using Pseudomonas pseudoflava polyhydroxyalkanoate synthase enzyme activity 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis the enzyme mutant S326T/Q482K might be useful for production of distinct polyhydroxyalkanoate copolymers with improved material properties 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis the PHA synthase is suitable for the biosynthesis of PHAs that can be used in various biomedical applications due to its ability to incorporate the lipase-degradable monomer sequences of 4-hydroxybutyrate (4HB) and 5-hydroxyvalerate (5HV) 2.3.1.304 poly[(R)-3-hydroxyalkanoate] polymerase synthesis the enzyme can be used for polyhydroxyalkanoate production from synthetic wastewater 2.3.2.2 gamma-glutamyltransferase synthesis synthesis of theanine from 5-L-glutamyl-4-nitroanilide and ethylamine 2.3.2.2 gamma-glutamyltransferase synthesis the enzyme can catalyze the transfer of L-glutamine to Gly-Gly to synthesize Gln-Gly-Gly, which is promising for the synthesis of valuable gamma-glutamyl peptides 2.3.2.2 gamma-glutamyltransferase synthesis high potentiality of BlGGT-catalyzed reaction in the synthesis of gamma-glutamyl compounds in variety of biotechnological and pharmaceutical applications 2.3.2.2 gamma-glutamyltransferase synthesis in pharmaceutical and medical sectors GGT enzyme can be exploited in the synthesis of pro-drugs and therapeutic compounds such as gamma-glutamyl DOPA, gamma-D-glutamyl-L-tryptophan, and gamma-glutamyl-taurine 2.3.2.2 gamma-glutamyltransferase synthesis the recombinant extracellular enzyme is used for enzymatic synthesis of L-theanine, which is performed at optimal conditions: 20 mM L-Gln, 100 mM ethylamine-HCl, 0.5 U/ml BaGGT, incubated at 40°C for 6 h, 200 rpm 2.3.2.2 gamma-glutamyltransferase synthesis using recombinant Bacillus licheniformis gamma-glutamyltranspeptidase (BlGGT) in a straightforward procedure for the biocatalytic synthesis of gamma-glutamyl-phenylalanine (gamma-Glu-Phe), qualitative analysis of reaction products, mass spectrometry/nuclear magnetic resonance analyses 2.3.2.6 lysine/arginine leucyltransferase synthesis use of enzyme to link non-natural amino acids to the N termini of target proteins through the use of tRNAPhes aminoacylated with various types of non-natural amino acids 2.3.2.6 lysine/arginine leucyltransferase synthesis leucyl/phenylalanyl(L/F)-tRNA-protein transferase mediates aminoacyl transfer of a nonnatural amino acid to the N-terminus of peptides and proteins and the bioorthogonal reactive group can be converted to a variety of functional groups 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis selective introduction of aminated compounds into proteins 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis soy flour as a source of transglutaminase substrates to prepare hydrocolloid films together with pectin 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis high-yield expression of His-tagged enzyme in Escherichia coli and single-step purification protocol giving high specific activity 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis method for on-column activation of His-tagged enzyme by trypsin. About 89% of pro-MTG-His6 can be transferred to mature MTG-His6 with storage stabilization 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis microbial transglutaminase (MTG) is a practical tool to enzymatically form isopeptide bonds between peptide or protein substrates, crosslinking the side-chains of reactive glutamine and lysine residues is solidly rooted in food and textile processing. MTG-reactive glutamines can be readily introduced into a protein domain for fluorescent labeling, method evaluation, overview 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis TGase can be used for the development of site-specific derivatives of IFN alpha-2b possessing interesting antiviral and pharmacokinetic properties 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis the enzyme can be a useful biocatalyst for the synthesis of desirable bioactive molecules 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis the enzyme is used for biomaterial fabrication for tissue engineering, e.g. from gelatin, evaluation, overview 2.3.2.13 protein-glutamine gamma-glutamyltransferase synthesis the Kutzneria albida microbial transglutaminase enabling highly site-specific labeling of proteins. Its site specificity, favorable kinetics, ease of use, and cost-effective production render KalbTG an attractive tool for a broad range of applications, including production of therapeutic antibody-drug conjugates. The high activity and low molecular mass of KalbTG signifies a key advantage for mass production and enzymatic labeling purposes 2.3.2.20 cyclo(L-leucyl-L-phenylalanyl) synthase synthesis CDPSs are good candidates for the biological production of 2,5-diketopiperazines (2,5-DKPs) because their heterologous expression in Escherichia coli is easy to implement and leads up to high amounts of cyclodipeptides recovered in culture supernatants. CDPSs are often found within biosynthetic gene clusters containing diverse tailoring enzymes responsible for further chemical modifications of the produced cyclodipeptides 2.3.2.20 cyclo(L-leucyl-L-phenylalanyl) synthase synthesis cyclodipeptide synthases (CDPSs) use two aminoacyl-tRNAs to catalyze the formation of two peptide bonds leading to cyclodipeptides that can be further used for the synthesis of diketopiperazines 2.3.2.21 cyclo(L-tyrosyl-L-tyrosyl) synthase synthesis cyclodipeptide synthases (CDPSs) use two aminoacyl-tRNAs to catalyze the formation of two peptide bonds leading to cyclodipeptides that can be further used for the synthesis of diketopiperazines 2.3.2.22 cyclo(L-leucyl-L-leucyl) synthase synthesis cyclodipeptide synthases (CDPSs) use two aminoacyl-tRNAs to catalyze the formation of two peptide bonds leading to cyclodipeptides that can be further used for the synthesis of diketopiperazines 2.3.2.26 HECT-type E3 ubiquitin transferase synthesis construcution of a soluble HECT domain truncation of isoform WWP2 which is amendable for preparation scale expression in Escherichia coli. A relatively simple purification process achieves highly pure protein by employing immobilized metal-affinity chromatography followed by salting out, ion exchange chromatography and finally, size exclusion chromatography. Procedure allows to obtain about 60 mg/L of the soluble protein 2.3.3.8 ATP citrate synthase synthesis metabolic engineering of ATP-citrate lyase improves methylenesuccinic acid production in Aspergillus niger due to an increase in glycolytic flux 2.3.3.10 hydroxymethylglutaryl-CoA synthase synthesis use of crystal structure of enzyme in complex with inhibitor (E,E)-11-(3-hydroxymethyl-4-oxo-2-oxytanyl)-3,5,7-trimethyl-2,4-undecadienoic acid as starting point for structure-based design of inhibitors 2.3.3.14 homocitrate synthase synthesis novel glutarate biosynthetic pathway by incorporation of a +1 carbon chain extension pathway from 2-oxoglutarate in combination with 2-oxo acid decarboxylation pathway in Escherichia coli. Introduction of homocitrate synthase, homoaconitase and homoisocitrate dehydrogenase from Saccharomyces cerevisiae into Escherichia coli enables +1 carbon extension from 2-oxoglutarate to 2-oxoadipate, which is subsequently converted into glutarate by a promiscuous 2-oxo acid decarboxylase (KivD) and a succinate semialdehyde dehydrogenase (GabD). The recombinant Escherichia coli coexpressing all five genes produces 0.3 g/l glutarate from glucose. To further improve the titers, 2-oxoglutarate is rechanneled into carbon chain extension pathway via the clustered regularly interspersed palindromic repeats system mediated interference (CRISPRi) of essential genes sucA and sucB in tricarboxylic acid cycle. The final strain can produce 0.42 g/l glutarate, which is increased by 40% compared with the parental strain. Glutarate is one of the most potential building blocks for bioplastics 2.3.3.16 citrate synthase (unknown stereospecificity) synthesis involved in poly 3-hydroxybutyrate formation 2.3.3.16 citrate synthase (unknown stereospecificity) synthesis over-expression of citrate syntase gltA or phosphoenolpyruvate carboxylase ppc increases the maximum cell dry weight by 23% and 91% resp. No acetate excretion is detected at these increased cell densities 2.3.3.21 (R)-citramalate synthase synthesis the autrotrophic micro-organism may be engineered for robust butanol and propanol production from 2-ketobutyrate, which is an intermediate in the isoleucine biosynthesis pathway 2.3.3.21 (R)-citramalate synthase synthesis synthesis of citramalate at high yield by Escherichia coli overexpressing citramalate synthase. Citramalate is a chemical precursor to the industrially important methacrylic acid. Acetate is an undesirable by-product potentially formed from pyruvate and acetyl-CoA, the precursors of citramalate during aerobic growth of Escherichia coli. Gene deletions critical to reducing acetate accumulation during aerobic growth and citramalate production are ifdentified in metabolically engineered Escherichia coli strains. The key knockouts critical to minimizing acetate formation are identified as pta, ackA and poxB 2.3.3.21 (R)-citramalate synthase synthesis synthesis of citramalic acid from glycerol by metabolically engineered Escherichia coli 2.3.3.21 (R)-citramalate synthase synthesis in a fedbatch process an exponential feeding strategy using expression of CimA and the citrate synthase GltA F383M variant, over 60 g/l citramalate with a yield of 0.53 g citramalate/g glucose are generated in 132 hr 2.4.1.1 glycogen phosphorylase synthesis production of D-glucose-1-phosphate with yield of 47% w/w from a reaction containing 5% w/v soluble starch in 0.7 M potassium phosphate. Purification of D-glucose-1-phosphate by ethanol preciptitation 2.4.1.1 glycogen phosphorylase synthesis immobilized enzyme is used for the production of glucose-1-phosphate, a cytostatic compound used in cardiotherapy 2.4.1.1 glycogen phosphorylase synthesis maltodextrin phosphorylase from Pyrococcus furiosus is fused with the cellulose-binding domain of Clostridium cellulovorans serving as an aggregation module. After molecular cloning of the corresponding gene fusion construct and controlled expression in Escherichia coli BL21, 83% of total maltodextrin phosphorylase activity is displayed in active inclusion bodies. These active inclusion bodies are easily isolated by nonionic detergent treatment and directly used for maltodextrin conversion to alpha-D-glucose-1-phosphate in a repetitive batch mode. Only 10% of enzyme activity is lost after ten conversion cycles at optimum conditions 2.4.1.1 glycogen phosphorylase synthesis immobilization of partially purified enzyme using egg shell as solid support with a percentage retention of the enzyme on egg shell of nearly 56%. After immobilization, specific activity of the enzyme increases from 0. 0225 to 0.0452 with a concomitant slight alkaline shift in the optimum pH when assayed in both the directions. The immobilized enzyme also displays increased optimum temperature and thermo-stability and can be reused number of times. Use of immobilized enzyme for the production of glucose-1-phosphate 2.4.1.1 glycogen phosphorylase synthesis use of enzyme for alpha-glucosaminylation of maltooligosaccharides 2.4.1.2 dextrin dextranase synthesis enzyme is utilized for bioconversions of glycosides in diverse purposes, overview, and for production of dextran and oligodextran, no product inhibition, enzyme is robust in high level production 2.4.1.4 amylosucrase synthesis mutant enzyme R226A, that is activated by the products it forms and yields twice as much insoluble glucan and lower quantities of by-products as the wild-type enzyme is a very promising enzyme for industrial synthesis of amylose-like polymers 2.4.1.4 amylosucrase synthesis potentiality of amylosucrase in the design of amylodextrins with controlled morphology, structure, and physicochemical properties 2.4.1.4 amylosucrase synthesis potential of amylosucrase in the design of original carbohydrate-based dendritic nanoparticles 2.4.1.4 amylosucrase synthesis the enzyme can be used for synthesis of salicin glycosides with sucrose serving as the glucopyranosyl donor and salicin as the acceptor molecule 2.4.1.4 amylosucrase synthesis the enzyme can efficiently be used for synthesis of salicin glycosides with sucrose serving as the glucopyranosyl donor and salicin as the acceptor molecule 2.4.1.4 amylosucrase synthesis the enzyme might be useful for important tailoring reactions for the generation of bioactive compounds by glycosylation 2.4.1.4 amylosucrase synthesis amylosucrase has great potential in the biotechnology and food industries, due to its multifunctional enzyme activities. It can synthesize alpha-1,4-glucans, like amylose, from sucrose as a sole substrate. It can also utilize various other molecules as acceptors. In addition, amylosucrase produces sucrose isomers such as turanose and trehalulose. It also efficiently synthesizes modified starch with increased ratios of slow digestive starch and resistant starch, and glucosylated functional compounds with increased water solubility and stability. It produces turnaose more efficiently than other carbohydrate-active enzymes. Amylose synthesized by amylosucrase forms microparticles and these can be utilized as biocompatible materials with various bio-applications, including drug delivery, chromatography, and bioanalytical sciences 2.4.1.4 amylosucrase synthesis immobilization enhances the efficiency of the glycosylation reaction and is therefore considered effective for industrial application in sustainable production of dihydroxybenzene glucosides 2.4.1.4 amylosucrase synthesis isoquercitrin (quercetin-3-O-beta-D-glucopyranoside) has diverse biological functions, such as anti-oxidant and anticancer activity, but its use is limited by poor solubility and bioavailability. Enzymatically modified isoquercitrin (EMIQ) is a mixture of transglycosylated isoquercitrins that have better solubility and bioavailability than do quercetin and isoquercitrin. Amylosucrase (ASase), has transglycosylation activity to produce EMIQ. Both enzymes produce a variety of EMIQs including isoquercitrin, isoquercitrin-glucoside, isoquercitrin-diglucoside, and isoquercitrin-triglucoside. The enzyme has a higher bioconversion yield from isoquercitrin to EMIQ (97%). The yield of soquercitrin-triglucoside, which is the most bioavailable form is 46%. The enzyme can be used to synthesize EMIQ in a simple and specific process 2.4.1.4 amylosucrase synthesis mutant enzyme R226K/I228V/A289I/F290Y/E300I/V331T/Q437S/N439D/C445A only produces soluble oligosaccharides as no insoluble high molecular weight amylose is observed. The mutant enzyme is an attractive enzymatic tool that could offer interesting opportunities for the design of amylodextrins with controlled size 2.4.1.4 amylosucrase synthesis the batch-feeding whole-cell biocatalysis by Amy-1 is a promising technology for alpha-arbutin production with enhanced yield and molar conversion rate 2.4.1.4 amylosucrase synthesis the beta-carotene embedded amylose microparticles are prepared in one-step by utilizing the unique catalytic activity of amylosucrase from Deinococcus geothermalis, which synthesizes linear amylose chains using sucrose as the sole substrate. Synthesized amylose chains self-assembled with b-carotene to form well-defined spherical microparticles with an encapsulation yield of 65%. The synthetic method enables microparticle formation and beta-carotene encapsulation in one-step using amylosucrase and sucrose as the sole substrates, which indicates that the devised process may be cost-effective and suitable for industrial applications 2.4.1.4 amylosucrase synthesis transglycosylation reactions with amylosucrase from Deinococcus geothermalis constitute an efficient and economical method to produce alpha-glucosyl flavonoids 2.4.1.5 dextransucrase synthesis enzymatic synthesis of salicin prodrugs by the reaction of cyclomaltodextrin glucanyltransferase from Bacillus macerans with cyclomaltohexaose and salicyl alcohol which gives a salicin as a major product and the reaction of Leuconostoc mesenteroides B-742CB dextransucrase with sucrose and salicyl alcohol which gives a isosalicin as the major product 2.4.1.5 dextransucrase synthesis alpha-glycosylation by GTFR with sucrose and different alcohols and amino acid derivatives for the synthesis of glycoethers and glycosylated amino acids, which are not easy to obtain by chemical or enzymatic synthetic methods. These products can be used for solid-phase synthesis to generate glycopeptides 2.4.1.5 dextransucrase synthesis potential use of the enzyme in production of glucooligosaccharides containing alpha(1,2) bonds for the dermocosmetic industry 2.4.1.5 dextransucrase synthesis production of dextran 2.4.1.5 dextransucrase synthesis the use of cashew apple juice as substrate is an interesting alternative to grow Leconostoc mesenteroides and to produce dextransucrase. High enzyme activities are obtained even when the substrate is used without yeast extract or phosphate addition 2.4.1.5 dextransucrase synthesis engineered enzyme variants are capable to produce isomalto-oligosaccharides and dextrans of controlled molecular weight of about 10-40 kDa in a one-step process, overview 2.4.1.5 dextransucrase synthesis the enzyme is usable in the production of isomaltooligosaccharide, a promising dietary component with prebiotic effect, the long-chain IMOs are preferred to short chain ones owing to the longer persistence in the colon, optimization of synthesis of long-chain IMOs, overview 2.4.1.5 dextransucrase synthesis Leuconostoc mesenteroides dextransucrase is useful for enzymatic synthesis of alkyl alpha-D-glucosides, best yield is 50% using 1-butyl alpha-D-glucoside with 0.9 M 1-butanol 2.4.1.5 dextransucrase synthesis one-step synthesis of isomalto-oligosaccharides by a fusion enzyme of dextransucrase and dextranase 2.4.1.5 dextransucrase synthesis the enzyme is useful for production of L-ascorbic acid 2-glucoside for use as an antioxidant in industrial applications 2.4.1.5 dextransucrase synthesis the strain Leuconostoc citreum strain B/110-1-2 is used for industrial production of dextran and dextran derivatives 2.4.1.5 dextransucrase synthesis purified Weissella confusa Cab3 dextransucrase (WcCab3-DSR) is used for in vitro synthesis of dextran and glucooligosaccharides 2.4.1.5 dextransucrase synthesis combined use of dextransucrase and dextranase and the maltose acceptor is a simple and effective method to promote the high-quality of functional isomaltooligosaccharides 2.4.1.5 dextransucrase synthesis synthesis of glucosylated steviosides, which are more stable at pH 2, 60°C for 48 h than stevioside 2.4.1.5 dextransucrase synthesis the mutant enzyme is highly suitable for the synthesis of different flavonoid glucosides. For substrates such as quercetin and its glycosides, a high glucosylation efficiency is achieved, whereas the incubation of e.g. neohesperidin dihydrochalcone and naringin only yields low portions of glucoconjugates. The different substrates are not only glucosylated with varying efficiencies but also at different positions. While glucosylation of the phenolic hydroxyl groups of the flavonoid B-ring occurrs almost exclusively at position O4', flavonoid bound beta-glucosyl units are conjugated at position O3, O4, and O6 depending on the substrate. It is possible to demonstrate that flavonoids with a rutinose and a neohesperidose residue can be glucosylated either at position O6 of the glucose unit or at position O4 of the alpha-rhamnose unit 2.4.1.7 sucrose phosphorylase synthesis cheap and very efficient synthesis of N-acetyllactosamine, one-pot reaction combined with galactosyltransferase 2.4.1.7 sucrose phosphorylase synthesis enzymatic production of a chemically stable ascorbic acid derivative 2-O-alpha-D-glucopyranosyl-L-ascorbic acid 2.4.1.7 sucrose phosphorylase synthesis LmSPase is immobilised onto Eupergit C and used for the continuous production of alpha-D-glucose 1-phosphate from sucrose 2.4.1.7 sucrose phosphorylase synthesis the enzyme is useful as transglucosylation catalyst for synthesis of alpha-D-glucosides as industrial fine chemicals, overview. The enzyme is also used in the industrial process for production of 2-O-(alpha-D-glucopyranosyl)-sn-glycerol as active ingredient of cosmetic formulations 2.4.1.7 sucrose phosphorylase synthesis sucrose phosphorylase can glycosylate a variety of small molecules using sucrose as cheap ut efficient donor substrate. The immobilized enzyme is optimized due to a higher temperature tolerance compared to the soluble enzyme from Bifidobacterium adolescentis, overview 2.4.1.7 sucrose phosphorylase synthesis sucrose phosphorylase is a promising biocatalyst for the production of special sugars and glycoconjugates, but its transglycosylation activity rarely exceeds the competing hydrolytic reaction 2.4.1.7 sucrose phosphorylase synthesis 2-O-(alpha-D-glucopyranosyl)-sn-glycerol (alphaGG) is a natural osmolyte. alphaGG is produced industrially for application as an active cosmetic ingredient. Immobilized preparations of the enzyme on Sepabeads and TRISOPERL controlled pore glass (CPG) supports are useful catalysts for alphaGG production, for they show the same high selectivity in the transglucosylation from sucrose to glycerol as the soluble enzyme does. Up to 90 mg enzyme loaded on solid support to give highly active immobilizate. Product yields of 85% and product titers of 800 mM reach at high reaction selectivity. High productivity (500 mM/h) obtained with immobilized enzyme in a microstructured flow reactor 2.4.1.7 sucrose phosphorylase synthesis effective kojibiose synthesis i as performed using the crude enzyme solution from the supernatant of fermentation broth of Bacillus subtilis expressing the enzyme from Bifidobacterium adolescentis, providing a basis for potential industrial-scale preparation of kojibiose. Kojibiose is prebiotic element, a low-calorie sweetener and exhibits antitoxic activity, thereby promoting the emergence of new drugs 2.4.1.7 sucrose phosphorylase synthesis the enzyme is a promising biocatalyst for the glycosylation of a wide variety of carbohydrates and non-carbohydrate molecules. It is important for enzymatic synthesis of alpha-D-glycosides, which are widely used in food, medicine, cosmetics and other industries. It generally reversibly catalyses the conversion of sucrose and phosphate into fructose and alpha-D-glucose-1-phosphate, and it can also transfer glucosyl groups from sucrose to a hydroxy group in a suitable acceptor, generating new alpha-D-glucosidic products such as ascorbic acid 2-glucoside, alpha-arbutin, 2-O-(alpha-D-glucopyranosyl)-sn-glycerol. 2-O-alpha-D-glucopyranosyl-L-ascorbic acid is produced after incubation of ascorbic acid sodium salt and sucrose (1:2) with 19.76 U/ml of the enzyme at pH 7.0 and 50°C for 24 h, with a maximum yield of 19.7% (39.94 g/l) 2.4.1.8 maltose phosphorylase synthesis enzymatic synthesis of alpha-anomer-selective D-glucosides using maltose phosphorylase 2.4.1.8 maltose phosphorylase synthesis simple expression and purification protocol and the use of maltose as an inexpensive starting material make this maltose phosphorylase from Emticicia oligotrophica a valuable biocatalyst for the synthesis of glucose-containing glycosides 2.4.1.B9 D-Glc-alpha-1-diphosphoundecaprenol 4-beta-glucosyltransferase synthesis the enzyme is involved in biosynthesis of xanthan, an industrially important exopolysaccharide 2.4.1.10 levansucrase synthesis immobilization of recombinant free levansucrase on magnetite leads to production of low molecular weight levan, increased thermal stability of the enzyme 2.4.1.10 levansucrase synthesis enzyme is an important biocatalyst for production of fructose homopolymers, potential usage in bioindustrial fields 2.4.1.10 levansucrase synthesis production of the artificial sweetener, lactosucrose 2.4.1.10 levansucrase synthesis D-glucose acts as an inhibitor of the transfructosylation reaction, Candida cacaoi selectively removes glucose from the reaction medium, the decrease of glucose concentration by yeast in the medium by 16-19% results in the increased degree of levan polymerisation (by 6 to 9%) and efficiency of levan synthesis (by 9 to 11%) 2.4.1.10 levansucrase synthesis the enzyme is useful in levan production for application in the food and pharmaceutical industry 2.4.1.10 levansucrase synthesis levansucrase Lsc3 of Pseudomonas syringae pv. tomato has very high catalytic activity and stability making it a promising biotechnological catalyst for FOS and levan synthesis 2.4.1.10 levansucrase synthesis the enzyme is interesting for levan production due to its ability to directly use the free energy of cleavage of non-activated sucrose to transfer the fructosyl group to a variety of acceptors including monosaccharides (exchange), oligosaccharides (fructooligosaccharides synthesis) or a growing fructan chain (polymer synthesis). Levansucrase from Bacillus amyloliquefaciens is a promising biocatalyst for the synthesis of beta-(2-6)-linked-fructose-based carbohydrates (fructooligosaccharides, oligolevans, levans) targeting specific structures and functional properties 2.4.1.13 sucrose synthase synthesis a biocatalytic cascade of polyphosphate kinase and sucrose synthase is developed for synthesis of nucleotide-activated derivatives of glucose 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis - 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis application of immobilized enzyme is practically important for continous production of cyclodextrins, broad activity maximum is advantageous for industrial operations, immobilized enzyme is able to work at maximum efficiency at lower temperatures 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis ATCC 21783, enzyme is becoming commercially important since cyclodextrins have found various practical applications 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis thermostable, useful for industrial utilization 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis industrial strain E 192, industrial production of cyclodextrins 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis used in industrial applications 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis strain 1011, Y195L and Y195V CGTases acquired better characteristics for industrial use 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis constructing proteins having new properties of industrial importance 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis chemical industry 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis enzymatic synthesis of glycosides such as maltooligosyl sucrose, glycosyl stevioside and glycosyl ascorbic acid 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis enzymatic synthesis of salicin prodrugs by the reaction of cyclomaltodextrin glucanyltransferase from Bacillus macerans with cyclomaltohexaose and salicyl alcohol which gives a salicin as a major product and the reaction of Leuconostoc mesenteroides B-742CB dextransucrase with sucrose and salicyl alcohol which gives a isosalicin as the major product 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis production of alpha-cyclodextrin. Cyclodextrins serve as molecular hosts, are used in the food industry for capturing and retaining flavors and are also used in the formulation of pharmaceuticals. 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis potential industrial application of this CGTase in processes in which thermal stability is required. This enzyme could be used after starch gelatinization without cooling the solution to temperatures lower than 60°C 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis synthesis of cyclodextrins with multiple applications in the food, pharmaceutical, cosmetic, agricultural and chemical industries. Conditions used to produce cyclodextrins with cyclodextrin glycosyltransferase from Bacillus circulans DF 9R are optimized using experimental designs 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the CGTase-glyoxyl derivative allow the production of cyclodextrin at 85°C, giving twice the production rate compared to the free enzyme, 17.5% higher mass selectivity for beta-cyclodextrin and may help avoid microbial contamination. These characteristics are important considerations for the development of an industrial CD production continuous process 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme can be applied in biotechnology for the production of cyclodextrins and oligosaccharides with novel properties 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme is of interest for industrial cyclodextrin production 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme is useful for industrial production of cyclodextrins 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme is useful for production of alpha-, beta-, and gamma-cyclodextrins from soluble starch 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis CGTase is an important industrial enzyme, unique in its capacity to convert starch and related substrates into cyclodextrins through cyclization, an intramolecular transglycosylation reaction 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis CGTases is used to catalyze cyclomaltooligosaccharides/cyclodextrins production in important industrial processes 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis cyclodextrin glucanotransferases are industrially important enzymes that produce a mixture of cyclic alpha-(1,4)-linked oligosaccharides, cyclodextrins, from starch, overview. Use of complexing agents during cyclodextrin synthesis and the variation in solubility of the different cyclodextrins to allow selective precipitation. Usage of the enzyme as immobilized biocatalyst 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis efficient synthesis of a long carbohydrate chain alkyl glycoside catalyzed by CGTase, the equilibrium lays to 80% on the side of dodecyl-beta-D-maltooctaoside production when the enzyme from Bacillus macerans is used as biocatalyst, method evaluation, overview 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis manipulation of the enzyme by molecular imprinting to preferentially produce large-ring cyclodextrins, overview 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis synthesis of a long carbohydrate chain alkyl glycoside catalyzed by CGTase, the equilibrium lays to 80% on the side of dodecyl-beta-D-maltooctaoside production when the enzyme from Bacillus macerans is used as biocatalyst, method evaluation, overview 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme as membrane biocatalyst is applied for a direct cyclodextrin production 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis the enzyme is used for cyclodextrin production, generation of a modified enzyme with increased alpha-cyclization specificity, overview 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis enzyme achieves 47% conversion of an insoluble raw commercial corn starch into cyclodextrins with production of only beta- and gamma-cyclodextrins, in a ratio of 80%:20% in alkaline pH 9.0 2.4.1.19 cyclomaltodextrin glucanotransferase synthesis enzyme achieves 50.7% conversion of raw corn starch into 81.6% beta- and 18.4% gamma-cyclodextrins after 24 h enzyme reaction at 60°C and pH 8.0 2.4.1.20 cellobiose phosphorylase synthesis ability of the phosphorylase to utilize alpha-D -glucose 1-fluoride as alternate glucosyl donor in place of alpha-D-glucose 1-phosphate for the synthesis of alpha-1,4-glucosides under thermodynamic control in close to 100% yield 2.4.1.20 cellobiose phosphorylase synthesis the cellobiose phosphorylases in the cell extract is used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from non-radioactive glucose-1-phosphate and radioactive glucose. For cellobiose synthesis, the reaction is carried out at 60°C for 30 min with crude enzyme. After an impluse feed of radiolabeled cellobiose to a Clostridium thermocellum culture, the intracellular sugar levels are measured: the largest amount of radioactivity is cellobiose with lesser amounts of glucose, cellotriose, and cellotetraose, and an average degree of polymerization of intracellular cellodextrins is ca. 2 2.4.1.20 cellobiose phosphorylase synthesis the enzyme is useful for production of D-glucose 1-phosphate, an expensive substrate for other enzymatic syntheses 2.4.1.20 cellobiose phosphorylase synthesis enzymatic cellobiose synthesis from starch using two phosphorylases: glucan phosphorylase, from Klebsiella pneumoniae, converts glucose residues in the starch into glucose-1-phosphate, removal of phosphate, and glucose-1-phosphate is incubated with alpha-D-glucose and is converted into cellobiose by recombinant cellobiose phosphorylase, removal of phosphate, 60% cellobiose yield from glucose 1-phosphate and, at least, 23.7% cellobiose yield from starch, method optimization, overview 2.4.1.20 cellobiose phosphorylase synthesis the enzyme is useful for synthesis of alkyl beta-glucosides, overview 2.4.1.20 cellobiose phosphorylase synthesis expression of enzyme in Escherichia coli and study on the role of molecular chaperones and growth temperature on the solubilization of enzyme overexpressed in Escherichia coli. The growth of host at low temperature enhances enzyme in soluble fraction. Similarly, induction of target gene at low level of IPTG also yields more enzyme in the soluble fraction. Coexpression of the target gene with molecular chaperones GroESL and KODH does not enhance the solubilization under in vivo conditions 2.4.1.24 1,4-alpha-glucan 6-alpha-glucosyltransferase synthesis the enzyme is used in starch processing for the production of isomaltodextrins 2.4.1.25 4-alpha-glucanotransferase synthesis potential applications in the starch industry 2.4.1.25 4-alpha-glucanotransferase synthesis cycloamylose will be used in the food, pharmaceutical and chemical industries 2.4.1.25 4-alpha-glucanotransferase synthesis production of cycloamylose 2.4.1.25 4-alpha-glucanotransferase synthesis production of cycloamylose starch gels 2.4.1.25 4-alpha-glucanotransferase synthesis production of isomalto-oligosaccharides 2.4.1.25 4-alpha-glucanotransferase synthesis the enzyme is useful in the enzymatic synthesis of dimaltosyl-beta-cyclodextrin via a transglycosylation reaction 2.4.1.25 4-alpha-glucanotransferase synthesis the recombinant enzyme is used to enzymatically-synthesized glycogen as a food ingredient 2.4.1.25 4-alpha-glucanotransferase synthesis combination of maltogenic amylase reactions from Bacillus stearothermophilus and 4-alpha-glucanotransferase from Thermus scotoductus to increase the water solubility of puerarin, an isoflavonoid derived from Radix puerariae. The puerarin transfer products, including maltosyl-alpha-(1->6)-puerarin as a major product, are reacted with alpha-glucanotransferase in the presence of amylose. The maltosyl-alpha-(1->6)-puerarin-cycloamylose complex is formed by an elongation reaction and cyclization by alpha-glucanotransferase. The encapsulation of puerarin or glycosylated puerarin with a macrocyclic amylose is widely applicable both to improving the water solubility of the compound and stabilizing it during cold storage 2.4.1.25 4-alpha-glucanotransferase synthesis development of an efficient biocatalytic production process of cycloamyloses directly from sucrose by use of Synechocystis sp. 4-alpha-glucanotransferase and Neisseria polysaccharea amylosucrase. From one-pot synthesis, the maximum cycloamylose yield of 9.6%, w/w with 0.3 M sucrose is achieved with 10 units/ml of amylosucrase and 0.1 unit/ml of 4-alpha-glucanotransferase at 40°C for a 3 h reaction in a simultaneous dual enzyme reaction mode. The size of linear alpha-(1,4)-glucan is positively related to the cycloamylose productivity by 4-alpha-glucanotransferase in a hyperbolic manner. The dual enzyme reaction converts sucrose directly to cycloamyloses via in situ transient linear alpha-(1,4)-glucan as an soluble intermediate 2.4.1.30 1,3-beta-oligoglucan phosphorylase synthesis synthesis of laminarioligosaccharides, degree of polymerization is dependent on the glucose/glucose 1-phosphate ratio, overview 2.4.1.30 1,3-beta-oligoglucan phosphorylase synthesis synthesis of laminarioligosaccharides by combined action of laminaribiose phosphorylase EC 2.4.1.31 and beta-1,3-oligoglucan phosphorylase EC 2.4.1.30 2.4.1.38 beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase synthesis - 2.4.1.38 beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase synthesis use of transformed whole yeast cells, expressing the human N-acetylglucosamine beta-1,4-galactosyltransferase, in synthesis of N-acetyllactosamine 2.4.1.38 beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase synthesis preparation of a series of specific derivatives of the complex protopanaxadiol glycoside ginsenoside Rb1 2.4.1.38 beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase synthesis the enzyme is useful as catalyst for quantitative synthesis of Gal-beta-1,4-Man-pNP, overview 2.4.1.38 beta-N-acetylglucosaminylglycopeptide beta-1,4-galactosyltransferase synthesis the enzyme is useful as catalyst for quantitative synthesis of the thiodisaccharide Gal-betaS-1,4-GlcNAcpNP as well as Gal-beta-1,4-Man-pNP, overview 2.4.1.40 glycoprotein-fucosylgalactoside alpha-N-acetylgalactosaminyltransferase synthesis the enzyme possesses flexible substrate specificity and can therefore be used to synthesize five different types of A antigens with high efficiency, engineering of bacterial cell surface polysaccharides by BgtA, overview 2.4.1.B46 4-O-(alpha-L-rhodosaminyl)aklavinone L-2-deoxyfucosyltransferase synthesis AknK may be a useful enzyme for the chemoenzymatic synthesis of anthracycline variants 2.4.1.49 cellodextrin phosphorylase synthesis enzyme is a good tool for the synthesis of cellodextrins 2.4.1.49 cellodextrin phosphorylase synthesis synthesis of cellulase inhibitors from reaction of 4-O-beta-D-glucopyranosyl-1-deoxynojirimycin or 6-O-beta-cellobiosyl-1-deoxynojirimycin as acceptors with glucose 1-phosphate as donors 2.4.1.49 cellodextrin phosphorylase synthesis simple procedures for synthesizing various sugar length radioactive cellodextrins (G2–G6) are developed using the Clostridium thermocellum cellobiose and cellodextrin phosphorylases from small building blocks of nonradioactive glucose-1-phosphate and radioactive glucose 2.4.1.49 cellodextrin phosphorylase synthesis biocatalytic production of cellodextrins with a degree of polymerization. Kinetic and thermodynamic restrictions upon alpha-Glc1-P utilization (200 mM; 45°C, pH 7.0) are effectively overcome by in situ removal of the phosphate released via precipitation with Mg2+. The product's degree of polymerization is controlled by the molar ratio of glucose/alpha-Glc1-P used in the reaction. In optimized conversion, 36 g/l soluble cellodextrins are obtained. By keeping the glucose concentration low, the reaction is shifted completely towards insoluble product formation 2.4.1.49 cellodextrin phosphorylase synthesis to increase the reaction rate of cellodextrin phosphorolysis, synthetic enzyme complexes containing CDP and phosphoglucomutase are constructed to convert G1P to glucose 6-phosphate rapidly. The CDP-phosphoglucomutase enzyme complex with the highest enhancement of initial reaction rate is integrated into an in vitro synthetic enzymatic biosystem for generating bioelectricity from cellodextrin, exhibiting a much higher current density (3.35fold) and power density (2.14fold) than its counterpart biosystem containing free CDP and phosphoglucomutase mixture 2.4.1.B53 hesperetin 7-O-glucoside 6''-O-rhamnosyltransferase synthesis naringin and narirutin are produced from naringenin-7-O-glucoside using engineered fission yeast expressing the Arabidopsis thaliana rhamnose synthase RHM2 and the Citrus maxima alpha1,2-rhamnosyltransferase or the Citrus sinensis alpha16rhamnosyl transferase genes as a whole-cell-biocatalyst 2.4.1.54 undecaprenyl-phosphate mannosyltransferase synthesis useful for enzymatic synthesis of a wide variety of mannosylphosphopolyisoprenyl products 2.4.1.56 lipopolysaccharide N-acetylglucosaminyltransferase synthesis enzyme promise to be a useful catalyst in the preparation of both GlcNAcbeta1-3Gal and GalNAc1-3Gal linkages 2.4.1.64 alpha,alpha-trehalose phosphorylase synthesis synthesis of trehalose from maltose by a coupled enzyme system with trehalose phosphorylase and maltose phosphorylase 2.4.1.65 3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase synthesis the soluble form of fucosyltransferase III secreted by stably transfected cells may be used for in vitro synthesis of the Lewis 1 determinant on carbohydrates and glycoproteins 2.4.1.65 3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase synthesis by constructing yeast cells that display human FUT6 on the cell wall by fusion of FUT6 with the yeast cell wall proteins Pir1 and Pir2, the fucosylated oligosaccharides can be easily synthesized by the incubation of yeast cells with an appropriate donor and acceptor. It will thus be possible to prepare a large amount of immobilized FUT6 fused with Pir proteins in an inexpensive medium lacking the serum that is required for mammalian cell cultivation 2.4.1.65 3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase synthesis the enzyme possesses a broad tolerance toward nonnatural type I acceptor substrate analogs and therefore represents a valuable tool for the chemoenzymatic synthesis of Lewis A, sialyl Lewis A as well as mimetics thereof 2.4.1.65 3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase synthesis alpha1-3/4-fucosyltransferase is expressed in Escherichia coli at a level of 30 mg/l culture and used as a diverse catalyst in a one-pot multienzyme system for high-yield production of L-fucose-containing carbohydrates including Lewis antigens such as Lewis a, b, x, O-sulfated Lewis x, and sialyl Lewis x as well as human milk fucosides such as 3-fucosyllactose, lacto-N-fucopentaose III, lacto-N-difuco-hexaose II and III. While difucosylation of tetrasaccharides is readily achieved using an excess amount of donor, synthesis of lacto-N-fucopentaose III is achieved by alpha1-3/4-fucosyltransferase-catalyzed selective fucosylation of the N-acetyllactoamine component in lacto-N-neotetraose 2.4.1.B65 4-hydroxy-3-methoxybenzyl alcohol alpha-D-glucosyltransferase synthesis at optimum reaction temperature of 37°C, optimal maltose concentration of 60% (w/v), optimal pH 6.6, and optimal concentration of vanillyl alcohol of 158 mM, yield of vanillyl alcohol alpha-D-glucoside is 90 mM with no by product formation. This compound shows good antioxidant activity as well as stability in gastrointestinal tract. It is hydrolyzed on brush border membrane of enterocytes, so it can serve in protecting the gastrointestinal system from oxidation 2.4.1.68 glycoprotein 6-alpha-L-fucosyltransferase synthesis mutants of the general acid/base residue E274, including E274A, E274S, and E274G, act as efficient glycoligases able to fucosylate a wide variety of complex N-glycopeptides and intact glycoproteins by using alpha-fucosyl fluoride as a simple donor substrate. The alpha-1,6-fucosidase mutants can introduce an alpha1,6-fucose moiety specifically at the Asn-linked GlcNAc moiety to GlcNAc-peptide, and also to high-mannose and complex type N-glycans in the context of N-glycopeptides, N-glycoproteins, and intact antibodies 2.4.1.69 type 1 galactoside alpha-(1,2)-fucosyltransferase synthesis fast and efficient synthesis of a novel homologous series of L-fucosylated trisaccharides using the alpha-(1->2)-L-galactosyltransferase 2.4.1.69 type 1 galactoside alpha-(1,2)-fucosyltransferase synthesis large-scale synthesis of H-antigen oligosaccharides by expressing Helicobacter pylori alpha1,2-fucosyltransferase in metabolically engineered Escherichia coli cells 2.4.1.69 type 1 galactoside alpha-(1,2)-fucosyltransferase synthesis introduction of the WcfB gene plus the genes for guanosine 5'-diphosphate-L-fucose biosynthetic enzymes allows the lacZ-deleted Escherichia coli to produce 2-fucosyllactose. By fedbatch fermentation, 15.4 g/l (extracellular concentration) of 2-fucosyllactose can be obtained (yield of 0.858 g/g lactose and productivity of 0.530 g/l/h) 2.4.1.B77 alpha-(1->2) branching sucrase synthesis alpha(1->2) or alpha(1->3) branched dextrans with high molar masses and controlled architecture are synthesized using dextransucrase from Oenococcous kitahare DSM 17330 (DSR-OK), the branching sucrase from Leuconostoc citreum NRRL B-1299 (BRS-A) and the branching sucrase from Leuconostoc citreum NRRL B-742 (BRS-BDELTA1), all in cell-free extract from recombinant Escherichia coli strain BL21. Their molecular structure, solubility, conformation, film-forming ability, as well as their thermal and mechanical properties are determined, detailed overview 2.4.1.79 globotriaosylceramide 3-beta-N-acetylgalactosaminyltransferase synthesis synthesis of globoside and isogloboside tetrasaccharides by using beta(1->3) N-acetylgalactosaminyltransferase/UDP-N-acetylglucosamine C4 epimerase fusion protein 2.4.1.79 globotriaosylceramide 3-beta-N-acetylgalactosaminyltransferase synthesis synthesis of globotetraose (GalNAcb-3Gala-4Galb-4Glc)by the high cell density culture of an Escherichia coli strain over-expressing the Neisseria meningitidis lgtC gene for alpha-1,4-Gal transferase. The strain which is devoid of both alpha and beta galactosidase activity is fed with glycerol as the energy and carbon source and with lactose as precursor for globotriose synthesis. After complete exhaustion of lactose, globotriose could serve as an alternative acceptor for LgtC and the formation of a series of polygalactosylated compounds is observed. The system is extended to the synthesis of globotetraose (GalNAcbeta-3Gala-4Galbeta-4Glc) by overexpressing two additional genes: lgtD from Haemophilus influenzae Rd which encodes a beta-1,3-GalNAc transferase and wbpP from Pseudomonas aeruginosa which encodes a UDP-GalNAc C4 epimerase. Globotetraose could also be produced from exogenous globotriose which is actively taken up by the cells 2.4.1.81 flavone 7-O-beta-glucosyltransferase synthesis biotransformation of eriodictyol using Escherichi coli expressing AtGT-2 2.4.1.82 galactinol-sucrose galactosyltransferase synthesis use of an in vitro multienzyme system using sucrose synthase (SUS), UDP-glucose 4-epimerase (GalE), galactinol synthase (GS), raffinose synthase (RS), and stachyose synthase (STS) and intermedia UDP and inositol, which can be recycled, to synthesize raffinose,. The reaction system produces 11.1mM raffinose using purified enzymes under optimal reaction conditions and substrate concentrations. Optimization further increases raffinose production to 86.6 mM and enables the synthesis of 61.1 mM stachyose. The UDP turnover number reaches 337. Inositol in the reaction system is recycled five times, and 255.8 mM raffinose (128.9 g/liter) is obtained 2.4.1.85 cyanohydrin beta-glucosyltransferase synthesis integration of genes CYP79A1, CYP71E1, and UGT85B1 in Nicotiana tabacum chloroplast genome and functional expression, the enzymes convert endogenous tyrosine into dhurrin using electrons derived directly from the photosynthetic electron transport chain, without the need for the presence of an NADPH-dependent P450 oxidoreductase. The dhurrin produced in the engineered plants amounted to 0.1-0.2% of leaf dry weight compared to 6% in the origin Sorghum bicolor. Plant P450s involved in the synthesis of economically important compounds can be engineered into the thylakoid membrane of chloroplasts, and their full catalytic cycle can be driven directly by photosynthesis-derived electrons 2.4.1.87 N-acetyllactosaminide 3-alpha-galactosyltransferase synthesis - 2.4.1.87 N-acetyllactosaminide 3-alpha-galactosyltransferase synthesis enzymic synthesis of octadecameric saccharides of mutiply branched blood group I-type, carrying 4 distal alpha-1,3-galactose residues 2.4.1.87 N-acetyllactosaminide 3-alpha-galactosyltransferase synthesis useful tool for production of a range of compounds to further investigate the binding site of anti-Gal and other proteins with Galalpha(1-3)Gal binding sites 2.4.1.87 N-acetyllactosaminide 3-alpha-galactosyltransferase synthesis attaching beta p-nitrophenyl to galactose converts the complex from a poor into a good substrate, the group mimics a monosaccharide like N-acetylglucosamine 2.4.1.87 N-acetyllactosaminide 3-alpha-galactosyltransferase synthesis production of novel complex glycans with role in biological recognition processes with engineered enzyme with modified substrate specificity 2.4.1.90 N-acetyllactosamine synthase synthesis - 2.4.1.90 N-acetyllactosamine synthase synthesis use of transformed whole yeast cells, expressing the human N-acetylglucosamine beta-1,4-galactosyltransferase, in synthesis of N-acetyllactosamine 2.4.1.90 N-acetyllactosamine synthase synthesis preparation of a series of specific derivatives of the complex protopanaxadiol glycoside ginsenoside Rb1 2.4.1.90 N-acetyllactosamine synthase synthesis enzyme can be useful in the glycosylation of cytokines, enzymes or other glycosylated compounds modifying their functions for use in medical therapies 2.4.1.90 N-acetyllactosamine synthase synthesis development of a method to tag glycoproteins carrying terminal GlcNAc using the enzyme, overview 2.4.1.92 (N-acetylneuraminyl)-galactosylglucosylceramide N-acetylgalactosaminyltransferase synthesis expression of enzyme both as complete membrane-bound enzyme and as a soluble form in the baculovirus insect cell expression system 2.4.1.92 (N-acetylneuraminyl)-galactosylglucosylceramide N-acetylgalactosaminyltransferase synthesis stable expression of enzyme in CHO-Lec8 cells, production of complex-type N-glycans quantitatively bearing LacdiNAc-structures on their antennae and containing repeating LacdiNAc-structures 2.4.1.92 (N-acetylneuraminyl)-galactosylglucosylceramide N-acetylgalactosaminyltransferase synthesis the immobilized enzyme might be useful in chemoenzymatical ceramide glycoconjugate synthesis 2.4.1.99 sucrose:sucrose fructosyltransferase synthesis high transferase/hydrolase ratio confers it a great potential for the industrial production of prebiotic fructooligosaccharides 2.4.1.100 2,1-fructan:2,1-fructan 1-fructosyltransferase synthesis because inulin-type fructans with a high degree of polymerization show superior properties for specific food and non-food applications, the hDP 1-FFT gene from Viguiera discolor has potential for the production of inulin with a high degree of polymerization in vitro or in transgenic crops 2.4.1.100 2,1-fructan:2,1-fructan 1-fructosyltransferase synthesis the enzyme might be suitable for both fructo-oligosaccharides and high degree of polymerization inulin production in bioreactors 2.4.1.102 beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-N-acetylglucosaminyltransferase synthesis chemical-enzymatic synthesis of sialyl-Lex-containing hexasaccharides found on O-linked glycoproteins, process involves several enzymes of the pathway 2.4.1.115 anthocyanidin 3-O-glucosyltransferase synthesis expression of enzyme in Escherichia coli, together with dihydroflavonol 4-reductase, anthocyanidin synthase, flavanone 3beta-hydroxylase. production of pelargonidin 3-O-glucoside or cyanidin 3-O-glucoside from naringenin or eriodyctol, resp., via this recombinant plant pathway. Yields of 0.0056 mg/l pelargonidin 3-O-glucoside, or of 0.006 mg/l cyanidin 3-O-glucoside 2.4.1.140 alternansucrase synthesis modification of bacterial cellulose using sucrose, dextransucrase from Leuconostoc mesenteroides B-742CMB and alternansucrase from Leuconostoc B-1355C. This modification method produces a new bacterial cellulose that has a unique structure and probably new property 2.4.1.140 alternansucrase synthesis alternan polymer is more water soluble and less viscous than dextrans, making it a good substitute for Arabic gum. Nanoparticles and films have been obtained with alternan produced by Leuconostoc citreum ABK-1 alternansucrase, thereby opening up other potential applications in nanotechnology 2.4.1.140 alternansucrase synthesis alternansucrase enzyme from Leuconostoc citreum SK24.002 is used to produce di-glycosyl-stevioside through acceptor reaction, optimized method, overview 2.4.1.143 alpha-1,6-mannosyl-glycoprotein 2-beta-N-acetylglucosaminyltransferase synthesis the baculoviris-host system constitutively expressing beta-1,4-galactosyltransferase and beta-1,2-N-acetylglucosaminyltransferase II can be used to produce a recombinant glycoprotein with fully galactosylated, biantennary N-glycans 2.4.1.144 beta-1,4-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase synthesis overexpression of enzyme in human hepatoma cells fails to reduce branch formation of N-glycans, coexpression of caveolin-1 leads to a dramatic decrease in the extent of branching with no enhancement of enzyme activity 2.4.1.144 beta-1,4-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase synthesis RC12 cells, apheochromocytoma cell line, transfected with enzyme gene, result in suppression of neurite outgrowth induced by costimulation of epidermal growth factor and integrins. Epidermal growth factor receptor-mediated ERK-activation is blocked in transfectants. Epidermal growth factor receptor of transfectants is modified by bisecting GlcNAc in its N-glycan structures 2.4.1.144 beta-1,4-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase synthesis transfection of mouse aorta endothelial cells with enzyme results in reduction of their susceptibility to complement-mediated cell lysis by normal human serum and suppresses the antigenicity of cells to human natural antibodies. Reactivity of glycoproteins of transfectants to normal human serum and GSIB4 lectin is reduced 2.4.1.144 beta-1,4-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase synthesis introduction of human N-acetylglucosaminyltransferase gene into tobacco plants leads to highly efficient synthesis of bisected N-glycans. The majority of N-glycans of an antibody produced in a plant expressing GnT-III is also bisected. This might improve the efficacy of therapeutic antibodies produced in this type of transgenic plant 2.4.1.145 alpha-1,3-mannosyl-glycoprotein 4-beta-N-acetylglucosaminyltransferase synthesis it is necessary to select appropriate host cell lines according to their glycosyltransferases activity balance to produce glycoproteins with an objective sugar chain structure. The descibed method of grouping cell lines according to their glycosyltransferases activity balance is useful to select appropriate host cells 2.4.1.146 beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,3-N-acetylglucosaminyltransferase synthesis use of the recombinant enzyme for construction of the 6-sulfo sialyl Lewis x on extended core1 O-glycans 2.4.1.150 N-acetyllactosaminide beta-1,6-N-acetylglucosaminyltransferase synthesis cIGnT6 transfers multiple GlcNAc branches to long linear polylactosamines, a prerequisite for improving enzyme-assisted in vitro synthesis of a type of multivalent sialyl Lewis x glycans that are high affinity inhibitors of lymphocyte L-selectin, enzyme allows general polylactosamine synthesis 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis recombinant yeast cells which provide an inexpensive, self-perpetuating source of fucosyltransferase activity immobilized in the cell wall, useful for in vitro synthesis of sLex 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis stable expression of beta1,4-N-acetylgalactosaminyltransferase from Caenorhabditis elegans and human FucT9 in CHO cells, producing fucosylated poly-LacdiNAc N-glycans 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis a chemoenzymatic model for the preparative-scale synthesis of a diverse array of GDP-fucose derivatives is reported 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis a KO line lacking two XylT genes and four FucT genes expressing a human IgG2 antibody shows an IgG2 expression level as high as in a control transformant. The IgG glycosylation profile shows no beta(1,2)-xylose or alpha(1,3)-fucose present on the glycosylation moiety. The dominant glycoform is the GnGn structure 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis expression of bacterial alpha-1,3-fucosyltransferase in engineered Escherichia coli deficient in beta-galactosidase activity and expressing the essential enzymes for the production of guanosine 5'-diphosphate-L-fucose, for synthesis of 3-fucosyllactose. The fucT gene gives the best 3-fucosyllactose production in a simple batch fermentation process using glycerol as a carbon source and lactose as an acceptor. With concomitant blocking the colanic acid biosynthetic pathway, the recombinant strain exhibits the highest concentration (11.5 g/l), yield (0.39 mol/mol), and productivity (0.22 g/l/h) of 3-fucosyllactose in glycerol-limited fed-batch fermentation 2.4.1.152 4-galactosyl-N-acetylglucosaminide 3-alpha-L-fucosyltransferase synthesis to overcome the poor solubility of FutA upon expression in Escherichia coli, codon optimization, and systematic truncation of the protein at the C-terminus with only one heptad repeat remaining yield 150-200 mg/l of soluble protein of FutA and result in more than an 18fold increase in the 3-fucosyllactose yield. Mutant A128N obtained by focused directed evolution mutant displays a 3.4fold higher catalytic activity than wild-type FutA. Mutant A128N/H129E/Y132I exhibits a 9.6fold improvement in specific activity when compared to wild-type. Mutants A128N/H129E/S46F and A128N/H129E/Y132I/S46F show synergistic effects, that is 14.5- and 15.5fold improvement in specific activity relative to wild-type. The mutations increase the binding affinity for lactose and kcat values for lactose and GDP-fucose. The quadruple mutant A128N/H129E/Y132I/S46F synthesizes 3-fucosyllactose with an improved yield and productivity (more than 96% yield based on 5 mM of GDP-Fuc within 1 h) 2.4.1.173 sterol 3beta-glucosyltransferase synthesis - 2.4.1.173 sterol 3beta-glucosyltransferase synthesis synthesis of steryl beta-D-monoglucosides that are constituents of the cell membrane of higher plants 2.4.1.173 sterol 3beta-glucosyltransferase synthesis synthesis of steryl beta-D-monoglucosides possible function as glucose carriers through the membrane and intercellular transport of sterols 2.4.1.173 sterol 3beta-glucosyltransferase synthesis synthesis of poriferasterol monoglucoside may be involved in differentiation 2.4.1.177 cinnamate beta-D-glucosyltransferase synthesis GT2 enzymes might contribute to the production of ellagic acid/ellagitannins in strawberry and raspberry, and are useful to develop strawberry fruit with additional health benefits and for the biotechnological production of bioactive polyphenols 2.4.1.177 cinnamate beta-D-glucosyltransferase synthesis trans-cinnamic acid (tCA) and hydrocinnamyl alcohol (HcinOH) are valuable aromatic compounds with applications in the flavour, fragrance and cosmetic industry. Production in transgenic Saccharomyces cerevisiae strain strain MGY0.06 expressing phenylalanine ammonia lyase (PAL, from Photorhabdus luminescens) and an aryl carboxylic acid reductase, as well as UDP-glucose:cinnamate glucosyltransferase FaGT2 from from Fragaria ananassa. Overview of the heterologous pathway 2.4.1.182 lipid-A-disaccharide synthase synthesis synthesis of a variety of artificial fluorinated lipid A precursor analogues on a preparative scale for investigation of structure-function relationships of lipid A derivatives 2.4.1.186 glycogenin glucosyltransferase synthesis the COOH-terminal fragment of glycogenin can be used as a effective high affinity reagent for the purification of glycogen synthase from skeletal muscle and liver 2.4.1.211 1,3-beta-galactosyl-N-acetylhexosamine phosphorylase synthesis production of dissaccharidic structures instead of chemical synthesis 2.4.1.211 1,3-beta-galactosyl-N-acetylhexosamine phosphorylase synthesis suitable catalyst for practical syntheses of oligosaccharides 2.4.1.211 1,3-beta-galactosyl-N-acetylhexosamine phosphorylase synthesis large scale production of lacto-N-biose I, a building block of type I human milk oligosaccharides 2.4.1.212 hyaluronan synthase synthesis optimization of the recombinant enzyme expression in Escherichia coli for large scale production of hyaluronan polymers for usage in basic studies, and for biotechnological creation of functional carbohydrates in medical purposes, engineering of produced product chain length 2.4.1.212 hyaluronan synthase synthesis it may be possible to generate compounds that will selectively inhibit the binding of hyaluronan to one particular hyaladherin species without perturbing other species. Such sugar molecules may have future utility as selective therapeutics with minimal side effects for diseases such as cancer, autoimmune disease, inflammation, and infection 2.4.1.212 hyaluronan synthase synthesis multi-enzyme strategy for the in vitro one-pot synthesis of high-molecular-weight hyaluronic acid from substrates sucrose and N-acetylglucosamine (GlcNAc) with in situ regeneration of nucleotide sugars. With optimized reaction conditions, hyaluronic acid with a molecular mass above 2 MDa is synthesized from catalytic UDP concentrations and 10 mM GlcNAc in less than 10 h 2.4.1.212 hyaluronan synthase synthesis the hasA gene is cloned and expressed in Escherichia coli BL21 and the use this recombinant Escherichia coli strain BL21 is used as host in order to explore the biosynthetic capabilities for hyaluronan production (concentration and molecular weight) in batch fermentation using shake flask culture. High hyaluronan production (0.115 g/l) is achieved in nutrient rich media with 50 g/l glucose concentration. With addition of 1.0 mM isopropyl beta-D-1-thiogalactopyranoside, the highest hyaluronan production (0.532 g/l) and molecular weight at 34600 Da is achieved which is around fivefold higher compared to without isopropyl beta-D-1-thiogalactopyranoside 2.4.1.214 glycoprotein 3-alpha-L-fucosyltransferase synthesis a KO line lacking two XylT genes and four FucT genes expressing a human IgG2 antibody shows an IgG2 expression level as high as in a control transformant. The IgG glycosylation profile shows no beta(1,2)-xylose or alpha(1,3)-fucose present on the glycosylation moiety. The dominant glycoform is the GnGn structure 2.4.1.216 trehalose 6-phosphate phosphorylase synthesis efficient one-pot enzymatic synthesis of trehalose 6-phosphate. The one-pot reaction of trehalose 6-phosphate phosphorylase and maltose phosphorylase enables production of 65 mM trehalose 6-phosphate from 100 mM maltose, 100 mM Glc6P, and 20 mM inorganic phosphate. Addition of beta-phosphoglucomutase to this reaction produces glucose 6-phosphate from beta-glucose 1-phosphate and thus reduces requirement of glucose 6-phosphate as a starting material. Within the range of 20-469 mM inorganic phosphate tested, the 54 mM concentration yielded the highest amount of trehalose 6-phosphate (33 mM). Addition of yeast increases the yield because of its glucose consumption. From 100 mmol maltose and 60 mmol inorganic phosphate, the production of 37.5 mmol trehalose 6-phosphate in a one-pot reaction (100 ml) is achieved, and 9.4 g trehalose 6-phosphate dipotassium salt is obtained 2.4.1.218 hydroquinone glucosyltransferase synthesis formation of arbutin, which is a potent inhibitor of human melanin biosynthesis with commercial value 2.4.1.218 hydroquinone glucosyltransferase synthesis an artificial pathway is established in Escherichia coli for high-level production of arbutin from simple carbon sources in Escherichia coli for high-level production of arbutin from simple carbon sources. Introduction of the genes for 4-hydroxybenzoate 1-hydroxylase from Candida parapsilosis CBS604 and hydroquinone glucosyltransferase from Rauvolfia serpentina into Escherichia coli leads to the production of 54.71 mg/l of arbutin from glucose. Further redirection of carbon flux into arbutin biosynthesis pathway by enhancing shikimate pathway genes enables production of 3.29 g/l arbutin. Final optimization of glucose concentration added in the culture medium is able to further improve the titer of arbutin to 4.19 g/l in shake flasks experiments 2.4.1.222 O-fucosylpeptide 3-beta-N-acetylglucosaminyltransferase synthesis use of manic fringe in engineering of a mammalian O-glycosylation pathway in the yeast Saccharomyces cerevisiae for production of O-fucosylated epidermal growth factor domains. In the system, manic fringe facilitates the addition of N-acetylglucosamine to the EGF domain from factor IX but not from factor VII 2.4.1.227 undecaprenyldiphospho-muramoylpentapeptide beta-N-acetylglucosaminyltransferase synthesis overexpression of enzyme results in formation of vesicular intracellular membranes enriched in cardiolipin. Cardiolipin content of cell is about 7fold increased 2.4.1.230 kojibiose phosphorylase synthesis synthesis of selaginose by kojibiose phosphorylase gene and trehalose phosphorylase gene intracellularly hyperexpressed under the control of the Bacillus amyloliquefaciens alpha-amylase promoter in Bacillus subtilis 2.4.1.230 kojibiose phosphorylase synthesis the phosphorylase can efficiently catalyze the reverse reaction with high specificity, and thus can be applied to the practical synthesis of alpha-glucosyl oligosaccharides 2.4.1.231 alpha,alpha-trehalose phosphorylase (configuration-retaining) synthesis sucrose is converted to alpha,alpha-trehalose in about 60% yield using a coupled enzyme system composed of sucrose phosphorylase from Leuconostoc mesenteroides, glucose isomerase from Streptomyces murinus and the appropriately stabilized PoTPase 2.4.1.236 flavanone 7-O-glucoside 2''-O-beta-L-rhamnosyltransferase synthesis metabolically engineered plant cell suspension cultures expressing recombinant Cm1,2RhaT can biotransform hesperetin, the hesperidin aglycone, into neohesperidin 2.4.1.245 alpha,alpha-trehalose synthase synthesis the enzyme may be useful for the regeneration of NDP-alpha-D-glucose from NDP, especially for ADP-alpha-D-glucose from ADP, with trehalose as a glucose resource 2.4.1.245 alpha,alpha-trehalose synthase synthesis trehalose synthase from Pyrococcus horikoshii can be applied to the sugar nucleotide cycling process for the synthesis of functional alpha-galactosyl oligosaccharides, alpha-galactose epitopes and globotriose, using the effective regeneration of UDP-galactose 2.4.1.245 alpha,alpha-trehalose synthase synthesis maltose, NaCl, K2HPO4 and beef extract are the most significant factors affecting trehalose synthase production in Corynebacterium glutamicum. Trehalose synthase activity increases to 580 U/ml, an approximate 1.76-fold increase over the previous activity (330 U/ml) using an optimized fermentation medium with the main composition maltose (22g/l), beef extract (7.49g/l), NaCl (13.36g/l), K2HPO4(11.33g/l) and peptone (15.00g/l) 2.4.1.245 alpha,alpha-trehalose synthase synthesis synthesis of the trehalase analogue alpha-D-Glc-(1->1)-alpha-D-Glc, that might be effective at inhibiting intestinal brush-order disaccharidases 2.4.1.245 alpha,alpha-trehalose synthase synthesis the enzyme is a useful thermostable enzyme particularly for the production of the trehalose analogue containing a mannose that acts as the inhibitor of intestinal sucrase 2.4.1.247 beta-D-galactosyl-(1->4)-L-rhamnose phosphorylase synthesis suitable catalyst for practical syntheses of oligosaccharides 2.4.1.248 cycloisomaltooligosaccharide glucanotransferase synthesis immobilization of enzyme for production of cycloisomaltooligosaccharides. Immobilization on Chitopearl BCW-3505 results in 35% remaining activity, on Chitopearl BCW-3005 35%, Chitopearl SH-3505 28%, and Diaion HP-20 on 22% remaining activity. The maximum cycloisomaltooligosaccharides yield in batch reactions at 0.2, 2 and 10% dextran was 28, 24 and 12%, respectively. The concentration of linear oligosaccharides, the byproducts in the reaction mixture, isgreater with the immobilized CITase than the free enzyme. The immobilized CITase is less thermostable than the free enzyme by about 10°C 2.4.1.248 cycloisomaltooligosaccharide glucanotransferase synthesis immobilization of enzyme on anion-exchange porous hollow-fiber membrane with a degree of enzyme multilayer-binding of 0.3-9.8 and production of seven- to nine-glucose-membered cycloisomaltooligosaccharides from dextran at a maximum yield of 28% in weight at a space velocity of 10 per h during the permeation of 2.0% w/w dextran solution across the enzyme-immobilized porous hollow-fiber membrane. Yield increases with increasing degree of enzyme multilayering 2.4.1.248 cycloisomaltooligosaccharide glucanotransferase synthesis optimal conditions for production of cyclo-(-6)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-4)-alpha-D-Glc-(1-) from starch are substrate concentration 1%, pH 5.5, temperature 55°C, 24 h reaction time, enzyme concentration 1 unit/g-dry solid, isoamylase 2500 units/g-dry-solid 2.4.1.248 cycloisomaltooligosaccharide glucanotransferase synthesis use of enzyme for synthesis of cycloisomaltooligosaccharides from dextran. Enzyme immobilized on chitopearl BCW-3505 is not influenced by NaCl up to 2 M. When 2 M NaCl is included in the substrate solution during continuous production of cycloisomaltooligosaccharides by a column system packed with the immobilized enzyme, no microbial contamination appears and cycloisomaltooligosaccharides are produced for 40 days. The added NaCl has no influence on the life time of the system and is effective in suppressing the growth of contaminating microbes. The added NaCl can be separated easily from cycloisomaltooligosaccharides by an open column system packed with activated carbon 2.4.1.248 cycloisomaltooligosaccharide glucanotransferase synthesis the enzyme can be applied to produce relatively large CIs at high temperature 2.4.1.251 GlcA-beta-(1->2)-D-Man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphospho-ditrans,octacis-undecaprenol 4-beta-mannosyltransferase synthesis the enzyme is involved in biosynthesis of xanthan, an industrially important exopolysaccharide 2.4.1.252 GDP-mannose:cellobiosyl-diphosphopolyprenol alpha-mannosyltransferase synthesis the enzyme is involved in biosynthesis of xanthan, an industrially important exopolysaccharide 2.4.1.257 GDP-Man:Man2GlcNAc2-PP-dolichol alpha-1,6-mannosyltransferase synthesis engineering of a synthetic pathway in Escherichia coli for the production of eukaryotic trimannosyl chitobiose glycans and the transfer of these glycans to specific asparagine residues in target proteins. Glycan biosynthesis is enabled by four eukaryotic glycosyltransferases, including the yeast uridine diphosphate-N-acetylglucosamine transferases Alg13 and Alg14 and the mannosyltransferases Alg1 and Alg2. By including the bacterial oligosaccharyltransferase PglB from Campylobacter jejuni, glycans are successfully transferred to eukaryotic proteins 2.4.1.258 dolichyl-P-Man:Man5GlcNAc2-PP-dolichol alpha-1,3-mannosyltransferase synthesis genetic engineering of N-glycan biosynthesis in Yarrowia lipolytica so that it produces Man(3)GlcNAc(2) structures on its glycoproteins. Disruption of the ALG3 gene, EC 2.4.1.258, results in modification of proteins mainly with Man(5)GlcNAc(2) and GlcMan(5)GlcNAc(2) glycans, and to a lesser extent with Glc(2)Man(5)GlcNAc(2) glycans. To avoid underoccupancy of glycosylation sites, Alg6, EC 2.4.1.267, is concomitantly overexpressed. Overexpression of the heterodimeric Aspergillus niger glucosidase II results in removal the terminal glucose residues. Overexpression of an alpha-1,2-mannosidase leads to Man(3)GlcNAc(2) structures, which are substrates for the synthesis of complex-type glycans. The final Yarrowia lipolytica strain produces proteins glycosylated with the trimannosyl core N-glycan (Man(3)GlcNAc(2)), which is the common core of all complex-type N-glycans 2.4.1.264 D-Man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphosphoundecaprenol 2-beta-glucuronosyltransferase synthesis the enzyme is involved in biosynthesis of the industrially important exopolysaccharide xanthan 2.4.1.264 D-Man-alpha-(1->3)-D-Glc-beta-(1->4)-D-Glc-alpha-1-diphosphoundecaprenol 2-beta-glucuronosyltransferase synthesis the enzyme is involved in biosynthesis of xanthan, an industrially important exopolysaccharide 2.4.1.265 dolichyl-P-Glc:Glc1Man9GlcNAc2-PP-dolichol alpha-1,3-glucosyltransferase synthesis expression of the genes encoding for alginate-lyase algL and the catalytic subunit of the alginate polymerase complex, dolichyl-P-Glc:Glc1Man9GlcNAc2-PP-dolichol alpha-1,3-glucosyltransferase alg8, in chemostat cultures of Azotobacter vinelandii ATCC 9046. Increased alginate production of 2.4 g per l and a higher specific alginate production rate of 0.1 g per g and h are obtained at an inlet sucrose concentration of 15 g per l. Carbon recovery of about 100% is obtained at an inlet sucrose concentration of 10 g per l, whereas it is close to 50% at higher inlet sucrose concentrations. An increase in the molecular weight of alginate is linked to higher alg8 gene expression 2.4.1.267 dolichyl-P-Glc:Man9GlcNAc2-PP-dolichol alpha-1,3-glucosyltransferase synthesis genetic engineering of N-glycan biosynthesis in Yarrowia lipolytica so that it produces Man(3)GlcNAc(2) structures on its glycoproteins. Disruption of the ALG3 gene, EC 2.4.1.258, results in modification of proteins mainly with Man(5)GlcNAc(2) and GlcMan(5)GlcNAc(2) glycans, and to a lesser extent with Glc(2)Man(5)GlcNAc(2) glycans. To avoid underoccupancy of glycosylation sites, Alg6, EC 2.4.1.267, is concomitantly overexpressed. Overexpression of the heterodimeric Aspergillus niger glucosidase II results in removal the terminal glucose residues. Overexpression of an alpha-1,2-mannosidase leads to Man(3)GlcNAc(2) structures, which are substrates for the synthesis of complex-type glycans. The final Yarrowia lipolytica strain produces proteins glycosylated with the trimannosyl core N-glycan (Man(3)GlcNAc(2)), which is the common core of all complex-type N-glycans 2.4.1.276 zeaxanthin glucosyltransferase synthesis biosynthesis of zeaxanthin diglucoside is obtained when the gene crtX is co-transformed into Escherichia coli containing the plasmids carrying crtE, crtB, crtI, crtY, and crtZ genes required for zeaxanthin beta-D-diglucoside biosynthesis 2.4.1.276 zeaxanthin glucosyltransferase synthesis production of rare beta-carotene-modified carotenoids possessing 2-O, 2-H or 2-glucosyl and/or 3-O, 3-H or 3-glucosyl functionalities in their beta-ionone rings using a recombinant Escherichia coli approach, involving expression of carotenoid biosynthesis genes crtE, crtB, crtI, crtY, crtZ, crtX and crtG. From the cells of the recombinant Escherichia coli, caloxanthin i.e. (beta,beta-carotene-2,3,2',3'-tetrol)-3'-beta-D-glucose, zeaxanthin i.e. (beta,beta-carotene-3,3'-diol) 3,3'-beta-D-diglucoside, and nostoxanthin i.e. (beta,beta-carotene-2,3,3'-triol) are isolated and identified. Caloxanthin 3'-beta-D-glucoside displays potent 1O2 quenching activity 2.4.1.276 zeaxanthin glucosyltransferase synthesis Escherichia coli cells that express the full six carotenoid biosynthesis genes (crtE, crtB, crtI, crtY, crtZ, and crtX) of the bacterium Pantoea ananatis synthesizes zeaxanthin 3,3'-beta-D-diglucoside and 3-beta-glucosyl-3'-beta-quinovosyl zeaxanthin. The singlet oxygen-quenching activity of zeaxanthin 3,3'-beta-D-diglucoside is superior to that of zeaxanthin 2.4.1.276 zeaxanthin glucosyltransferase synthesis Escherichia coli strain CAR001 carries the crtEXYIB operon of Enterobacter agglomerans to produce beta-carotene. Deletion of genes encoding zeaxanthin glucosyltransferase (crtX) and lycopene beta-cyclase (crtY) from the operon leads to production of 10.5 mg lycopene/l. Further optimization results in a strain producing 3.52 g lycopene/l in fed-batch fermentation 2.4.1.277 10-deoxymethynolide desosaminyltransferase synthesis expression of different deoxysugar biosynthetic gene cassettes and the gene encoding a substrate-flexible glycosyltransferase DesVII in Streptomyces venezuelae YJ003 mutant strain bearing a deletion of a desosamine biosynthetic (des) gene cluster. The resulting recombinants produce macrolide antibiotic YC-17 analogs possessing unnatural sugars replacing native D-desosamine. These metabolites are D-quinovosyl-10-deoxymethynolide, L-rhamnosyl-10-deoxymethynolide, L-olivosyl-10-deoxymethynolide, and D-boivinosyl-10-deoxymethynolide 2.4.1.308 GDP-Fuc:beta-D-Gal-1,3-alpha-D-GalNAc-1,3-alpha-GalNAc-diphosphoundecaprenol alpha-1,2-fucosyltransferase synthesis stepwise enzymatic synthesis of human blood group B antigen tetrasaccharide. Biosynthetic pathway of B antigen is the same in Escherichia coli O86 as in humans, thus providing us an attractive way of synthesizing blood group B antigen with microbial glycosyltransferases 2.4.1.309 UDP-Gal:alpha-L-Fuc-1,2-beta-Gal-1,3-alpha-GalNAc-1,3-alpha-GalNAc-diphosphoundecaprenol alpha-1,3-galactosyltransferase synthesis stepwise enzymatic synthesis of human blood group B antigen tetrasaccharide. Biosynthetic pathway of B antigen is the same in Escherichia coli O86 as in humans, thus providing an attractive way of synthesizing blood group B antigen with microbial glycosyltransferases 2.4.1.310 vancomycin aglycone glucosyltransferase synthesis in vitro sequential glycosylations of the vancomycin aglycon catalyzed by the glycosyltransferases GtfE and GtfD and total synthesis of vancomycin and derivatives. For GtfE, presence of reductant tris(2-carboxyethyl)phosphine (TCEP, 2 mM) is crucial 2.4.1.318 demethyllactenocin mycarosyltransferase synthesis the disruption of the tylCV gene results in the accumulation of desmycosin, the direct precursor of tilmicosin. After optimization of the culture conditions, the yield of desmycosin reaches 7.3 g/l in a 5 l jar fermentor 2.4.1.321 cellobionic acid phosphorylase synthesis an engineered strain of Neurospora crassa (F5) with six of seven beta-glucosidase (bgl) genes knocked out produces cellobiose and cellobionate directly from cellulose without the addition of exogenous cellulases. Further modification by knock-out of catabolite repression genes, cre-1 and ace-1, leads to improved cellobiose dehydrogenase and exoglucanase expression but not to an improvement in cellobiose or cellobionate production. Deletion of the cellobionate phosphorylase gene NdvB from the genome of F5 lacking ace-1 and cre-1 to prevent the consumption of cellobiose and cellobionate results in a slightly reduced hydrolysis rate and in convertsion of 75% of the cellulose consumed to the desired products, cellobiose and cellobionate, compared to 18% converted by the strain F5 lackong only ace-1 cre-1 2.4.1.321 cellobionic acid phosphorylase synthesis engineering of recombinant Neurospora crassa strains to produce cellobionate. Recombinant strains heterologously express laccase genes from different sources in Neurospora crassa which has six out of seven beta-glucosidase, two transcription factors, and the cellobionate phosphorylase (ndvB) genes deleted. The engineered strain produces 47.4 mM cellobionate from cellulose without any enzyme addition 2.4.1.322 devancosaminyl-vancomycin vancosaminetransferase synthesis in vitro sequential glycosylations of the vancomycin aglycon catalyzed by the glycosyltransferases GtfE and GtfD and total synthesis of vancomycin and derivatives 2.4.1.330 beta-D-glucosyl crocetin beta-1,6-glucosyltransferase synthesis synthesis of crocetin and crocin-5 can be obtained in zeaxanthin-producing Escherichia coli by expression of dioxygenase CCD2 from Crocus sativus, glycosyltransferases UGT94E5 and UGT75L6 from Gardenia jasminoides, aldehyde dehydrogenase ALD8 from Crocus sativus and glycosyltransferases YjiC, YdhE and YojK from Bacillus subtilis 2.4.1.331 8-demethyltetracenomycin C L-rhamnosyltransferase synthesis expression of NDP-glucose-synthase OleS, NDP-glucose-4, 6-dehydratase OleE, 2, 3-dehydratase OleV, 3, 5-epimerase OleL and 3-ketoreductase OleW in combination with elloramycin glycosyltransferase ElmGt in Streptomyces lividans results in accumulation of 8-demthyltetracenomycin C plus 8-demethyl-8-(4-dehydro)-alpha-L-olivosyl-tetracenomycin and 8-demethyl-8-alpha-L-olivosyl-tetracenomycin 2.4.1.331 8-demethyltetracenomycin C L-rhamnosyltransferase synthesis generation of L- and D-stereoisomers of amicetose by combining sugar biosynthesis genes from four different antibiotic gene clusters and transfer of both sugars to the elloramycin aglycone by the sugar flexible ElmGT glycosyltransferase to yield antitumor tetracenomycins 2.4.1.333 1,2-beta-oligoglucan phosphorylase synthesis synthesis of beta-1,2-glucan with sophorose as an acceptor through three synthesis steps using sucrose phosphorylase and 1,2-beta-oligoglucan phosphorylase. The first step is performed in 200 microl reaction solution containing 5 mM sophorose and 1.0 M sucrose. beta-1,2-Glucan in a part of the resultant solution is hydrolyzed to beta-1,2-glucooligosaccharides by beta-1,2-glucanase. The second synthesis is performed in 25 times the volume for the first synthesis. The hydrolysate solution (1% volume of the reaction solution) is used as an acceptor. After treatment with the beta-1,2-glucanase again, the third synthesis is performed in 200 times the volume of the second synthesis (1 l). More than 140 g of beta-1,2-glucan is synthesized using approximately 20 microg of sophorose as the starting acceptor material 2.4.1.337 1,2-diacylglycerol 3-alpha-glucosyltransferase synthesis optimization of protein expression through controlling a few basic expression parameters, including temperature and growth media. The final expression level can be increased by two orders of magnitude, reaching 170 mg of pure protein per litre culture 2.4.1.344 type 2 galactoside alpha-(1,2)-fucosyltransferase synthesis enzyme is applied in an efficient one-pot multienzyme fucosylation system for high-yield synthesis of human blood group H antigens containing beta1-3-linked galactosides and an important human milk oligosaccharide lacto-N-fucopentaose I in preparative and gram scales 2.4.1.359 glucosylglycerol phosphorylase (configuration-retaining) synthesis glucosylglycerol phosphorylase might be an attractive biocatalyst for the production of the osmolyte glucosylglycerol, which is currently produced on industrial scale by exploiting a side activity of the closely related sucrose phosphorylase 2.4.1.360 2-hydroxyflavanone C-glucosyltransferase synthesis biosynthesis of natural and novel C-glycosylflavones utilising recombinant Oryza sativa C-glycosyltransferase and Desmodium incanum root proteins 2.4.1.360 2-hydroxyflavanone C-glucosyltransferase synthesis production of C-glucosides of flavonoids and related compounds (i.e., 2-hydroxyflavanone, dihydrochalcone, and trihydroxyacetophenone) by Escherichia coli expressing C-glucosyltransferase from Fagopyrum esculentum. The substrates in their respective cultures were taken up by the cells and C-glucosylated, and the products were released into the culture media. The bioconversion process was completed in 1-2 h, but products were already observed immediately after addition of the substrates (0.2 mM). The conversion rates of these substrates reaches 80-95% 2.4.1.362 alpha-(1->3) branching sucrase synthesis using a suitable sucrose/dextran ratio, a comb-like dextran with 50% ofx02alpha-(1->3) branching can be synthesized 2.4.1.362 alpha-(1->3) branching sucrase synthesis alpha(1->2) or alpha(1->3) branched dextrans with high molar masses and controlled architecture are synthesized using dextransucrase from Oenococcous kitahare DSM 17330 (DSR-OK), the branching sucrase from Leuconostoc citreum NRRL B-1299 (BRS-A) and the branching sucrase from Leuconostoc citreum NRRL B-742 (BRS-BDELTA1), all in cell-free extract from recombinant Escherichia coli strain BL21. Their molecular structure, solubility, conformation, film-forming ability, as well as their thermal and mechanical properties are determined, detailed overview 2.4.1.372 mutansucrase synthesis one-pot synthesis and development of unnatural-type bio-based polysaccharide, alpha-1,3-glucan. Synthesis can be achieved by in vitro enzymatic polymerization with GtfJ utilizing sucrose as a glucose monomer source, via one-pot water-based reaction. The structure of alpha-1,3-glucan is completely linear without branches with weight-average molecular weight of 700 kDa. Acetate and propionate esters of alpha-1,3-glucan are additionally synthesized and characterized 2.4.1.373 alpha-(1->2) branching sucrase synthesis use of enzyme to generate a platform for flavonoid glucosylation. Engineered variants show remarkable ability for luteolin, morin and naringenin glucosylation with conversion ranging from 30% to 90% 2.4.1.389 solabiose phosphorylase synthesis production of solabiose from lactose and sucrose. Lactose is hydrolyzed to D-galactose and D-glucose by beta-galactosidase. Phosphorolysis of sucrose and synthesis of solabiose are then coupled by adding sucrose, sucrose phosphorylase, and enzyme PBOR_28850 to the reaction mixture. Using 210 mmol lactose and 280 mmol sucrose, 207 mmol of solabiose is produced. Yeast treatment degrades the remaining monosaccharides and sucrose without reducing solabiose. Solabiose with a purity of 93.7% is obtained without any chromatographic procedures 2.4.2.1 purine-nucleoside phosphorylase synthesis efficient overexpression of thermostable enzyme in Escherichia coli. The increase of the cultivation temperature from 30°C to 42°C allows a more than 10fold increase of activity per cell and optimizing the 5 mRNA gene sequence further increases the activity per cell 1.7fold at 42°C 2.4.2.1 purine-nucleoside phosphorylase synthesis synthesis of ribavirin by the enzyme and comparison with Escherichia coli enzyme. A cold-adapted Pseudoalteromonas enzyme shows approximately 15 degrees lower optimum temperature and 1.80fold higher catalytic efficiency at 37°C within substrate inosine than its homolog from Escherichia coli 2.4.2.1 purine-nucleoside phosphorylase synthesis two-step efficient synthesis of 5-methyluridine, a intermediate in the synthesis of anti-AIDS drug such as stavudine and zidovudine 2.4.2.1 purine-nucleoside phosphorylase synthesis use of agar from four different polymer carriers as a suitable matrix for whole recombinant cell entrapment. Although the immobilization process reduces the substrate affinity and catalytic efficiency of recombinant cells, it can significantly enhance the stability and reusability of these cells. The immobilized whole cell biocatalyst is applied to produce ribavirin, as a model nucleoside synthesis reaction showing relative high productivity of rabavirin and quick reaction time 2.4.2.1 purine-nucleoside phosphorylase synthesis the enzyme is a valuable industrial biocatalysts for high-temperature reactions that produce nucleoside drugs in high yields 2.4.2.1 purine-nucleoside phosphorylase synthesis enzymatic ribosylation of fluorescent 8-azapurine derivatives, like 8-azaguanine and 2,6-diamino-8-azapurine, with purine-nucleoside phosphorylase (PNP) as a catalyst, leads to N9, N8, and N7-ribosides. The final proportion of the products may be modulated by point mutations in the enzyme active site. As an example, ribosylation of the latter substrate by wild-type calf PNP gives N7- and N8-ribosides, while the N243D mutant directs the ribosyl substitution at N9- and N7-positions. The same mutant allows synthesis of the fluorescent N7-D-ribosyl-8-azaguanine. The mutated form of the Escherichia coli PNP, D204N, can be utilized to obtain non-typical ribosides of 8-azaadenine and 2,6-diamino-8-azapurine as well. The N7- and N8-ribosides of the 8-azapurines can be analytically useful, as illustrated by N7-D-ribosyl-2,6-diamino-8-azapurine, which is a good fluorogenic substrate for mammalian forms of PNP, including human blood PNP, while the N8-riboside is selective to the Escherichia coli enzyme 2.4.2.1 purine-nucleoside phosphorylase synthesis enzymatic ribosylation of fluorescent 8-azapurine derivatives, like 8-azaguanine and 2,6-diamino-8-azapurine, with purine-nucleoside phosphorylase (PNP) as a catalyst, leads to N9, N8, and N7-ribosides. The final proportion of the products may be modulated by point mutations in the enzyme active site. As an example, ribosylation of the latter substrate by wild-type calf PNP gives N7- and N8-ribosides, while the N243D mutant directs the ribosyl substitution at N9- and N7-positions. The same mutant allows synthesis of the fluorescent N7-D-ribosyl-8-azaguanine. The N7- and N8-ribosides of the 8-azapurines can be analytically useful, as illustrated by N7-D-ribosyl-2,6-diamino-8-azapurine, which is a good fluorogenic substrate for mammalian forms of PNP, while the N8-riboside is selective to the Escherichia coli enzyme 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme AmPNP is an excellent biocatalyst for the industrial production of purine nucleoside analogues, biosynthesis of purine nucleoside analogues, overview 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 2,6-diaminopurine 2'-deoxyriboside 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 2,6-diaminopurine riboside and 5-methyluridine 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 2,6-diaminopurine riboside, 2,6-dichloropurine riboside, 6-chloro-2-fluoropurine riboside, 2-chloroadenine riboside, 2-fluoroadenine riboside, 2,6-diaminopurine 2'-deoxyriboside, 2,6-dichloropurine 2'-deoxyriboside, 6-chloro-2-fluoropurine 2'-deoxyriboside, 2-chloro-2'-deoxyadenosine (cladribine), and 2-fluoro-2'-deoxyadenosine, as well as of 2,6-diaminopurine arabinoside, 2,6-dichloropurine arabinoside, and 6-chloro-2-fluoropurine arabinoside 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 2-chloro-2'-deoxyadenosine (Cladribine), and 2-fluoroadenine arabinoside 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 2-fluoro-2'-deoxyadenosine and 2'-deoxy-2'-fluoroadenosine 2.4.2.1 purine-nucleoside phosphorylase synthesis enzyme is useful in biocatalytic production of 5-methyluridine 2.4.2.1 purine-nucleoside phosphorylase synthesis important application of PNP is in chemo-enzymatic synthesis of bioactive nucleoside analogues, utilizing various types of PNP among others. This application may be extended to tricyclic nucleobase analogues, particularly to adenine, isoguanine, and guanine derivatives. In particular, N9-D-riboside of can be obtained quantitatively from 1,N2-ethenoguanine, and N9-beta-D- and N7-beta-D-ribosides of 1,N6-etheno-isoguanine as a mixture, using the Escherichia coli PNP as a biocatalyst 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis production of ribavirin, which is an antiviral drug 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis overexpression of enzyme in Escherichia coli, medium optimization and presence of uridine and 5-fluorouracil results in conversion of more than 90% of uridine to 5-fluorouridine by the cells 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis the enzyme is useful in biocatalytic production of 2,6-dichloropurine 2'-deoxyriboside and 6-chloro-2-fluoropurine 2'-deoxyriboside 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis the enzyme is useful in biocatalytic production of 2-chloro-2'-deoxyadenosine (cladribine), adenine-arabinoside, 2-fluoroadenine arabinoside, and 5-methyluridine 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis the enzyme is useful in biocatalytic production of 2-chloroadenine riboside, 2-fluoroadenine riboside, 2,6-diaminopurine 2'-deoxyriboside, 2-chloro-2'-deoxyadenosine, 2-fluoro-2'-deoxyadenosine, 2'-deoxy-2'-fluoroadenosine, 2'-deoxy-2'-fluoro-arabinofuranosyl adenine, 2,6-diaminopurine arabinoside, 2,6-dichloropurine arabinoside, 6-chloro-2-fluoropurine arabinoside, and 5-iododeoxyuridine 2.4.2.2 pyrimidine-nucleoside phosphorylase synthesis during enzymatic synthesis of adenine arabinoside from adenine and uracil arabinoside with wet cells of Klebsiella aerogenes induced by cytidine or CMP, the reaction time can be shortened from 36 to 6 h 2.4.2.3 uridine phosphorylase synthesis evolvement of a mutant enzyme by iterative saturation mutagenesis. Compared to the wild type enzyme, which has a temperature optimum of 40°C and a half-life of 9.89 h at 60°C, the selected mutant has a temperature optimum of 60°C and a half-life of 17.3 h at 60°C. Self-immobilization of the native enzyme as a Spherezyme shows a 3.3fold increase in thermostability while immobilized mutant enzyme shows a 4.4fold increase in thermostability. Combining the enzyme with the purine nucleoside phosphorylase from Bacillus halodurans allows for synthesis of 5-methyluridine (a pharmaceutical intermediate) from guanosine and thymine in a one-pot transglycosylation reaction. Replacing the wild type uridine phosphorylase with the mutant allows for an increase in reaction temperature to 65°C and increased the reaction productivity from 10 to 31 g per l and h 2.4.2.3 uridine phosphorylase synthesis two-step efficient synthesis of 5-methyluridine, a intermediate in the synthesis of anti-AIDS drug such as stavudine and zidovudine 2.4.2.3 uridine phosphorylase synthesis the enzyme is a valuable industrial biocatalysts for high-temperature reactions that produce nucleoside drugs in high yields 2.4.2.4 thymidine phosphorylase synthesis construction of Escherichia coli expression vector pDEOA and use of lactose instead of IPTG to induce expression. The use of lactose at concentrations above 0.5 mmol/l has an induction effect similar to that of IPTG but results in a longer initial induction time and better cell growth. The thymidine phosphorylase induced by lactose is very stable at 50°C. Intact pDEOA cells induced by lactose can be used as a source of thymidine phosphorylase. Under standard reaction conditions, several deoxynucleosides are effectively produced from thymidine 2.4.2.4 thymidine phosphorylase synthesis immobilization of enzyme on solid support with the aim to have a stable and recyclable biocatalyst for nucleoside synthesis. Immobilization by ionic adsorption on amine-functionalized agarose and Sepabeads results in more than 85% activity recovery. Cross-linking with aldehyde dextran, MW20 kDa, decreases the percentage of expressed activity after immobilization by about 25%, but results in up to 6fold and 3fold higher stability than the soluble enzyme and the non-cross-linked counterpart, respectively, at pH 10 and 37°C. The preparation can be successfully used for the one-pot synthesis of 5-fluoro-2'-deoxyuridine starting from 2'-deoxyuridine or thymidine and 5-fluorouracil. In both cases, the reaction proceeds at the same rate leading to 62% conversion in 1 h 2.4.2.5 nucleoside ribosyltransferase synthesis the enzyme is a thermophilic and halotolerant biocatalyst that is successfully employed in the synthesis of different purine ribonucleoside analogues, potential of AvNRT as an industrial biocatalyst for the synthesis of nucleoside analogues 2.4.2.6 nucleoside deoxyribosyltransferase synthesis development of a practical method for enzymatic synthesis of deoxyguanosine by the combination of transglycosylation with NdRT-II from thymidine to a 2-amino-6-substituted purine, and the hydrolysis reaction with bacterial adenosine deaminase 2.4.2.6 nucleoside deoxyribosyltransferase synthesis useful as biocatalyst, immobilization on calcium alginate or calcium pectate of the enzyme increases the 2'-deoxynucleoside synthesis effiency of the organism 2.4.2.6 nucleoside deoxyribosyltransferase synthesis covalent attachment of recombinant Lactobacillus reuteri 2'-deoxyribosyltransferase to Sepabeads EC-EP303. The immobilized enzyme retains 50% of its maximal activity after 17.9 h at 60°C, and 96% activity is observed after storage at 40°C for 110 h. Immobilized enzyme can be recycled for 26 consecutive batch reactions in the synthesis of 2,6-diaminopurine-2'-deoxyriboside with negligible loss of catalytic activity and can be employed in the enzymatic synthesis of different natural and therapeutic nucleosides such as 5-ethyl-2'-deoxyuridine and 5-trifluorothymidine 2.4.2.6 nucleoside deoxyribosyltransferase synthesis immobilization of 2'-deoxyribosyltransferase from Lactobacillus reuteri on Sepabeads. Immobilized enzyme activity is enhanced 1.2–1.4fold at 20% of methanol, ethanol, 2-propanol, diethylene glycol, and acetone, and at 10% and 30% acetonitrile. Highest increased activity is also obtained in presence of 20% acetonitrile. Immobilized enzyme is successfully used in the synthesis of 2'-deoxyxanthosine and 2'-deoxyguanosine using 2'-deoxyuridine as sugar donor and the corresponding poor water-soluble base in the presence of 30% of methanol, ethanol, 2-propanol, ethylene glycol, acetonitrile, and DMSO, giving high nucleoside yields at 4 h 2.4.2.6 nucleoside deoxyribosyltransferase synthesis compared with NPTs (EC 2.4.2.5), NDTs present the advantage of catalyzing transglycosylation reactions between purine or pyrimidine bases and nucleosides in one enzyme one-pot mode 2.4.2.6 nucleoside deoxyribosyltransferase synthesis potential of LmPDTas an industrial biocatalyst for enzymatic production of several natural and non-natural therapeutic nucleosides, such as vidarabine (ara A), didanosine (ddI), ddG, or 2'-fluoro-2'-deoxyguanosine 2.4.2.6 nucleoside deoxyribosyltransferase synthesis TbPDT is proficient in the biosynthesis of numerous therapeutic nucleosides, including didanosine, vidarabine, cladribine, fludarabine, and nelarabine. TbPDT has good potential as an industrial biocatalyst for the synthesis of a wide range of therapeutic nucleosides through an efficient and environmentally friendly methodology 2.4.2.6 nucleoside deoxyribosyltransferase synthesis the enzyme can be used for in-flow synthesis of nucleoside analogues (2'-deoxy, 2',3'-dideoxy and arabinonucleoside derivatives) of pharmaceutical interest in mono- and a bi-enzymatic analytical immobilized enzyme reactors (IMERs), method, overview 2.4.2.6 nucleoside deoxyribosyltransferase synthesis the enzyme is successfully employed in the enzymatic production of several therapeutic nucleosides such as didanosine, vidarabine, and cytarabine 2.4.2.6 nucleoside deoxyribosyltransferase synthesis the enzyme is used for a green bioprocess by employing an environmentally friendly methodology to produce floxuridine (5-fluoro-2'-deoxyuridine), a compound with proven anti-tumor activity. The enzyme as enzymatic biocatalyst meets the requirements of high activity, stability, and short reaction times needed for low-cost production in a future preparative application 2.4.2.7 adenine phosphoribosyltransferase synthesis purine nucleotide synthesis using TthAPRT and TthHPRT (EC 2.4.2.8) 2.4.2.8 hypoxanthine phosphoribosyltransferase synthesis a lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta) deletion strain is evolved for 2,000 h, resulting in a stable strain with 40:1 ethanol selectivity and a 4.2-fold increase in ethanol yield over the wild-type strain. In a coculture of organic acid-deficient engineered strains of both Clostridium thermocellum and Thermoanaerobacterium saccharolyticum, fermentation of 92 g/liter Avicel results in 38 g/liter ethanol, with acetic and lactic acids below detection limits, in 146 h. engineering is based on a phosphoribosyl transferase (Hpt) deletion strain, which produces acetate, lactate, and ethanol in a ratio of 1.7:1.5:1.0, similar to the 2.1:1.9:1.0 ratio produced by the wild type. The Hpt/Ldh double mutant strain does not produce significant levels of lactate and has a 1.4:1.0 ratio of acetate to ethanol. Similarly, the Hpt/Pta double mutant strain does not produce acetate and has a 1.9:1.0 ratio of lactate to ethanol. The Hpt/Ldh/Pta triple mutant strain achieves ethanol selectivity of 40:1 relative to organic acids 2.4.2.8 hypoxanthine phosphoribosyltransferase synthesis potential of ZgHGPRT/AMPK as biocatalyst for the synthesis of nucleoside-50-mono-, di-, and triphosphates 2.4.2.9 uracil phosphoribosyltransferase synthesis MtTtUPRT is successfully applied in the enzymatic synthesis of therapeutic NMP analogues. The immobilized enzyme is usable as a biocatalyst in the synthesis of different 5-modified uridine-5'-monophosphates at short times. MTtUPRT3 can be reused up to 8 times in the synthesis of uridine-5'-monophosphate (UMP) with maximum activity at 100°C and pH 7.0 (activity 968 IU/g support, retaining activity 100%). Potential application of MTtUPRT3 as industrial biocatalyst 2.4.2.38 glycoprotein 2-beta-D-xylosyltransferase synthesis potential of maize and the beta1,2-xylosyltransferase as a production system for heterologous glycoproteins 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis recombinant enzyme may be used for in vitro synthesis of oligosaccharides 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis enzyme can be exploited as biocatalyst in the synthesis of interesting non-natural compounds, especially in view of chemical, regiospecific sialylation, chemo-enzymatic synthesis of modified carbohydrate ligands, and their suitability as probes for studying molecular recognition phenomena 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis because this enzyme is most active at basic pH, sialyltransferase obtained from Photobacterium leiognathi JT-SHIZ-145 is a promising tool for the efficient production of sialosides and the modification of glycoconjugates 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis the Photobacterium sialyltransferase can be used in the synthesis of sialoside analogues having a large substituent at the 9-position of Neu5Ac 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis application of the alpha2,6-trans-sialidase activity of DELTA15Pd2,6ST in the synthesis of sialosides 2.4.3.1 beta-galactoside alpha-(2,6)-sialyltransferase synthesis the enzyme may be a powerful tool for the synthesis of sialosides and the modification of sialyl-glycoconjugates 2.4.3.3 alpha-N-acetylgalactosaminide alpha-2,6-sialyltransferase synthesis use of glycan microarrays for high-throughput acceptor specificity screening of recombinant sialyltransferases. Donor substrate is cytidine 5'-monophospho-N-acteylneuraminate, which has been biotinylated at position 9. human alpha2,6sialyltransferase-I also sialylates chitobiose structures including N-glycans 2.4.3.4 beta-galactoside alpha-2,3-sialyltransferase synthesis chemoenzymatic synthesis of sialylated oligosaccharides for their evaluation in a polysialyltransferase assay 2.4.3.4 beta-galactoside alpha-2,3-sialyltransferase synthesis the enzyme can be exploited for the enzymatic synthesis of diverse sialyl products 2.4.3.6 N-acetyllactosaminide alpha-2,3-sialyltransferase synthesis chemical-enzymatic synthesis of sialyl-Lewis x-containing hexasaccharides found on O-linked glycoproteins, process involves several enzymes of the pathway 2.4.3.6 N-acetyllactosaminide alpha-2,3-sialyltransferase synthesis enzymatic synthesis of 4-methylumbelliferyl derivatives of N-acetyllactosaminide and sialyl N-acetyllactosaminides from 4-methylumbelliferyl N-acetyl-beta-D-glucosaminide by beta-galactosidase and sialyltransferase. The fluorescent synthesized compounds are useful for the assay of neuraminidase, sialyltransferase, and fucosyltransferase 2.4.3.6 N-acetyllactosaminide alpha-2,3-sialyltransferase synthesis the enzyme is useful for chemoenzymatic synthesis of artificial glycopolypeptides containing multivalent sialyloligosaccharides with a gamma-polyglutamic acid backbone. These sialyloligosaccharides are effective in inhibition of infections of cells by influenza viruses, which bind preferentially to alpha2,3-linked Neu5Ac 2.5.1.6 methionine adenosyltransferase synthesis improved production of erythromycin A by expression of a heterologous gene encoding S-adenosylmethionine synthetase in Saccharopolyspora erythraea 2.5.1.6 methionine adenosyltransferase synthesis recombination of MAT genes from Escherichia coli, Saccharomyces cerevisiae, and Streptomyces spectabilis by DNA shuffling and transformation into Pichia pastoris. In the two best recombinant strains, the MAT activities are respectively 201% and 65% higher than the recombinant strains containing the starting MAT genes, and the SAM concentration increases by 103% and 65%, respectively. A 6.14 g/l of SAM production is reached in a 500 l bioreactor with the best recombinant strain 2.5.1.6 methionine adenosyltransferase synthesis recombination of MAT genes from Escherichia coli, Saccharomyces cerevisiae, and Streptomyces spectabilis by DNA shuffling and transformation into Pichia pastoris. In the two best recombinant strains, the MAT activities are respectively 201% and 65% higher than the recombinant strains containing the starting MAT genes, and the SAM concentration increases by 103% and 65%, respectively. The K18R mutation of Streptomyces spectabilis probably result in the increased activity of the best MAT. A 6.14 g/l of SAM production is reached in a 500 l bioreactor with the best recombinant strain 2.5.1.19 3-phosphoshikimate 1-carboxyvinyltransferase synthesis enzyme is a target for development and synthesis of antimicrobial drugs 2.5.1.20 rubber cis-polyprenylcistransferase synthesis metal ion concentration regulates rubber initiation rate, biosythetic rate and molecular weight of product 2.5.1.20 rubber cis-polyprenylcistransferase synthesis natural rubber is one of the most important raw materials, and is industrially utilized for a huge variety of rubber products such as car tires. Natural rubber is synthesized by the rubber transferase on rubber particles in latex. The purified rubber particles show increasing levels of the rubber transferase activities with decreasing particle size 2.5.1.21 squalene synthase synthesis the engineered enzym emutant mMaSQS?C17 (E186K) is a potential candidate of the terpenes and steroids synthesis employed for synthetic biology 2.5.1.27 adenylate dimethylallyltransferase synthesis method for enzymatic preparation of isopentenyladenine-type and trans-zeatin-type cytokinins with radioisotope-labeling. The method is based on EC 2.5.1.27 from Arabidopsis thaliana, EC 3.1.3.1 from calf intestine and EC 2.4.2.1 from Escherichia coli 2.5.1.29 geranylgeranyl diphosphate synthase synthesis the enzyme in Xanthophyllomyces is used for production of astaxanthin, i.e. 3,3'-hydroxy-4,4'-diketo-beta-carotene, which is an industrially important carotenoid used for feeding salmon or trout in farming 2.5.1.29 geranylgeranyl diphosphate synthase synthesis to engineer a host that has the capability to supply geranylgeranyl diphosphate, a common precursor of isoprenoids, isopentenyl diphosphate isomerase (encoded by idi) from Escherichia coli and geranylgeranyl diphosphate synthase (encoded by gps) from Archaeoglobus fulgidus are cloned and overexpressed. The latter is shown to be a multifunctional enzyme converting dimethylallyl diphosphate to geranylgeranyl diphosphate. These two genes and the gene cluster (crtBIYZW) of the marine bacterium Agrobacterium aurantiacum are introduced into Escherichia coli to produce astaxanthin, an orange pigment and antioxidant. The metabolically engineered strain produces astaxanthin at a very high rate 2.5.1.29 geranylgeranyl diphosphate synthase synthesis enzyme IdsA is a target for metabolic engineering of carotenoid production in Corynebacterium glutamicum 2.5.1.B31 5-dimethylallyltryptophan synthase synthesis prenyltransferases of the DMATS superfamily are successfully used for production of prenylated compounds including analogues of their natural substrates such as simple indoles, tryptophan-containing cyclic dipeptides, and tyrosine derivatives, overview 2.5.1.B31 5-dimethylallyltryptophan synthase synthesis possibility of producing prenylated analogues of ardeemin fumiquinazoline, a precursor of the multidrug resistance (MDR) export pump inhibitor ardeemin, by using dimethylallydiphosphate transferase enzymes, e.g. 5-DMATS, overview 2.5.1.B31 5-dimethylallyltryptophan synthase synthesis possibility of producing prenylated analogues of ardeemin fumiquinazoline, a precursor of the multidrug resistance (MDR) export pump inhibitor ardeemin, by using dimethylallydiphosphate transferase enzymes, e.g. 5-DMATSSc, overview 2.5.1.B31 5-dimethylallyltryptophan synthase synthesis potential usage of tryptophan prenylating enzymes as biocatalysts for Friedel-Crafts alkylation 2.5.1.34 4-dimethylallyltryptophan synthase synthesis prenyltransferases of the DMATS superfamily are successfully used for production of prenylated compounds including analogues of their natural substrates such as simple indoles, tryptophan-containing cyclic dipeptides, and tyrosine derivatives, overview 2.5.1.34 4-dimethylallyltryptophan synthase synthesis possibility of producing prenylated analogues of ardeemin fumiquinazoline, a precursor of the multidrug resistance (MDR) export pump inhibitor ardeemin, by using dimethylallydiphosphate transferase enzymes, e.g. 4-DMATS, overview 2.5.1.47 cysteine synthase synthesis synthesis of L-Cys, therefore immobilization 2.5.1.47 cysteine synthase synthesis overexpression of enzyme in cytosol, chloroplast, or both of Nicotiana tabacum, transgenic plants show significantly more tolerance than wild-type against Cd2+, Se2+, and Ni2+. Application of transgenic plants to phyto-remediation of Cd2+ from contaminated soils 2.5.1.47 cysteine synthase synthesis enzme OASS produces novel beta-substituted L-amino acids when offered unnatural nucleophiles instead of sulfid, which is useful in producing pharmaceuticals, such as mucolytic agent L-carbocisteine and building blocks for the synthesis of pharmaceuticals and agrochemicals 2.5.1.47 cysteine synthase synthesis CysK overexpressing cells grow fast during log phase, and produce 26.5% more L-isoleucine in flask fermentation and 23.5% more L-isoleucine in fed-batch fermentation. The key genes aspC, lysC, hom, thrB, ilvA, and ilvBN involved in L-isoleucine biosynthesis are all upregulated in CysK overexpressing Corynebacterium glutamicum IWJ001. Overexpressing cells are longer and thicker than control cells, and their membrane permeability increases by 15.8% and biofilm formation ability decreases by 71.3% 2.5.1.48 cystathionine gamma-synthase synthesis production of alpha or beta deuterated amino acids 2.5.1.48 cystathionine gamma-synthase synthesis production of L-cystathionine 2.5.1.48 cystathionine gamma-synthase synthesis construction of an Escherichia coli strain for production of O-succinyl-L-homoserine. The metJ transcriptional repressor and metI subunit of DL-methionine transporter are deleted, metL (encodes bifunctional aspartate kinase/homoserine dehydrogenase II) is overexpressed and metB (encodes cystathionine gamma-synthase) is inactivated. The O-succinyl-L-homoserine titer thus increases to 7.30 g/l. Release of the feedback regulation leads to production of 9.31 g/l O-succinyl-L-homoserine from 20 g/l glucose in batch fermentation 2.5.1.49 O-acetylhomoserine aminocarboxypropyltransferase synthesis overexpression of gene metY increases the in vitro activity of O-acetylhomoserine aminocarboxypropyltransferase to 1780 mU/mg and is beneficial for methionine production, since the intracellular methionine pool increases 2fold in the engineered strain 2.5.1.54 3-deoxy-7-phosphoheptulonate synthase synthesis synthesis of L-phenylalanine (an important amino acid that is widely used in the production of food flavors and pharmaceuticals) by engineered Escherichia coli. Coexpression of Vitreoscilla hemoglobin gene, driven by a tac promoter, with the genes encoding 3-deoxy-D-arabinoheptulosonate-7-phosphate synthetase (aroF) and feedback-resistant chorismate mutase/prephenate dehydratase (pheAfbr), leads to increased productivity of L-phenylalanine and decreased demand for aeration by Escherichia coli CICC10245 2.5.1.54 3-deoxy-7-phosphoheptulonate synthase synthesis a strain carrying a mutant protein lacking 81 N-terminal amino acids shows an increased 2-phenylethanol production 2.5.1.56 N-acetylneuraminate synthase synthesis development of a novel microbial transformation for the synthesis of N-acetyl-D-neuraminic acid using bacterial cells expressing N-acetyl-D-glucosamine 2-epimerase and N-acetylneuraminate synthase 2.5.1.56 N-acetylneuraminate synthase synthesis construction of a recombinant Escherichia coli simultaneously overexpressing GlcNAc 2-epimerase and N-acetylneuraminic acid synthase. The recombinant strain produces 25 g/l N-acetylneuraminic acid in 22 h without the addition of Corynebacterium ammoniagenes cells. For manufacturing on an industrial scale, it is preferable to use unconcentrated culture broth as the source of enzymes. The N-acetylneuraminic acid aldolase gene of the host strain is disrupted yielding a strain which cannot degrade N-acetylneuraminic acid. After a 22 h reaction with 120 g/l N-acetylglucosamine in a 5 l jar fermenter, the culture broth of contains 53 g/l N-acetylneuraminic acid 2.5.1.56 N-acetylneuraminate synthase synthesis the enzyme might be useful for production of sialic acids and derivates at low temperatures 2.5.1.56 N-acetylneuraminate synthase synthesis design of an in vivo NeuAc biosensor and application for genetic screening of a mutant library of NeuAc producers. A Ni2+-based selection system couples the cell growth with in vivo NeuAc concentration. The final strain produces up to 8.31 g/l NeuAc in minimal medium using glucose as sole carbon source 2.5.1.56 N-acetylneuraminate synthase synthesis NeuAc production in Bacillus subtilis is enhanced to 46.04 g/l, by expressing NanA from Staphylococcus hominis. Enhanced expression of NanAand blocking formation of the by-product acetoin from pyruvate, result in 68.75 g/l NeuAc production with a molar conversion rate of 55.57% from GlcNAc 2.5.1.57 N-acylneuraminate-9-phosphate synthase synthesis synthesis of CMP-N-acetylneuraminic acid in Sf9 expression system by coexpression of enzyme with UDP-GlcNAc-2-epimerase/ManNAc kinase and CMP-sialic acid synthetase plus addition of N-acetylglucosamine 2.5.1.62 chlorophyll synthase synthesis expression of functional Arabidopsis thaliana chlorophyll synthase in cyanobacterial host Synechocystis sp. Synthesis of the plant chlorophyll synthase allows deletion of the otherwise essential native cyanobacterial gene. The interaction with membrane insertase YidC is maintained for the eukaryotic enzyme, HliD or Ycf39 do not copurify with ChlG 2.5.1.62 chlorophyll synthase synthesis expression of functional Chlamydomonas reinhardtii chlorophyll synthase in cyanobacterial host Synechocystis sp. Synthesis of the algal chlorophyll synthase allows deletion of the otherwise essential native cyanobacterial gene. The interaction with membrane insertase YidC is maintained for the eukaryotic enzyme, HliD or Ycf39 do not copurify with ChlG 2.5.1.63 adenosyl-fluoride synthase synthesis the enzyme is used in biotransformation reaction for production of fluorinated compounds, biotransformation protocols for coupled reaction systems, overview 2.5.1.63 adenosyl-fluoride synthase synthesis induced production of fluorosalinosporamide by replacing the chlorinase gene salL from Salinispora tropica with the fluorinase gene flA. Maximum yields of 4 mg/l fluorosalinosporamide production are detected at pH 6.0. The fluorosalinosporamide production yields are comparable to those attained through mutasynthesis. The major fluorinated component in the Salinispora tropica salL- flA+ extract is fluorosalinosporamide 2.5.1.63 adenosyl-fluoride synthase synthesis one-pot three-step continuous enzymatic synthesis of 5-fluoro-5-deoxy-D-ribose from ATP and L-methionine using S-adenosyl-L-methionine synthase, fluorinase and methylthioadenosine nucleosidase in the presence of fluoride ions. The conversion yield is 22.6% of 5-fluoro-5-deoxy-D-ribose from ATP, while the fluoride ions are generated from BF4 ionic liquids and/or the biodegradation of benzotrifluoride in the synthetic process 2.5.1.63 adenosyl-fluoride synthase synthesis preparation of sodium [18F]-fluoroacetate by generation of 5'-[18F]-fluoro-5'-deoxyadenosine by a fluorinase catalysed reaction of S-adenosyl-L-methionine with no carrier added [18F]-fluoride and then oxidation to [18F]-fluoroacetate by a Kuhn-Roth oxidative degradation. Na [18F]-fluoroacetate can be synthesized in 96% radiochemical purity 2.5.1.63 adenosyl-fluoride synthase synthesis a two-step radiolabelling protocol of a cancer relevant cRGD peptide is described where the fluorinase enzyme is used to catalyse a transhalogenation reaction to generate [18F]-5'-fluoro-5'-deoxy-2-ethynyladenosine, followed by a click reaction to an azide tethered cRGD peptide. This protocol offers efficient radiolabelling of a biologically relevant peptide construct in water at pH 7.8, 37°C in 2 hours, which is metabolically stable in rats and retains high affinity for alphavbeta3 integrin 2.5.1.63 adenosyl-fluoride synthase synthesis substrate tolerance allows a peptide cargo to be tethered to a 5'-chloro-5'-deoxynucleoside substrate for transhalogenation by the enzyme to a 5'-fluoro-5-deoxynucleoside. The reaction is successfully extended from that previously reported for a monomeric cyclic peptide (cRGD) to cargoes of dendritic scaffolds carrying two and four cyclic peptide motifs. The RGD peptide sequence is known to bind upregulated alphaVbeta3 integrin motifs on the surface of cancer cells and it is demonstrated that the fluorinated products have a higher affinity to alphaVbeta3 integrin than their monomeric counterparts. Tolerance of the fluorinase to these large multimeric peptides suggests that the C-2 position of a chlorinated nucleoside represents a site for the attachment of a diverse range of peptide cargos for use in enzymatic fluorination 2.5.1.63 adenosyl-fluoride synthase synthesis the enzyme is applicated in positron emission tomography (PET) as a result of its ability to catalyze C-18F bond formation in the presence of [18F]fluoride as the nucleophile. The enzymatic process has a technical advantage in the PET context because [18F]fluoride is generated in the cyclotron as a dilute solution in [18O]water, and the enzyme can use this form of aqueous fluoride directly. This feature obviates the usual requirement to secure dry [18F]fluoride by ion-exchange chromatography. The enzymatic process therefore offers the possibility to directly radiolabel biomolecules in the aqueous phase 2.5.1.63 adenosyl-fluoride synthase synthesis deveopment of a coupled chlorinase-fluorinase system for rapid transhalogenation. The chlorinase shares a substrate tolerance with the fluorinase, enabling these two enzymes to cooperatively produce 5'-fluorodeoxy-2-ethynyladenosine (5'-FDEA) in up to 91.6% yield in 1 h 2.5.1.66 N2-(2-carboxyethyl)arginine synthase synthesis clavulanic acid intermediates: deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid are heterologously produced in Streptomyces venezuelae recombinant using four sets of early genes from the clavulanic acid biosynthetic pathway 2.5.1.66 N2-(2-carboxyethyl)arginine synthase synthesis in an industrial clavulanic acid-overproducer Streptomyces clavuligerus DEPA, carboxyethylarginine synthase (Ceas2), clavaldehyde dehydrogenase (Car) and carboxyethyl-arginine betalactam synthase (Bls2) are overrepesented, whereas the enzymes of two other secondary metabolites are underrepresented 2.5.1.66 N2-(2-carboxyethyl)arginine synthase synthesis synthesis of clavulanic acid intermediates in Streptomyces venezuelae by expression of clavulanic acid biosynthesis genes carboxyethylarginine synthase (Ceas2), beta-lactam synthetase (Bls2), clavaminate synthase (Cas2), and proclavaminate amidinohydrolase (Pah2). The recombinant strain produces deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid 2.5.1.67 chrysanthemyl diphosphate synthase synthesis construction of the trans-chrysanthemic acid biosynthetic pathway in tomato fruits by expressing the chrysanthemyl diphosphate synthase gene, as well as an alcohol dehydrogenease (ADH) gene and aldehyde dehydrogenase (ALDH) gene from a wild tomato species, all under the control of the PG promoter. Trangenic tomato fruits show a concentration of trans-chrysanthemic acid that is about 1.7fold higher (by weight) than the levels of lycopene present in non-transgenic fruit, the level of lycopene in the transgenic plants is reduced by 68%. Ninety seven percent of the diverted DMAPP is converted to trans-chrysanthemic acid, but 62% of this acid is further glycosylated 2.5.1.67 chrysanthemyl diphosphate synthase synthesis transient coexpression of CDS with ADH2 which encodes an enzyme that oxidizes trans-chrysanthemol to trans-chrysanthemal, and ALDH1 which encodes an enzyme that oxidizes trans-chrysanthemal into trans-chrysanthemic acid, in Nicotiana benthamiana leaves results in the production of trans-chrysanthemic acid as well as several other side products. The majority (58%) of trans-chrysanthemic acid is glycosylated or otherwise modified 2.5.1.72 quinolinate synthase synthesis production of quinolinic acid from L-aspartate, dihydroxyacetone phosphate, and O2 by use of enzymes NadA and NadB 2.5.1.78 6,7-dimethyl-8-ribityllumazine synthase synthesis Bacillus subtilis is used for riboflavin production, also involving the enzyme RibH 2.5.1.80 7-dimethylallyltryptophan synthase synthesis 7-DMATS catalyzes regio- and stereospecific prenylations and can be used as efficient catalysts for chemoenzymatic synthesis of prenylated compounds 2.5.1.91 all-trans-decaprenyl-diphosphate synthase synthesis batch and fed-batch production of coenzyme Q10 2.5.1.91 all-trans-decaprenyl-diphosphate synthase synthesis for biotechnological production of coenzyme Q10 in recombinant Escherichia coli, three genetic manipulations are performed: heterologous expression of decaprenyl diphosphate synthase (Dps) from Agrobacterium tumefaciens, deletion of endogenous octaprenyl diphosphate synthase (IspB), and overexpression of 1-deoxy-D-xylulose synthase (Dxs). Expression of the dps gene and deletion of the ispB gene in Escherichia coli BL21(DE3)DELTAispB/pAP1 allows production of CoQ10 only. Coexpression of the dxs gene increases the specific content of CoQ10 2.5.1.91 all-trans-decaprenyl-diphosphate synthase synthesis ubiquinone-10 production using Agrobacterium tumefaciens dps gene in Escherichia coli by coexpression system. The expression coupled by dps and ubiCA is effective for increasing UQ-10 production by five times than that by expressing single dps gene in the shake flask culture 2.5.1.91 all-trans-decaprenyl-diphosphate synthase synthesis heterologous overexpression in Escherichia coli results in a level of ubiquinone-10 in the transformants that is greater than that of intrinsic ubiquinone-8 2.5.1.91 all-trans-decaprenyl-diphosphate synthase synthesis recombinant Escherichia coli harboring the decaprenyl diphosphate synthase gene produces CoQ10 as well as CoQ8 and CoQ9. The recombinant Escherichia coli harboring only the decaprenyl diphosphate synthase gene produces 0.21 mg/l of CoQ10, whereas Escherichia coli coexpressing decaprenyl diphosphate synthase and 1-deoxy-D-xylulose 5-phosphate synthase produces 0.37 mg/l of CoQ10. The CoQ10 fraction is increased from 15.86% for only decaprenyl diphosphate synthase to 29.78% for coexpression of decaprenyl diphosphate synthase and 1-deoxy-D-xylulose 5-phosphate synthase 2.5.1.94 adenosyl-chloride synthase synthesis deveopment of a coupled chlorinase-fluorinase system for rapid transhalogenation. The chlorinase shares a substrate tolerance with the fluorinase, enabling these two enzymes to cooperatively produce 5'-fluorodeoxy-2-ethynyladenosine in up to 91.6% yield in 1 h 2.5.1.96 4,4'-diapophytoene synthase synthesis construction of an in vitro C30 carotenoid pathway based on recombinant Escherichia coli farnesyl diphosphate synthase and 4,4'-diapophytoene synthase. The 4,4'-diapophytoene product is unstabel in aqueous buffer. In situ extraction using an aqueous-organic two-phase system results in a 100% conversion of isopentenyl diphosphate and dimethylallyl diphosphate into 4,4'-diapophytoene synthase 2.5.1.109 brevianamide F prenyltransferase (deoxybrevianamide E-forming) synthesis regularly C7-prenylated cyclic dipeptides are successfully obtained by using the member of the DMATS family, CdpC7PT from Aspergillus terreus 2.5.1.111 4-hydroxyphenylpyruvate 3-dimethylallyltransferase synthesis NovQ is capable of catalyzing the transfer of a dimethylallyl group to both phenylpropanoids, such as p-coumaric acid and caffeic acid, and the B-ring of flavonoids. NovQ can serve as a useful biocatalyst for the synthesis of prenylated phenylpropanoids and prenylated flavonoids 2.5.1.115 homogentisate phytyltransferase synthesis constitutive expression of genes encoding homogentisate phytyltransferase HPT and tocopherol cyclase TC individually and in combination in tobacco. The alpha-tocopherol content in transgenic tobacco plants expressing HPT, TC, and both HPT and TC is increased by 5.4-, 4.0-, and 7.1fold, respectively, when compared to the wild type 2.5.1.122 4-O-dimethylallyl-L-tyrosine synthase synthesis the specific 4-O-prenylation of tyrosine derivatives by SirD could find usage for production of prenylated tyrosine derivatives by chemoenzymatic synthesis 2.5.1.122 4-O-dimethylallyl-L-tyrosine synthase synthesis the broad alkyl donor specificity of SirD opens the door for the interrogation of the alkyl donor specificity of other prenyltransferases for potential utility as biocatalysts for differential alkylation applications 2.6.1.1 aspartate transaminase synthesis mutant enzyme R292E/L18H can use L-lysine as inexpensive amino donor for the production of L-homophenylalanine. The low solubility of product L-homophenylalanine and spontaneous cyclization of 2-keto-6-aminocaproate drive the reaction completely towards production of L-homophenylalanine 2.6.1.1 aspartate transaminase synthesis codon-optimized expression in Escherichia coli with His6-tag and purification. The enzyme activity of purified aspartate aminotransferase reaches 150000 U/l. The preparation shows high stability during long-term storage at -20ºC 2.6.1.1 aspartate transaminase synthesis expression in 2-phenylethanol producing strain Saccharomyces cerevisiae YS58, and use of the strain for the coproduction of 2-phenylethanol and L-homophenylalanine via a fermentation process. The L-homophenylalanine productivity of the recombinant Saccharomyces cerevisiae improves by 78.9% in comparison to the wild-type. High yields of 43.7 mM L-homophenylalanine and 32.4 mM 2-phenylethanol are achieved 2.6.1.2 alanine transaminase synthesis for synthesis of monoclonal antibodies in Chinese hamster ovary cells, cooverexpression of alanine aminotransferase in a taurine transporter-overexpressing host cell line gives a higher monoclonal anitbody yield in a shorter culture period. Forced cooverexpression of taurine transporter TAUT and ALT1 in results in a higher proliferation, with an ideal balance between cell viability and productivity 2.6.1.16 glutamine-fructose-6-phosphate transaminase (isomerizing) synthesis the enzyme is useful in bioproduction of N-acetylglucosamine (GlcNAc), a nutraceutical that has various applications in healthcare 2.6.1.B16 amine transaminase synthesis amine transaminase (ATA) catalyzing stereoselective amination of prochiral ketones is an attractive alternative to transition metal catalysis 2.6.1.B16 amine transaminase synthesis the covalently immobilized enzyme is used for amine bioproduction in batch and flow reaction systems, method evaluation, overview. Produced vanillylamine is a valuable building block for the synthesis of natural products, such as capsaicinoids 2.6.1.17 succinyldiaminopimelate transaminase synthesis overexpression of enzyme, increase in production of L-lysine, 10fold increase in enzyme activity of strain 2.6.1.B17 4-hydroxyphenylglycine transaminase synthesis engineering of Escherichia coli for biosynthesis of L-phenylglycine from glucose. The enzymes HmaS (L-4-hydroxymandelate synthase), Hmo (L-4-hydroxymandelate oxidase) and HpgT (L-4-hydroxyphenylglycine transaminase) are heterologously expressed in Escherichia coli. HpgT conversing phenylglyoxylate to L-phenylglycine uses an unusual aminodonor L-phenylalanine, which releases another phenylpyruvate as the substrate of HmaS. A recycle-reaction maximizes the utilization of precursor phenylpyruvate. After deletionof tyrB and aspC, L-phenylglycine yield increases by 12.6fold. The L-phenylglycine yield is further improved by 14.9fold after enhancing hmaS expression 2.6.1.18 beta-alanine-pyruvate transaminase synthesis enzymatic synthesis of enantiomerically pure (R)-1-phenylethanol and (R)-alpha-methylbenzylamine from racemic alpha-methylbenzylamine in a coupled assay using the omega-transaminase, alcohol dehydrogenase, and glucose dehydrogenase 2.6.1.18 beta-alanine-pyruvate transaminase synthesis enzymatic synthesis of enantiomerically pure D-beta-amino-n-butyric acid from racemic beta-amino-n-butyric acid 2.6.1.18 beta-alanine-pyruvate transaminase synthesis optically pure valinol (ee >99%) is prepared employing different omega-transaminases from the corresponding prochiral hydroxy ketone. By the choice of the enzyme the (R)- as well as the (S)-enantiomer are accessible 2.6.1.18 beta-alanine-pyruvate transaminase synthesis the metabolic removal of pyruvate can be exploited to shift the unfavorable equilibrium of the omega-transaminase-catalyzed amination of ketones using alanine as an amino donor. Vigorous aeration should lead to substantial loss of a volatile ketone substrate without a condenser incorporated into a reactor set-up. A reactor design to support aerobic conditions without undesirable loss of a ketone substrate should be considered 2.6.1.18 beta-alanine-pyruvate transaminase synthesis a prominent example for beta-amino acids in pharmaceuticals is beta-phenylalanine, present in the natural substance paclitaxel (Taxol), which is a prominent anti-cancer agent. Method development for production of chiral beta-phenylalanine ethyl ester, which can be chemically or enzymatically hydrolyzed to (R)- or (S)-phenylalanine. Asymmetric synthesis of the beta-phenylalanine ethyl ester an upscaling, overview 2.6.1.B21 (R)-amine:2-oxo acid transaminase synthesis amine transaminase (ATA) catalyzing stereoselective amination of prochiral ketones is an attractive alternative to transition metal catalysis 2.6.1.B21 (R)-amine:2-oxo acid transaminase synthesis development of a one-pot, trienzymatic cascade comprising an (R)-specific omega-transaminase, an amine dehydrogenase, and a formate dehydrogenase for the economical and ecofriendly synthesis of (R)-chiral amines. Using inexpensive ammonium formate as the sole sacrificial agent, the established cascade system enables efficient omega-transaminase-mediated (R)-amination of various ketones, with high conversions and excellent enantiomeric excess (over 99%), water and CO2 are the only waste products 2.6.1.B21 (R)-amine:2-oxo acid transaminase synthesis in another reaction for deracemization using enantiocomplementary transaminases, the oxidative deamination step is carried out by alpha-transaminase such as branched-chain transaminase (EC 2.6.1.42) and D-amino acid transaminase (EC 2.6.1.21). It is notable that substitution of alpha-transaminase with alanine dehydrogenase in the deracemization method for production of D-amino acids using L-alanine dehydrogenase (AlaDH), D-selective omega-transaminase (omega-TA) ARTAmut, and NADH oxidase (NOX) eliminates the need for an expensive 2-oxoacid cosubstrate. Feasibility of the stereoinversion reaction is dependent on enzyme activities of AlaDH and onega-TA for L-amino acids and its keto acids, respectively. ARTAmut substrate specificity allows various oxoacids as substrates 2.6.1.B21 (R)-amine:2-oxo acid transaminase synthesis pyridoxal-5'-phosphate (PLP)-dependent transaminases are industrially important enzymes catalyzing the stereoselective amination of ketones and keto acids. Transaminases of PLP fold type IV are characterized by (R)- or (S)-stereoselective transfer of amino groups, depending on the substrate profile of the enzyme 2.6.1.28 tryptophan-phenylpyruvate transaminase synthesis one-pot synthesis of di-demethylasterriquinone D from L-tryptophan and phenylpyruvic acid by use of enzymes TdiA and TdiD 2.6.1.29 diamine transaminase synthesis biocatalytic transamination with near-stoichiometric inexpensive amine donors mediated by bifunctional mono- and di-amine transaminases. Both monoamine and diamine transaminase activity is reported, allowing conversion of a wide range of target ketone substrates with just a small excess of amine donor. The diamine co-substrates (putrescine, cadaverine or spermidine) are bio-derived and the enzyme system results in very little waste, making it a greener strategy for the production of valuable amine fine chemicals and pharmaceuticals 2.6.1.29 diamine transaminase synthesis diamine donors are used for effective equilibrium displacement of transaminase-mediated biotransformations. Whole-cell biotransformation using engineered Corynebacterium glutamicum cells, that produce diamine donors, is evaluated 2.6.1.30 pyridoxamine-pyruvate transaminase synthesis pyridoxamine production by bioconversion is generally preferable for environmental and energetic aspects compared to chemical synthesis. Pyridoxamine is produced from pyridoxine, a readily and economically available starting material, by bioconversion using a Rhodococcus expression system 2.6.1.33 dTDP-4-amino-4,6-dideoxy-D-glucose transaminase synthesis engineering of Escherichia coli to divert the flow of carbon flux from glucose-1-phosphate to thymidine diphosphate 4-keto 4,6-dideoxy-D-glucose (dTKDG), an intermediate of various dTDP-sugars. Glucose phosphate isomerase, glucose-6-phosphate dehydrogenase and uridylyltransferase genes are deleted while dTDP-D-glucose synthase and dTDP-Dglucose 4,6-dehydratase are overexpressed to produce a pool of dTKDG in the cell cytosol. The flow of dTKDG is further diverted to dTDP-D-viosamine, dTDP 4-amino 4,6-dideoxy-D-galactose, and dTDP 3-amino 3,6-dideoxy-D-galactose sugars using sugar aminotransferases gerB, wecE, and fdtB, respectively, from different sources. These sugar moieties are transferred to the 3-hydroxyl position of quercetin and kaempferol with the help of Arabidopsis thaliana glycosyltransferase ArGT3 2.6.1.42 branched-chain-amino-acid transaminase synthesis enzyme can be used for asymmetric synthesis of a range of non-natural amino acids such as L-norleucine, L-norvaline and L-neopentylglycine 2.6.1.42 branched-chain-amino-acid transaminase synthesis isobutanol and other branched-chain higher alcohols (BCHAs) are promising advanced biofuels derived from the degradation of branched-chain amino acids (BCAAs). The yeast Saccharomyces cerevisiae is a particularly attractive host for the production of BCHAs due to its high tolerance to alcohols and prevalent use in the bioethanol industry. Degradation of BCAAs begins with transamination reactions, catalyzed by branched-chain amino acid transaminases (BCATs) located in the mitochondria (Bat1p) and cytosol (Bat2p) 2.6.1.42 branched-chain-amino-acid transaminase synthesis pyridoxal-5'-phosphate (PLP)-dependent transaminases are industrially important enzymes catalyzing the stereoselective amination of ketones and keto acids. Transaminases of PLP fold type IV are characterized by (R)- or (S)-stereoselective transfer of amino groups, depending on the substrate profile of the enzyme 2.6.1.42 branched-chain-amino-acid transaminase synthesis usage of PsBCAT in a coupled reaction with Bacillus subtilis ornithine aminotransferase (BsOrnAT), applied to synthesize L-tert-leucine and L-norvaline. The reaction mixture contains 0.5 M Tris-HCl buffer, pH 8.5, 0.1 M trimethylpyruvate or 2-oxovalerate, 20 mM sodium glutamate, 80 mM L-ornithine monohydrochloride, 0.02 mM PLP, 0.2 mg PsBCAT, and 0.05 mg BsOrnAT enzyme in a total volume of 5 ml at 30°C 2.6.1.48 5-aminovalerate transaminase synthesis production of 5-aminovalerate and glutarate in Escherichia coli. Endogenous over-production of the precursor, lysine, is first achieved through metabolic deregulation of its biosynthesis pathway by introducing feedback resistant mutants of aspartate kinase III and dihydrodipicolinate synthase. Further disruption of native lysine decarboxylase activity limits cadaverine by-product formation. Co-expression of lysine monooxygenase and 5-aminovaleramide amidohydrolase then results in the production of 0.86 g/l 5-aminovalerate in 48 h. The additional co-expression of glutaric semialdehyde dehydrogenase and 5-aminovalerate aminotransferase leads to the production of 0.82 g/l glutarate under the same conditions. Yields on glucose are 71 and 68 mmol/mol for 5-aminovalerate and glutarate, respectively 2.6.1.48 5-aminovalerate transaminase synthesis production of 5-aminovalerate and glutarate in recombinant Escherichia coli. When the davAB genes encoding delta-aminovaleramidase and lysine 2-monooxygenase, respectively, are introduced into a recombinant Escherichia coli strain allowing enhanced L-lysine synthesis, 0.27 and 0.5 g/l of 5-aminovalerate are produced directly from glucose by batch and fed-batch cultures, respectively. Further conversion of 5-aminovalerate into glutarate can be demonstrated by expression of the Pseudomonas putida gabTD genes encoding 5-aminovalerate aminotransferase and glutarate semialdehyde dehydrogenase. A recombinant Eschrerichia coli strain expressing the davAB and gabTD genes cultured in a medium containing 20 g/l glucose,10 g/l L-lysine and 10 g/l alpha-ketoglutarate, produces 1.7 g/l of glutarate 2.6.1.48 5-aminovalerate transaminase synthesis expression of the Pseudomonas putida davAB genes encoding delta-aminovaleramidase and lysine 2-monooxygenase, respectively, in Escherichia coli. When the davAB genes are introduced into recombinant E. coli strain XQ56 allowing enhanced L-lysine synthesis, 0.27 and 0.5g/l of 5-aminovalerate are produced directly from glucose by batch- and fed-batch cultures, respectively. Further conversion of 5-aminovalerate into glutarate can be achieved by expression of the Pseudomonas putida gabTD genes encoding 5-aminovalerate aminotransferase and glutarate semialdehyde dehydrogenase. In a medium containing 20g/l glucose, 10g/l L-lysine and 10g/l alpha-ketoglutarate, this strain produces 1.7g/l of glutarate 2.6.1.52 phosphoserine transaminase synthesis deletion of L-serine dehydratase in combination with overexpression of enzyme, L-serine insensitive 3-phosphoglycerate dehydrogenase, and phosphoserine phosphatase yields up to 86 mM L-serine in the culture medium 2.6.1.57 aromatic-amino-acid transaminase synthesis asymmetric synthesis of L-homophenylalanine and L-phenylglycine from 2-oxo-4-phenylbutyrate and phenylglyoxylate, respectively, using L-glutamate as an amino donor. AroATBs can be used as a biocatalyst for the synthesis of unnatural L-amino acids by taking advantage of the large solubility difference between substrate and product at higher temperatures to accelerate an equilibrium shift favoring product formation 2.6.1.57 aromatic-amino-acid transaminase synthesis asymmetric synthesis of the nonproteinogenic amino acids (2S)-2-amino-4-oxo-4-phenylbutyric and (3E,2S)-2-amino-4-phenylbutenoic acid by recombinant Escherichia coli whole cells overexpressing aromatic transaminase from Enterobacter sp. BK2K-1 (AroATEs) in high yields (68–78%) and high enantiomeric purity (>99%) using L-aspartic acid as an amino donor 2.6.1.57 aromatic-amino-acid transaminase synthesis deletion of the Aro8 gene stimulates phenylethanol production when combined with other mutations that deregulate aromatic amino acid biosynthesis, i.e. expression of phenylalanine and tyrosine feedback-insensitive DAHP synthase. The resulting engineered Saccharomyces cerevisiae strain produces more than 3 mM phenylethanol from glucose during growth on a simple synthetic medium 2.6.1.70 aspartate-phenylpyruvate transaminase synthesis L-phenylalanine production as a building block of aspartame 2.6.1.72 D-4-hydroxyphenylglycine transaminase synthesis metabolic engineering of the Escherichia coli L-phenylalanine pathway for the production of D-phenylglycine. Expression of hydroxymandelate synthase from Amycolatopsis orientalis, hydroxymandelate oxidase from Streptomyces coelicolor and hydroxyphenylglycine aminotransferase from Pseudomonas putida in Escherichia coli 2.6.1.72 D-4-hydroxyphenylglycine transaminase synthesis functional expression in Pichia pastoris. Co-expression of Escherichia coli chaperonins GroEL-GroES with the enzyme gene dramatically improves the soluble active enzyme production. Increasing gene dosage of both the enzyme and those of the chaperones further increases functional protein yield up to 14400fold higher than when the enzyme is expressed alone. Optimization of cultivation condition further increases activity yield from the best co-expressing strain by 1.2fold 2.6.1.72 D-4-hydroxyphenylglycine transaminase synthesis N-terminus of PhgAT is genetically fused with short peptides from a ferredoxin enzyme of halophilic archaeon, Halobacterium salinarum. The fused enzymes display a reduced pI and increase in solubility in TEMP (pH 7.6) storage, and in CAPSO (pH 9.5) reaction buffers, respectively. All the fused PhgAT display higher enzymatic reaction rates than the wild-type at all concentrations of L-glutamate used. the halophilic fusion significantly increases the tolerance of PhgAT in the presence of NaCl and KCl 2.6.1.76 diaminobutyrate-2-oxoglutarate transaminase synthesis production of ectoine in Escherichia coli. The Escherichia coli regulatory protein AraC is engineered to recognize ectoine as it snon-natural effector and to activate transcription upon ectoine binding. The ectoine biosynthetic cluster from Halomonas elongata is cloned into Escherichia coli. By engineering the rate-limiting enzyme L-2,4-diaminobutyric acid aminotransferase EctB, ectoine production and the specific activityof the EctB mutant are increased 2.6.1.82 putrescine-2-oxoglutarate transaminase synthesis a route for the production of GABA via putrescine in Corynebacterium glutamicum. A putrescine-producing recombinant Corynebacterium glutamicum strain is converted into a GABA producing strain by heterologous expression of putrescine transaminase PatA and gamma-aminobutyraldehyde dehydrogenase PatD genes from Escherichia coli. The resultant strain produces 5.3 g per l of GABA. GABA production is improved further by adjusting the concentration of nitrogen in the culture medium, by avoiding the formation of the by-product N-acetylputrescine and by deletion of the genes for GABA catabolism and GABA re-uptake. GABA accumulation by this strain is increased by 5% to 8.0 g per l, and the volumetric productivity is increased to 0.31 g per l and h 2.6.1.82 putrescine-2-oxoglutarate transaminase synthesis enzyme prefers diaminoalkanes as substrates and thereby generates cyclic imines from the omega-amino aldehyde intermediates. The addition of a mild chemical reducing agent rapidly reduces the imine intermediate in situ to furnish a range of N-heterocycle products 2.6.1.82 putrescine-2-oxoglutarate transaminase synthesis putrescine transaminase Pp-SpuC application in the transamination of a comprehensive range of ketones/aldehydes for the production of a variety of benzylamine derivatives. Applications of enzyme Pp-SpuC as a biocatalyst with broad substrate tolerance 2.6.1.90 dTDP-3-amino-3,6-dideoxy-alpha-D-galactopyranose transaminase synthesis synthesis of TDP-D-ravidosamine, necessary for future in vitro glycosylation assays. TDP-D-ravidosamine is the anticipated sugar donor substrate of RavGT (the glycosyltransferase that links D-ravidosamine to the polyketide derived backbone defuco-gilvocarcin V). Defuco-gilvocarcin V exhibits superior anticancer/antibacterial activities 2.6.1.90 dTDP-3-amino-3,6-dideoxy-alpha-D-galactopyranose transaminase synthesis engineering of Escherichia coli to divert the flow of carbon flux from glucose-1-phosphate to thymidine diphosphate 4-oxo 6-deoxy-D-glucose (dTKDG), an intermediate of various dTDP-sugars. Glucose phosphate isomerase, glucose-6-phosphate dehydrogenase and uridylyltransferase genes are deleted while dTDP-D-glucose synthase and dTDP-D-glucose 4,6-dehydratase are overexpressed to produce a pool of dTKDG in the cell cytosol. The flow of dTKDG is further diverted to dTDP-D-viosamine, dTDP 4-amino 4,6-dideoxy-D-galactose, and dTDP 3-amino 3,6-dideoxy-D-galactose sugars using sugar aminotransferases gerB, wecE, and fdtB, respectively, from different sources. These sugar moieties are transferred to the 3-hydroxyl position of quercetin and kaempferol with the help of Arabidopsis thaliana glycosyltransferase ArGT3 2.6.1.116 6-aminohexanoate aminotransferase synthesis a coupled system of NylD1 and adipate semialdehyde dehydrogenase NylE1 allows to quantify the aminotransferase activity and enables the conversion of 6-aminohexaoate to adipate via adipate semialdehyde with a yield of >90% 2.6.99.3 O-ureido-L-serine synthase synthesis expression of a D-cycloserine biosynthetic gene cluster consisting of 10 open reading frames (dcsA to dcsJ) from Streptomyces lavendulae ATCC 11924 in Escherichia coli. When L-serine and hydroxyurea, the precursors of D-cycloserine, are incubated together with the Escherichia coli resting cell suspension, the cells produce significant amounts of D-cycloserine (350 microM). To increase the productivity, the dcsJ gene, which might be responsible for the excretion, is connected downstream of the four genes, resulting in production of D-cycloserine at 660 microM. To repress the side catalytic activity of DcsD, i.e. the formation of L-cysteine from O-acetylserine and H2S, Escherichia coli chromosomal cysJ and cysK genes, encoding the sulfite reductase alpha subunit and O-acetylserine sulfhydrylase, respectively, are disrupted. The final strain produces 980 microM D-cycloserine 2.7.1.1 hexokinase synthesis overexpression of HXK1 in a highly modified strain of Yarrowia lipolytica W29 engineered to optimize oil production, combined with Saccharomyces cerevisiae invertase gene expression. For the recombinant strain, sucrose is a better substrate than either of its building blocks, glucose or fructose. Over its 96 h of growth in the bioreactors, this strain produces 9.15g/l of lipids, yielding 0.262g/g of biomass 2.7.1.4 fructokinase synthesis enzyme expression strategy for enhancing amino acid production 2.7.1.5 rhamnulokinase synthesis production of L-Rhamnulose (6-deoxy-L-arabino-2-hexulose) and L-fuculose (6-deoxy-L-lyxo-2-hexulose) from L-rhamnose and L-fucose. First, isomerization of L-rhamnose to L-rhamnulose, or L-fucose to L-fuculose is combined with a targeted phosphorylation reaction catalyzed by L-rhamnulose kinase (RhaB). The by-products (ATP and ADP) are selectively removed by silver nitrate precipitation method. In the second step, the phosphate group is hydrolyzed to produce L-rhamnulose or L-fuculose with purity exceeding 99% in more than 80% yield 2.7.1.6 galactokinase synthesis enzyme mutant Y371H/M173L, substrate range is much wider than wild-type, generation of unnatural sugar 1-phosphates 2.7.1.6 galactokinase synthesis LgGalK and galactose oxidase variant M1 are combined in a one-pot, two-step system to synthesize 6-oxogalactose-1-phosphate and 6-oxo-2-fluorogalactose-1-phosphate, which are subsequently used to produce a panel of 30 substituted 6-aminogalactose-1-phosphate derivatives by chemical reductive amination in a one-pot, three-step chemoenzymatic process 2.7.1.11 6-phosphofructokinase synthesis expression of a modified 6-phosphofructo-1-kinase in a citrate producing Aspergillus niger strain in combination with cis-aconitate decarboxylase CadA from Aspergillus terreus to study the effect on the production of itaconic acid. The combined expression of pfkA and cadA results in increased citrate levels, but does not show increased itaconic acid levels. The combined expression of pfkA with the itaconic acid biosynthetic cluster, consisiting of cis-aconitate decarboxylase cadA, a putative mitochondrial transporter mttA and a putative plasmamembrane transporter mfsA, results in significantly increased itaconic acid production at earlier time points and significant increase in itaconic acid productivity. The maximum itaconic acid productivity reached is 0.15 g per l and h 2.7.1.11 6-phosphofructokinase synthesis expression of a truncated version of the gene encoding 6-phospho-1-fructokinase (tpfkA) along with its activator pkaC in Lactobacillus reuteri. Growth of the transformants at elevated glucose concentrations in the presence of fructose results in improved assimilation of the provided carbohydrates and a significant increase in the overall fermentation productivities. At the highest tested levels of glucose and fructose (75 g/l each), the transformant strain shows a 4fold increase in 6-phosphofructo-1-kinase activity and a 2.3fold increase in the glycolytic flux. The mannitol yield is 56 g/l compared to 10 g/l in the parental strain, and the lactate yield is 21 g/l (3.5 g/l in the parental strain). A high NADH/NAD+ratio occurrs under increased glycolytic flux conditions and facilitates the efficient conversion of fructose to mannitol 2.7.1.11 6-phosphofructokinase synthesis overexpression of both the 6-phosphofructokinase pfkA and pyruvate kinase pykA genes increase intracellular concentrations of ATP and NADH and also resistance to butanol toxicity. Marked increases of butanol and ethanol production, but not acetone, are observed in batch fermentation. The butanol and ethanol concentrations are 29.4 and 85.5% higher, respectively, in the fermentation by the double-overexpressing strain than the wild-type strain. In fed-batch fermentation using glucose, the butanol and total solvent (acetone, butanol, and ethanol) concentrations reach as high as 19.12 and 28.02 g/l, respectively 2.7.1.11 6-phosphofructokinase synthesis sodium ions activated phosphofructokinase leads to enhanced D-lactic acid production by Sporolactobacillus inulinus using sodium hydroxide as a neutralizing agent 2.7.1.17 xylulokinase synthesis an effective conversion of xylulose to xylulose 5-phosphate catalyzed by the xylulokinase in Saccharomyces cerevisiae is considered to be essential for the development of an efficient and accelerated ethanol fermentation process from xylulose 2.7.1.17 xylulokinase synthesis commercial production of bioethanol from xylose using enzyme mutants with improved solubility expressed in presence of chaperonins GroEL-GroES 2.7.1.17 xylulokinase synthesis overexpression of enzyme, 3fold higher expression does not result in any increase in rate of growth or xylose metabolism 2.7.1.17 xylulokinase synthesis construction of a Saccharomyces cerevisiae strain overexpressing GRE3-encoded NADPH-dependent aldose reductase and xylulokinase with a mutated strictly NADP+-dependent Pichia stipitis xylitol dehydrogenase. The recombinant strain efficiently ferments xylose and glucose mixture and the ethanol production is 21.4% higher than that of an isogenic constructed reference strain expressing Pichia stipitis xylose reductase. The yield of ethanol production increases from 0.395 g ethanol/g sugar to 0.435 g ethanol/g sugar after glucose depletion. Xylitol accumulation (0.6% of total sugar) is considerably lower than that of the reference strain (4.8% of total sugar) 2.7.1.17 xylulokinase synthesis silencing of gene expression with an antisense construct decreases D-xylulokinase activity after 48 h of incubation, leading to an increase in xylitol production from undetectable levels in wild-type to 8.6 mM 2.7.1.17 xylulokinase synthesis xylulokinase and xylose isomerase are overexpressed in Klebsiella oxytoca HP1 to enhance hydrogen production by the fermentation of xylose. The recombinant strains exhibit higher enzyme activity compared with the wild-type strain. Hydrogen production from pure xylose, xylose/glucose mixtures and bamboo stalk hydrolysate is significantly enhanced. The hydrogen yield per mole substrate in strains expressing xylulokinase and xylose isomerase reaches 1.93 and 2.46 mol H2/mol xylose, respectively in pure xylose, while the value for the wild strain is 1.68 mol H2/mol xylose. Relative to the wild type, hydrogen yield from 1 g of preprocessed bamboo powder increases by 33.04 upon overexpression of xylulokinase 2.7.1.21 thymidine kinase synthesis enzyme can be useful in drug design 2.7.1.23 NAD+ kinase synthesis combined activation of PntAB and YfjB leads to 28% and 22% increase of aerobic isobutanol titer and yield, resulting in production of 10.8 g/l isobutanol in 24 h with a yield of 0.62 mol/mol 2.7.1.23 NAD+ kinase synthesis overexpressing NAD kinase is a useful metabolic engineering strategy to improve NADPH supply and isoleucine biosynthesis 2.7.1.23 NAD+ kinase synthesis preparation of a superparamagnetic NAD kinase catalyst to synthesize NADP in vitro 2.7.1.26 riboflavin kinase synthesis immobilized enzyme is effective for phosphorylating riboflavin and numerous riboflavin analogs and provides a facile method for preparing exclusively other synthetic methods, the 5'-phosphates 2.7.1.26 riboflavin kinase synthesis the enzyme is used for the preparation of flavin mononucleotide (FMN) and FMN analogues from their corresponding riboflavin precursors, which is performed in a two-step procedure. After initial enzymatic conversion of riboflavin to FAD by the bifunctional FAD synthetase, the adenyl moiety of FAD is hydrolyzed with snake venom phosphodiesterase to yield FMN. The engineered FAD synthetase from Corynebacterium ammoniagenes with deleted N-terminal adenylation domain is a biocatalyst that is stable and efficient for direct and quantitative phosphorylation of riboflavin and riboflavin analogues to their corresponding FMN cofactors at preparative-scale 2.7.1.30 glycerol kinase synthesis methodology of dosage of glycerol kinase by response surface modeling of the enzymatic reaction. This low cost method for glycerol kinase dosage in a sequence of reactions is of great importance for many industries, like food, sugar and alcohol. Response surface modeling shows to be an adequate approach for modeling the reaction and optimization of reaction conditions to maximize glycerol kinase activity 2.7.1.33 pantothenate kinase synthesis synthesis of (carboxyl-18O)phosphopantothenate 2.7.1.35 pyridoxal kinase synthesis production of cadaverine (1,5-diaminopentane) can be done by fermentation or direct bioconversion and plays an important role as a building block of polyamides. Lysine decarboxylase (CadA) transforms L-lysine to cadaverine and pyridoxal 5'-phosphate (PLP) can increase the conversion rate and yield as a cofactor. Biotransformation of cadaverine using whole Escherichia coli cells that overexpress lysine decarboxylase has many merits, such as the rapid conversion of L-lysine to cadaverine, possible application of high concentration reactions up to the molar level, production of less byproduct, and potential reuse of the enzyme by immobilization. But the supply of PLP, an essential cofactor of lysine decarboxylase, is the major bottleneck in this system. Among various PLP systems examined, pyridoxal kinase (PdxY) shows the highest conversion of PL to PLP, resulting in more than 60 % conversion of L-lysine to cadaverine with lysine decarboxylase. Method evaluation and optimization. Interconversion of interconvertion of pyridoxal and pyridoxamine has to be controlled 2.7.1.36 mevalonate kinase synthesis production of amorpha-4,11-diene by an engineered strain of Escherichia coli containing codon-optimized MevT and amorphadiene synthase operons, and additional copies of mevalonate kinase and amorphadiene synthase genes, which could be identified as rate-limiting enzymes 2.7.1.49 hydroxymethylpyrimidine kinase synthesis useful reagent for the preparation of intermediates on the thiamin biosynthetic pathway 2.7.1.52 fucokinase synthesis the enzyme is a valuable biochemical tool to prepare activated L-fucose derivatives for fucosylation reactions, enzyme is valuable for the formation of radiolabeled fucose 1-phosphate 2.7.1.63 polyphosphate-glucose phosphotransferase synthesis great potentials of directed evolution in obtaining high-performance BioBricks suitable for the in vitro synthetic biology platform 2.7.1.63 polyphosphate-glucose phosphotransferase synthesis production of myo-inositol from glucose by a novel trienzymatic cascade of polyphosphate glucokinase, inositol 1-phosphate synthase and inositol monophosphatase. myo-Inositol (inositol) is important in the cosmetics, pharmaceutical and functional food industries. The conversion ratio from glucose to inositol reaches 90%, which is promising for the enzymatic synthesis of inositol without ATP supplementation 2.7.1.66 undecaprenol kinase synthesis one-pot reaction leading to undecaprenyl diphosphate disaccharide and starting from undecaprenol, ATP, and the UDP-sugars using the kinase, PglC and PglA 2.7.1.71 shikimate kinase synthesis the aroK enzyme knockout strain of Escherichia coli is useful for shikimate production 2.7.1.73 inosine kinase synthesis the recombinant strain, which expresses gene gsk and has both inosine kinase activity and ATP-regenerating activity, is used to induce the phosphorylation of inosine to produce inosine 5'-monophosphate, which is widely used as a flavor enhancer 2.7.1.77 nucleoside phosphotransferase synthesis method for transformation of nucleosides into 5’-monophosphates based on the shift in the equilibrium state of the reaction. Extent of nucleoside transformation is 41-91% 2.7.1.78 polynucleotide 5'-hydroxyl-kinase synthesis synthesis of photoreactive oligonucleotides, containing active groups at the 5'-end phosphate, as tools for photoaffinity modification of DNA-metabolizing enzymes and factors, utilization of gamma-substituted ATP derivatives 2.7.1.78 polynucleotide 5'-hydroxyl-kinase synthesis due to reversibility of the reaction, the bacteriophage can be utilized for exchange of labeled phosphate groups between 2 substrates 2.7.1.78 polynucleotide 5'-hydroxyl-kinase synthesis enzyme is an important tool in the synthesis of genes corresponding to yeast alanine tRNAand to precursor tyrosine tRNA of Escherichia coli 2.7.1.78 polynucleotide 5'-hydroxyl-kinase synthesis production of beta,gamma-imidoadenylyl 5'-tetraphosphate by using beta,gamma-imidoadenylyl 5'-triphosphate as substrate in the reverse reaction 2.7.1.78 polynucleotide 5'-hydroxyl-kinase synthesis construction of a catalytically useful ribozyme 2.7.1.86 NADH kinase synthesis NADH kinase can be employed as an effective metabolic manipulation target to improve poly-3-hydroxybutyrate synthesis 2.7.1.140 inositol-tetrakisphosphate 5-kinase synthesis overexpression of enzyme does not increase the level of inositolpentakisphosphate, gene silencing of enzyme results in decreased inositol pentakisphosphate and inositol hexakisphosphate levels 2.7.1.145 deoxynucleoside kinase synthesis enzyme can be utilized in industrial large-scale production of dNTPs and analogues, used for DNA synthetic reactions, giving a higher yield and less toxic byproducts compared to chemical processes 2.7.1.145 deoxynucleoside kinase synthesis enzyme can be utilized in industrial large-scale production of dNTPs and analogues, used forDNA synthetic reactions, giving a higher yield and less toxic byproducts compared to chemical processes 2.7.1.145 deoxynucleoside kinase synthesis the cross-linked DmdNK preparation is used for the preparative synthesis of arabinosyladenine monophosphate (ara-AMP) and fludarabine monophosphate (FaraAMP) 2.7.1.157 N-acetylgalactosamine kinase synthesis synthesis of UDP-GalNAc with high yield from GalNAc, UTP and ATP using recombinant human GalNAc kinase GK2 and UDP-GalNAc pyrophosphorylase AGX1 2.7.1.162 N-acetylhexosamine 1-kinase synthesis an N-acetylhexosamine 1-kinase from Bifidobacterium infantis (NahK_15697), a guanosine 5'-diphosphate (GDP)-mannose pyrophosphorylase from Pyrococcus furiosus (PFManC), and an Escherichia coli inorganic pyrophosphatase (EcPpA) are used efficiently for a one-pot three enzyme synthesis of GDP-mannose, GDP-glucose, their derivatives, and GDP-talose from simple monosaccharides and derivatives in preparative scale 2.7.1.162 N-acetylhexosamine 1-kinase synthesis the enzyme is used in the chemoenzymatic synthesis of UDP-GlcNAc and UDP-GlcNTFA attempted by three recombinant enzymes in a fed batch system, overview. The process can be used for the development of an efficient scalable process for the supply of UDP-monosaccharide donors for oligosaccharide synthesis 2.7.1.165 glycerate 2-kinase synthesis synthesis of 2-phospho-D-glycerate, quickly, inexpensively 2.7.1.165 glycerate 2-kinase synthesis a microbial production platform in Escherichia coli to synthesize D-glyceric acid from D-galacturonate represents an alternative for the production of D-glyceric acid, an industrially relevant chemical, that addresses current challenges in using acetic acid bacteria for its synthesis: increasing yield, enantio-purity and biological stability, overview 2.7.1.165 glycerate 2-kinase synthesis straightforward phosphorylation reaction by enzyme GCK and subsequent product isolation enables the preparation of enantiomerically pure D-glycerate 2-phosphate. This phosphorylation reaction, using recombinant glycerate-2-kinase, yields D-glycerate 2-phosphate in fewer reaction steps and with higher purity than chemical routes 2.7.1.175 maltokinase synthesis enzymatic synthesis of maltose 1-phosphate 2.7.1.185 mevalonate 3-kinase synthesis isobutene production rates are 507 pmol/min*g cells using Escherichia coli cells expressing the enzyme and 2880 pmol/min*mg protein with the purified histidine-tagged enzyme. Isobutene is a small, highly reactive molecule, used extensively as a platform chemical to manufacture a wide variety of products including fuel additives, rubbers, and speciality chemicals 2.7.2.1 acetate kinase synthesis protocol for overproduction of enzyme 2.7.2.4 aspartate kinase synthesis the enzyme is a potential target for improved production of L-lysine. Recombinants of Corynebacterium glutamicum with feedback resistant aspartate kinase would be a potential option to increase the L-lysine production by biotechnological process for industrial application 2.7.2.7 butyrate kinase synthesis butanoate kinase and phosphotransbutyrylase are successfully exploited for in vitro synthesis of 3-hydroxybutyryl-CoA, 4-hydroxybutyryl-CoA, 4-hydroxyvaleryl-CoA and poly(hydroxyalkanoic acid). Combination of butanoate kinase, phosphotransbutyrylase and poly(hydroxyalkanoic acid) synthase of Chromatium vinosum establishes a new system for in vitro synthesis of poly(3-hydroxybutyric acid) 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis - 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis used for an ATP regeneration system for acetyl-CoA synthesis 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis used in place of pyruvate kinase and phosphoenol pyruvate for NTP regeneration followed by synthesis of sugar nucleotides in a cyclic synthesis system for oligosaccharides 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis the enzyme is used to establih a thermostable ATP regeneration system from polyphosphate substrate, this is then used for synthesis of D-Ala-D-Ala in a coupled system with the thermostable D-alanine-D-alanine ligase TmDdl from Thermotoga maritima, a useful biocatalyst for synthesizing D-amino acid dipeptides, method development and optimization, overview 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis the enzyme is useful for ATP production from polyphosphate 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis a biocatalytic cascade of polyphosphate kinase and sucrose synthase is developed for synthesis of nucleotide-activated derivatives of glucose 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis construction of an ATP regeneration system from AMP using PPK2, coupled with aminoacyl proline (Xaa-Pro) synthesis catalyzed by the adenylation domain of tyrocidine synthetase TycA-A. 0.87 mM of L-Trp-L-Pro is successfully synthesized from AMP after 72 h. Addition of inorganic diphosphatase increases the reaction rate by 14fold. When the coupling reaction is applied to whole-cell reactions in Escherichia coli, ATP is successfully regenerated from AMP, and 6.7 mM of L-Trp-L-Pro is produced after 24 h with the supplementation of 10 mM AMP. Also various other L-Xaa-L-Pro an be produced 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis efficient synthesis of gamma-glutamyl compounds by co-expression of gamma-glutamylmethylamide synthetase and polyphosphate kinase in engineered Escherichia coli 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis energy delivery is a critical aspect of cell-free protein synthesis. The single kinase-based regeneration system simplifies cell free protein synthesis from a three-kinase system to single-kinase system, and potentially cheapens the cost of reagent preparations by using polyP instead of phosphocreatine. Incorporation of the PPK2-based NTP regeneration system into synthetic biomembrane vesicles can lead to artificial cell and proto-cell systems more akin to their natural counterparts 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis enzymatic production of glutathione coupling with an ATP regeneration system based on polyphosphate kinase 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis potential application for ATP regeneration in cascade reaction 2.7.4.1 ATP-polyphosphate phosphotransferase synthesis PPK2 is used for ATP regeneration to produce glutathione by a two-enzyme cascade in vitro. 47.1 mM glutathione can be synthesized with a productivity of 13.5 mM/h. ATP is regenerated approximately 471 times in the system within 3.5 h 2.7.4.B7 polyphosphate kinase (NTP) synthesis the enzyme can used to replenish ATP in ATP-consuming biochemical reactions 2.7.4.26 isopentenyl phosphate kinase synthesis expression of genes encoding enzymes of the mevalonate pathway from mevalonate to isopentenyl diphosphate in an engineered Escherichia coli strain that produces red carotinoid pigment lycopene. The archaeal pathway can function in bacterial cells to convert mevalonate into isopentenyl diphosphate 2.7.4.33 AMP-polyphosphate phosphotransferase synthesis design of an ATP regeneration system with poylphosphate-AMP phosphotransferase and polyphosphate kinase. The ATP regeneration system can also be used as a GTP regeneration system 2.7.4.33 AMP-polyphosphate phosphotransferase synthesis PPT catalyzes a key reaction in the cell-free regeneration of ATP from AMP and polyphosphate. The PPT/AdK system provides an alternative to existing enzymatic ATP regeneration systems in which phosphoenolpyruvate and acetylphosphate serve as phosphoryl donors and has the advantage that AMP and polyP are stable, inexpensive substrates 2.7.4.33 AMP-polyphosphate phosphotransferase synthesis the phospho-conversion of AMP to ADP using polyphosphate as phosphate donor by polyphosphate-AMP phosphotransferase, PAP, followed by the synthesis of ATP from ADP through poylphosphate kinase, PPK, is used as an ATP regenrating system. The PAP-PPK ATP regeneration system can continously produce ATP in the coupled reaction from polyphosphate, whhich is a cheaper substrate than acetylphosphate, phosphoenolphosphate, or creatine phosphate. The system can also regenerate GTP from GMP. Method optimization, overview. ATP can be used for e.g. synthesis of acetly-CoA 2.7.6.1 ribose-phosphate diphosphokinase synthesis the enzyme is useful in industrial production of the vitamin riboflavin using the fungus Ashbya gossypii 2.7.6.1 ribose-phosphate diphosphokinase synthesis expression of Escherichia coli Prs protein in Rosetta (DE3) and BL21 (DE3) pLysE strains and detailed method for His-Prs and untagged Prs purification on nickel affinity chromatography columns. The protocol allows purification of proteins with high yield, purity and activity 2.7.7.2 FAD synthase synthesis the enzyme is used for the preparation of flavin mononucleotide (FMN) and FMN analogues from their corresponding riboflavin precursors, which is performed in a two-step procedure. After initial enzymatic conversion of riboflavin to FAD by the bifunctional FAD synthetase, the adenyl moiety of FAD is hydrolyzed with snake venom phosphodiesterase to yield FMN. The engineered FAD synthetase from Corynebacterium ammoniagenes with deleted N-terminal adenylation domain is a biocatalyst that is stable and efficient for direct and quantitative phosphorylation of riboflavin and riboflavin analogues to their corresponding FMN cofactors at preparative-scale 2.7.7.2 FAD synthase synthesis modular engineering of Escherichia coli to improve flavin production and the conversion ratio of riboflavin RF to FMN/FAD. The RF operon and the bifunctional RF kinase/FAD synthetase are divided into two separate modules and expressed at different levels to produce different RF:ribF. The best strain respectively produces 324.1 and 171.6 mg/l of FAD and FMN in shake flask fermentation. Error-prone of the RibF gene further increase production up to 586.1 mg/l in shake flask cultivation 2.7.7.2 FAD synthase synthesis purified recombinant FADSCf can be used for the biosynthesis of FAD. Under optimized conditions (0.5 mM FMN, 5 mM ATP and 10 mM Mg2+), the production of FAD reaches 80 mM per mg of enzyme after a 21-hour reaction 2.7.7.6 DNA-directed RNA polymerase synthesis the enzyme is useful for in vitro transcription reactions to produce preparative quantities of transcribed RNA and labeled RNA probes, method evaluation, overview 2.7.7.6 DNA-directed RNA polymerase synthesis the enzyme is useful for in vitro transcription reactions to produce preparative quantities of transcribed RNA and labeled RNA probes, transcripts thousands of nucleotides in length, as are the major applications of these reactions, method evaluation, overview. Phage T7 RNA polymerase is an extremely processive enzyme in high-yield transcription of DNA sequences inserted downstream from the corresponding T7 promoter, applications overview 2.7.7.6 DNA-directed RNA polymerase synthesis the enzyme is useful for in vitro transcription reactions to produce preparative quantities of transcribed RNAand labeled RNA probes, transcripts thousands of nucleotides in length, as are the major applications of these reactions, method evaluation, overview. Phage SP6 RNA polymerase is an extremely processive enzyme in high-yield transcription of DNA sequences inserted downstream from the corresponding SP6 promoter, applications overview 2.7.7.6 DNA-directed RNA polymerase synthesis the enzyme is useful for in vitro transcription reactions to produce preparative quantities of transcribed RNAand labeled RNA probes, transcripts thousands of nucleotides in length, as are the major applications of these reactions, method evaluation, overview. Phage T3 RNA polymerase is an extremely processive enzyme in high-yield transcription of DNA sequences inserted downstream from the corresponding T3 promoter, applications overview 2.7.7.6 DNA-directed RNA polymerase synthesis development of an in vitro transcription system that accurately initiates transcription from bona fide promoters of the Peptoclostridium difficile glutamate dehydrogenase gene and rRNA genes rrnC and rrnE operons 2.7.7.7 DNA-directed DNA polymerase synthesis the enzyme is useful in DNA amplification and PCR-based applications, particularly in clinical diagnoses using uracil-DNA glycosylase. A mixture of Nanoarchaeum equitans DNA polymerase and Thermus aquaticus DNA polymerase improves the performance of Neq DNA polymerase for long and accurate PCR 2.7.7.7 DNA-directed DNA polymerase synthesis the very good competition of 7-substituted 7-deazapurine dNTPs, and still reasonably good activity of 5-substituted pyrimidine dNTPs, in the presence of their natural counterparts is very encouraging for further development of methods of polymerase synthesis of modified DNA and for possible in cellulo and even in vivo applications if satisfactory delivery of modified dNTPs will be solved 2.7.7.8 polyribonucleotide nucleotidyltransferase synthesis at the optimal temperature, polynucleotide phosphorylase completely destroys RNAs that possess even a very stable intramolecular secondary structure, but leaves intact RNAs whose 3' end is protected by chemical modification or by hybridization with a complementary oligonucleotide. This allows individual RNAs to be isolated from heterogeneous populations by degrading unprotected species. If oligonucleotide is hybridized to an internal RNA segment, the Tth polynucelotide phosphorylase stalls eight nucleotides downstream of that segment. This allows any arbitrary 5'-terminal fragment of RNA to be prepared with a precision similar to that of run-off transcription, but without the need for a restriction site 2.7.7.9 UTP-glucose-1-phosphate uridylyltransferase synthesis easy available enzyme can be used for synthesis of nucleotide sugars in enzymic glycoconjugate synthesis 2.7.7.9 UTP-glucose-1-phosphate uridylyltransferase synthesis immobilization enzyme provides a highly efficient biocatalyst for the production of UDP-glucose 2.7.7.13 mannose-1-phosphate guanylyltransferase synthesis enzyme can be used for the synthesis of GDPmannose deoxy derivatives 2.7.7.13 mannose-1-phosphate guanylyltransferase synthesis an N-acetylhexosamine 1-kinase from Bifidobacterium infantis (NahK_15697), a guanosine 5'-diphosphate (GDP)-mannose pyrophosphorylase from Pyrococcus furiosus (PFManC), and an Escherichia coli inorganic pyrophosphatase (EcPpA) are used efficiently for a one-pot three enzyme synthesis of GDP-mannose, GDP-glucose, their derivatives, and GDP-talose from simple monosaccharides and derivatives in preparative scale 2.7.7.18 nicotinate-nucleotide adenylyltransferase synthesis overexpression of enzyme gene under anaerobic conditions in a strain lacking IdhA and pflB gene products leads to 3.21fold increase in NAD+ and 1.67fold increase in NADH in the recombinant strain. Enzyme expression restores cell growth and glucose utilization under anaerobic conditions. After 72 h, the recombinant strain can consume 14 g/l of lgucose and produce 6.23 g/l of succinic acid 2.7.7.23 UDP-N-acetylglucosamine diphosphorylase synthesis synthesis of UDP-N-acetyl-D-glucosamine 2.7.7.23 UDP-N-acetylglucosamine diphosphorylase synthesis biosynthesis of UDP-N-acetyl-alpha-D-glucosamine. As glycosylation is the most important modification for activating peptide drugs, the activated form of N-acetyl-alpha-D-glucosamine is thought to be important for future development of effective drugs 2.7.7.27 glucose-1-phosphate adenylyltransferase synthesis adding Azospirillum brasilense enhances AGPase activity, starch formation, and mitigation of stress in Chlorella vulgaris. AGPase activity during the first 72 h of incubation is higher in Chlorella vulgaris when immobilized with Azospirillum brasilense, concomitant with higher starch accumulation and higher carbon uptake by the microalgae. Either carbon source and starvation either by N or P has the same pattern of AGPase activity and starch accumulation. Under replete conditions, the population of Chlorella vulgaris immobilized alone is higher than when immobilized together, but under starvation conditions Azospirillum brasilense induces a larger population of Chlorella vulgaris 2.7.7.30 fucose-1-phosphate guanylyltransferase synthesis enzymatic synthesis of GDP-fucose 2.7.7.30 fucose-1-phosphate guanylyltransferase synthesis a chemoenzymatic model for the preparative-scale synthesis of a diverse array of GDP-fucose derivatives is reported 2.7.7.30 fucose-1-phosphate guanylyltransferase synthesis functional expression of enzyme gene in Saccharomyces cerevisiae. The yeast cells synthesize GDP-L-fucose or GDP-D-arabinose upon exogenous addition of L-fucose or D-arabinose, respectively 2.7.7.30 fucose-1-phosphate guanylyltransferase synthesis engineering of Escherichia coli for efficient production of 2'-fucosyllactose by introduction of the fkp gene coding for fucokinase/GDP-L-fucose diphosphorylase from Bacteroides fragilis and the fucT2 gene encoding alpha-1,2-fucosyltransferase from Helicobacter pylori to produce 2'-fucosyllactose from fucose, lactose and glycerol. The yield and productivity are further improved by deletion of the fucI-fucK gene cluster coding for fucose isomerase and fuculose kinase. Fed-batch fermentation of the final strain results in 23.1 g/l of extracellular 2'-fucosyllactose and a productivity of 0.39 g/l/h 2.7.7.43 N-acylneuraminate cytidylyltransferase synthesis since CMP-N-acylneuraminate is unstable and relatively expensive, the enzyme is valuable for the preparative enzymatic synthesis of silylated oligosaccharides 2.7.7.43 N-acylneuraminate cytidylyltransferase synthesis the high expressivity of the recombinant production clone, the high catalytic efficiency of the enzyme, and its broad substrate tolerance make this enzyme the preferred catalyst for the enzymatic synthesis of CMP-Neu5Ac 2.7.7.43 N-acylneuraminate cytidylyltransferase synthesis the enzyme is used for preparative synthesis of CMP-N-acylneuraminate. Optimization of the culture conditions for the production of the enzyme and the detection of a colony variant of Escherichia coli K-235 that is an even better producer of the synthetase 2.7.7.43 N-acylneuraminate cytidylyltransferase synthesis the enzyme is used for preparative synthesis of CMP-sialic acid derivatives in a one-pot two-enzyme system with EC 4.1.3.3 and EC 2.7.7.43 2.7.7.48 RNA-directed RNA polymerase synthesis RdRp from the bacteriophage phi6 can be used to generate dsRNA for RNA interference, RNAi, applications, resulting in in vivo gene silencing. RNA molecules that are radioactively or fluorescently labeled using poly(A) polymerase can be used as probes in a wide variety of applications 2.7.7.48 RNA-directed RNA polymerase synthesis the functional nature of multimeric arrays of RNA-dependent RNA polymerase provides an appreciation for enzymatic catalysis on membranous surfaces within cells 2.7.7.48 RNA-directed RNA polymerase synthesis the RNA-dependent RNA polymerase of the bacteriophage phi6 is used for synthesis of long double-stranded RNA from a cloned region of the genome of Coxsackie virus B3, i.e. CBV3, a member of the human enterovirus B species, for usage in RNA interference in antiviral experimental therapies and gene function studies, overview. Synthesis of GFP CBV3 siRNA 2.7.7.49 RNA-directed DNA polymerase synthesis reverse transcriptase is commonly used to synthesize DNA complementary to a variety of RNA templates, synthesis of cDNA. Reverse transcriptase can utilize single-stranded DNA or RNA-DNA hybrid as template to synthesize double-stranded DNA. The reverse transcriptase, unlike the bacterial DNA polymerase, lacks the 3'-5' and 5'-3' exonuclease and can thus be efficiently used for end labeling or gap filling 2.7.7.64 UTP-monosaccharide-1-phosphate uridylyltransferase synthesis the enzyme is useful for the highly efficient production of UDP-galacturonic acid for studies on pectin biosynthesis 2.7.7.64 UTP-monosaccharide-1-phosphate uridylyltransferase synthesis use of UDP-sugar pyrophosphorylase (USP) from Arabidopsis thaliana with a galactokinase from Streptococcus pneumoniae TIGR4 (SpGalK) and an inorganic diphosphatase (PPase) to effectively synthesize UDP-sugars 2.7.7.64 UTP-monosaccharide-1-phosphate uridylyltransferase synthesis in mammals, uridine 5'-diphosphate-glucose (UDP-Glc) and uridine 5'-diphosphate-glucuronic acid (UDP-GlcA) are common building blocks of glycans and glycoconjugates. The commercial demand for these high-energy donors is increasing. To produce valuable UDP-GlcA in a cost-effective way, UDP-sugar pyrophosphorylase from Arabidopsis thaliana is constitutively expressed in Pichia pastoris and secreted into the extracellular medium. The synthesis of 4.2 g UDP-GlcA or 5.5 g UDP-Glc per liter of culture is revealed in the culture medium, without any need for purification. An anion exchange chromatography purification method for UDP-sugars is also developed. This route opens a door to large-scale production of the cheaper UDP-GlcA 2.7.7.65 diguanylate cyclase synthesis the thermophilic enzyme is a valuable tool for c-di-GMP synthesis as well as the preparation of c-di-GMP derivatives 2.7.7.105 phosphoenolpyruvate guanylyltransferase synthesis the 3PG-F420 biosynthetic gene cluster is fully functional in Escherichia coli, enabling convenient production of the cofactor by fermentation 2.7.7.106 3-phospho-D-glycerate guanylyltransferase synthesis the 3PG-F420 biosynthetic gene cluster is fully functional in Escherichia coli, enabling convenient production of the cofactor by fermentation 2.7.8.2 diacylglycerol cholinephosphotransferase synthesis heterologous expression in Camelina sativa of lychee phosphatidylcholine diacylglycerol cholinephosphotransferase PDCT. Camelina lines coexpressing PDCT and Escherichia coli cyclopropane synthase CPS show up to a 50% increase of cyclopropane fatty acids in mature seed, relative to the CPS background. The expression of PDCT strongly reduces the level of C18:1 substrate at phosphatidylcholine sn-1 and phosphatidylcholine sn-2 (i.e. the sites of cyclopropane fatty acids synthesis), while the levels of cyclopropane fatty acids increased in phosphatidylcholine sn-2, diacylglycerol sn-1 and diacylglycerol sn-2, and both sn-1/3 and sn-2 positions in triacylglycerol 2.7.8.7 holo-[acyl-carrier-protein] synthase synthesis the broad specificity of the single PPTase present in Pseudomonas can be used to 4'-phosphopantetheinylate various carrier proteins 2.7.8.7 holo-[acyl-carrier-protein] synthase synthesis biotechnological de novo production of m-cresol from sugar in complex yeast extract-peptone medium with the yeast Saccharomyces cerevisiae. A heterologous pathway based on the decarboxylation of the polyketide 6-methylsalicylic acid is introduced into a CEN.PK yeast strain. Overexpression of codon-optimized 6-methylsalicylic acid synthase from Penicillium patulum together with activating phosphopantetheinyl transferase npgA from Aspergillus nidulans results in up to 367 mg/l 6-methylsalicylic acid production. Additional genomic integration of the genes have a strongly promoting effect and 6-methylsalicylic acid titers reach more than 2 g/l. Simultaneous expression of 6-methylsalicylic acid decarboxylase patG from Aspergillus clavatus leads to the complete conversion of 6-methylsalicylic acid and production of up to 589 mg/L m-cresol 2.7.8.13 phospho-N-acetylmuramoyl-pentapeptide-transferase synthesis the enzyme is an attractive antibacterial drug target involved in peptidoglycan synthesis 2.7.8.13 phospho-N-acetylmuramoyl-pentapeptide-transferase synthesis heterologous production of enzyme in Escherichia coli membranes into styrene-maleic acid-wrapped nanodiscs and purification using detergent-free styrene-maleic acid copolymer system. Yield and the purity of the recombinant protein are comparable to enzyme extracted with a conventional detergent. The predominantly alpha-helical secondary structure of the protein in SMA-wrapped nanodiscs is more stable against heat denaturation compared to the micellar protein 2.7.8.31 undecaprenyl-phosphate glucose phosphotransferase synthesis the enzyme is involved in biosynthesis of xanthan, an industrially important exopolysaccharide 2.7.8.43 lipid A phosphoethanolamine transferase synthesis EptA remains active despite encapsulation within a nanostructured bicontinuous cubic phase and displays full transfer of the phosphoethanolamine group from a 1,2-dioleoyl-glycero-phosphoethanolamine doped lipid to monoolein, which makes up the bicontinuous cubic phase 2.7.10.1 receptor protein-tyrosine kinase synthesis methodology for generating milligram amounts of functional Eph tyrosine kinase receptor using the protein engineering approach of expressed protein ligation. Stimulation with ligand induces efficient autophosphorylation of the semisynthetic Eph construct. The in vitro phosphorylation of key Eph tyrosine residues upon ligand-induced activation follows a precise and unique order of phosphorylation of the Eph tyrosines in the kinase activation process 2.7.10.2 non-specific protein-tyrosine kinase synthesis a silent mutation at residue Ile229 changing the rare codon ATA in Escherichia coli to codon ATT or ATC allows for the fast expression and the purification of the unphosphorylated and phosphorylated kinase domains 2.7.10.2 non-specific protein-tyrosine kinase synthesis expression and a two-step purification procedure for the doubly tagged full-length expression construct, H6-FL-TYK-2-FLAG. In the presence of ATP and a peptide substrate, H6-FL-TYK-2-FLAG shows a marked lag in phosphopeptide product formation, while the wild-type shows no such lag. The lag can be eliminated by ATP pretreatment. The potencies of several nanomolar inhibitors are similar for TYK-2 KD and H6-FL-TYK-2-FLAG. However, these same inhibitors are about 1000 times less potent inhibiting the autophosphorylation of H6-FL-TYK-2-FLAG than they are inhibiting the phosphorylation of a peptide substrate modeled after the activation loop sequence of wild-type TYK-2 2.7.13.3 histidine kinase synthesis the deletion of histidine kinases cbei2073 and cbei4484 results in significant change in butanol biosynthesis, with butanol production increased by 40.8 and 17.3% (13.8 g/L and 11.5 g/L vs. 9.8 g/L), respectively, compared to the wild-type. Faster butanol production rates are observed, with butanol productivity greatly increased by 40.0 and 20.0%, respectively. The sporulation frequencies of two histidine kinases inactivated strains decrease by 96.9 and 77.4%, respectively 2.8.1.10 thiazole synthase synthesis synthesis of the thiazole moiety of thiamin from glycine, cysteine, and deoxy-D-xylulose-5-phosphate using overexpressed Bacillus subtilis ThiF, ThiS, ThiO, ThiG, and a NifS-like protein 2.8.2.1 aryl sulfotransferase synthesis Escherichia coli cells expressing human isoform SULT1A3 produce preferentially genistein 4'-sulfate, Escherichia coli cells expressing isoform SULT1C4 produce preferentially genistein 7-sulfate. Presence of glucose, SO42-, and incubation temperature are key factors that affect the efficiency of the production 2.8.2.1 aryl sulfotransferase synthesis generation of a complete set of recombinant fission yeast strains each expressing one of the 14 human SULT enzymes. The intracellular production of the cofactor 3'-phosphoadenosine 5'-phosphosulfate (PAPS) necessary for SULT activity is sufficiently high to support metabolite production 2.8.2.1 aryl sulfotransferase synthesis the enzyme is used for large scale chondroitin sulfate synthesis in a coupled system with chondroitin 4-sulfotransferase and chondroitin 6-sulfotransferase, overview 2.8.2.2 alcohol sulfotransferase synthesis cytosolic sulfotransferases (SULTs) acting as phase II metabolic enzymes can be used in the sulfonation of small molecules by transferring a sulfonate group from the unique co-factor 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to the substrates 2.8.2.2 alcohol sulfotransferase synthesis generation of a complete set of recombinant fission yeast strains each expressing one of the 14 human SULT enzymes. The intracellular production of the cofactor 3'-phosphoadenosine 5'-phosphosulfate (PAPS) necessary for SULT activity is sufficiently high to support metabolite production 2.8.2.4 estrone sulfotransferase synthesis generation of a complete set of recombinant fission yeast strains each expressing one of the 14 human SULT enzymes. The intracellular production of the cofactor 3'-phosphoadenosine 5'-phosphosulfate (PAPS) necessary for SULT activity is sufficiently high to support metabolite production 2.8.2.5 chondroitin 4-sulfotransferase synthesis the enzyme is used for large scale chondroitin sulfate synthesis in a coupled system with aryl sulfotransferase IV and chondroitin 6-sulfotransferase, overview 2.8.2.17 chondroitin 6-sulfotransferase synthesis the enzyme is used for large scale chondroitin sulfate synthesis in a coupled system with aryl sulfotransferase IV and chondroitin 4-sulfotransferase, overview 2.8.2.20 protein-tyrosine sulfotransferase synthesis usage of tyrosylprotein sulfotransferases for in vitro one-pot enzymatic synthesis of sulfated proteins/peptides 2.8.2.23 [heparan sulfate]-glucosamine 3-sulfotransferase 1 synthesis use of a chemoenzymatic synthetic approach to synthesize six 3-O-sulfated oligosaccharides, including three hexasaccharides and three octasaccharides. The synthesis is achieved by rearranging the enzymatic modification sequence to accommodate the substrate specificity of 3-O-sulfotransferase 3, analysis of the impact of 3-O-sulfation on the conformation of the pyranose ring of 2-O-sulfated iduronic acid using NMR spectroscopy, and on the correlation between ring conformation and anticoagulant activity. An octasaccharide interacts with antithrombin and displays anti factor Xa activity. The octasaccharide displays a faster clearance rate than fondaparinux, an FDA-approved pentasaccharide drug, in a rat model, making this octasaccharide a potential short-acting anticoagulant drug candidate that could reduce bleeding risk 2.8.2.30 [heparan sulfate]-glucosamine 3-sulfotransferase 3 synthesis use of a chemoenzymatic synthetic approach to synthesize six 3-O-sulfated oligosaccharides, including three hexasaccharides and three octasaccharides. The synthesis is achieved by rearranging the enzymatic modification sequence to accommodate the substrate specificity of 3-O-sulfotransferase 3, analysis of the impact of 3-O-sulfation on the conformation of the pyranose ring of 2-O-sulfated iduronic acid using NMR spectroscopy, and on the correlation between ring conformation and anticoagulant activity. An octasaccharide interacts with antithrombin and displays anti factor Xa activity. The octasaccharide displays a faster clearance rate than fondaparinux, an FDA-approved pentasaccharide drug, in a rat model, making this octasaccharide a potential short-acting anticoagulant drug candidate that could reduce bleeding risk 2.8.2.33 N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase synthesis use of enzyme for synthesis of chondroitin sulfate E from chondroitin sulfate A and of oversulfated dermatan sulfate with defined proportions of 4,6-sulfated residues 2.8.2.38 aliphatic desulfoglucosinolate sulfotransferase synthesis 3'-phosphoadenylyl-sulfate:aliphatic desulfoglucosinolate sulfotransferase from Arabidopsis thaliana ecotype Col-0 and myrosinase from Brevicoryne brassicae are expressed in Escherichia coli and successfully used for the biosynthesis of benzyl isothiocyanate by the combined expression of the optimized enzymes in vitro 2.8.3.1 propionate CoA-transferase synthesis engineered mutant Pct, coexpressed with an engineered Pseudomonas sp. MBEL 6-19 polyhydroxyalkanoate synthase 1, is able to produce poly(3-hydroxybutyrate-co-lactate) copolymers having 9-64 mol% of lactate. Polylactic acid might be a good alternative to petroleum-based plastic as it possesses several desirable properties such as biodegradability, biocompatibility, compostability, and low toxicity to humans 2.8.3.1 propionate CoA-transferase synthesis in an engineered Escherichia coli strain JLX7, the engineered enzyme mutant Pct540, coexpressed with an engineered Pseudomonas sp. MBEL 6-19 polyhydroxyalkanoate synthase 1, is able to effectively produce polylactic acid, a promising biomass-derived polymer, overview 2.8.3.1 propionate CoA-transferase synthesis a platform pathway for the production of widely used industrial chemicals 1,3-diols, e.g. 1,3-pentanediol, is engineered in Escherichia coli. The pathway is designed by modifying the previously reported (R)-1,3-butanediol synthetic pathway to consist of pct (propionate CoA-transferase) from Megasphaera elsdenii, bktB (thiolase), phaB (NADPH-dependent acetoacetyl-CoA reductase) from Ralstonia eutropha, bld (butyraldehyde dehydrogenase) from Clostridium saccharoperbutylacetonicum, and the endogenous alcohol dehydrogenase(s) of Escherichia coli. The recombinant Escherichia coli strains produce 1,3-pentanediol, 4-methyl-1,3-pentanediol, and 1,2,4-butanetriol, together with 1,3-butanediol, from mixtures of glucose and propionate, isobutyrate, and glycolate, respectively, in shake flask cultures 2.8.3.21 L-carnitine CoA-transferase synthesis production of L-carnithine in Escherichia coli. The overexpression of CoA-transferase CaiB and CoA-ligase CaiC leads to an increase in L-carnitine production. Under optimal concentrations of inducer and fumarate yields reach 10- and 50fold, respectively, that obtained for the wild type strain. Low levels of coenzyme A limit the activity of these two enzymes. The glyoxylate shunt and anaplerotic pathways limit the bioprocess since strains carrying deletions of isocitrate lyase and isocitrate dehydrogenase phosphatase/kinase yielded 20-25% more L-carnitine than the control. The deletion of phosphotransacetylase strongly inhibits the bioprocess 2.8.3.21 L-carnitine CoA-transferase synthesis production of L-carnithine in Escherichia coli. When the dilution rate and the initial crotonobetaine concentration are directly changed in the real experimental system to the optimum values, the system a 74% increase in production rate. Development of a model showing control points at reactor operation and molecular levels where conversion and productivity can be increased 2.8.4.1 coenzyme-B sulfoethylthiotransferase synthesis expression of methyl-coenzyme M reductase from an unculturable organism in Methanosarcina acetivorans to effectively run methanogenesis in reverse. Methanosarcina acetivorans cells heterologously producing methyl-coenzyme M reductase consume up to 9% of methane (corresponding to 109 ± 12 micromol of methane) after 6 weeks of anaerobic growth on methane and utilize 10 mM FeCl3 as an electron acceptor. When incubated on methane for 5 days, high-densities of cells consume 15% methane (corresponding to 143 ± 16 micromol of methane), and produce 10.3 mM acetate (corresponding to 52 ± 4 µmol of acetate) 3.1.1.1 carboxylesterase synthesis biocatalyst for esterification reactions 3.1.1.1 carboxylesterase synthesis enzyme is useful for deprotection of the compounds in the production of cetraxate and D-panothenate 3.1.1.1 carboxylesterase synthesis methyl hydrogen (1S,2R)-cyclohex-4-ene-1,2-dicarboxylate is an important educt for the synthesis of diverse biologically relevant products such as carbacyclin, prostaglandin synthon, brefeldin, anticapsin and others 3.1.1.1 carboxylesterase synthesis usage of the enzyme as biocatalyst for biotransformations in non-aqueous media in industrial applications at high and also at moderate temperatures 3.1.1.1 carboxylesterase synthesis effective removal of t-butyl protecting group in organic synthesis 3.1.1.1 carboxylesterase synthesis the enzyme is useful for the synthesis of 1-monocaprin, a widely used functional emulsifier in the food, cosmetic, and pharmaceutical industries and is recognized as a GRAS food additive 3.1.1.1 carboxylesterase synthesis synthesis of poly(delta-valerolactone) in organic solvents. The synthesized poly(delta-valerolactone) is of low molecular weight and narrow molecular weight distribution, and is expected to be widely used as the soft block of thermoplastic elastomers, or carriers for controlled drug delivery and release 3.1.1.1 carboxylesterase synthesis synthesis of poly(epsilon-caprolactone) of lower molecular weight, which can be used as the drug carrier or soft segment of polyurethanes 3.1.1.1 carboxylesterase synthesis the enzyme physically immobilized on hydrophobic macroporous resin is a biocatalyst for polyester synthesis 3.1.1.1 carboxylesterase synthesis the enzyme shows chiral resolution activity for (S)-ibuprofen, indicating that the enzyme can be used for the production of commercially important chiral drugs 3.1.1.1 carboxylesterase synthesis synthesis of (S)-alpha-arylpropionates (profens). The reaction proceeds via enantioselective hydrolysis of the malonic acid ester to the monoester. Cleavage of the carboxyl group at the active site of the enzyme leads to the formation of the prochiral intermediate, which is then protonated to give the enantioenriched monoacid. Engineering of the active site gives rise to a variant with good stereoselectivity. As the esterase-catalyzed domino reaction does not need external cofactors and allows a theoretical yield of 100% 3.1.1.1 carboxylesterase synthesis synthesis of chiral compounds 3.1.1.1 carboxylesterase synthesis the enzyme can be used for application in the synthesis of chiral compounds 3.1.1.1 carboxylesterase synthesis enzyme shows high enantioselectivity in the kinetic resolution of 2-carboxyethyl-3-cyano-5-methylhexanoic acid ethyl ester, which produce a valuable chiral intermediate-(3S)-2-carboxyethyl-3-cyano-5-methylhexanoic acid for the drug Pregabalin 3.1.1.1 carboxylesterase synthesis application of encapsulated hydrolases for enzymatic syntheses 3.1.1.1 carboxylesterase synthesis utilization of a marine bacillus esterase in the stereo-selective production of D-methyl lactate through enzymatic kinetic resolution reactions 3.1.1.3 triacylglycerol lipase synthesis immobilized enzyme catalyzes the esterification of oleic acid with butanol dissolved in hexane 3.1.1.3 triacylglycerol lipase synthesis enantioselective hydrolysis of chiral esters 3.1.1.3 triacylglycerol lipase synthesis immobilized enzyme is used for the transesterification reaction that replaces pamitic acid in palm oil with stearic acid 3.1.1.3 triacylglycerol lipase synthesis possible industrial application of the enzyme in continuous processes due to effective immobilization on diverse materials, overview 3.1.1.3 triacylglycerol lipase synthesis synthesis of enantiopure compounds by chemo-, regio-, and stereoselective transformations, catalysation of hydrolysis of water-immiscible triglycerides at water-liquid interface, transesterifications 3.1.1.3 triacylglycerol lipase synthesis the enzyme performs chemically selective hydrolysis, synthesis of optically active substances from achiral or racemic compounds 3.1.1.3 triacylglycerol lipase synthesis useful in application to the processing of industrial fats and oils containing eicosapentanoic acid and docosahexaenoic acid, such as fish oil splitting 3.1.1.3 triacylglycerol lipase synthesis production of (-)-4-endo-hydroxy-2-oxabicyclo[3.3.0]oct-7-en-3-one, which is an intermediate for the anti-HIV agent carbovir 3.1.1.3 triacylglycerol lipase synthesis production of (2R,3S)-3-(4-methoxyphenyl)glycidic acid methyl ester, which is an intermediate in the synthesis of diltiazem. diltiazem hydrochloride is a coronary vasodilator and a calcium channel blocker 3.1.1.3 triacylglycerol lipase synthesis production of (3R,4S)-cis-azetidinone acetate, which is an intermediate for the synthesis of paclitaxel 3.1.1.3 triacylglycerol lipase synthesis production of (S)-1-phenylethylamine and (R)-phenylethylmethoxyamide, which are intermediates for pharmaceuticals and pesticides and can also be used as chiral synthons in asymmetric synthesis 3.1.1.3 triacylglycerol lipase synthesis production of (S)-acetic acid 2-methyl-4-oxo-3-prop-2-ynyl-cyclopent-2-enyl ester, which is used as an intermediate in the synthesis of pyrethroids, which are used as insecticides. They show excellent insecticidal activities and a low toxicity in mammals. 3.1.1.3 triacylglycerol lipase synthesis production of (S)-acetic acid 4-(2,4-difluoro-phenyl)-2-hydroxymethyl-pent-4-enyl ester, which is used as an improved azole antifungal. It shows activity against systemic Candida and pulmonary Aspergillus infections 3.1.1.3 triacylglycerol lipase synthesis production of (S)-ester amide, which is useful as a lipophilic, hindered component of peptides. The amino acids are also useful building blocks for a number of chiral auxiliaries and ligands 3.1.1.3 triacylglycerol lipase synthesis production of Ibuprofen, which is an important nonsteroidal antiinflammatory drug 3.1.1.3 triacylglycerol lipase synthesis production of isopropyl palmitate, which is used in the preparation of soaps, skin creams, lubricants and grease 3.1.1.3 triacylglycerol lipase synthesis production of [4-[4a,6b(E)]]-6-[4,4-bis(4-fluorophenyl)-3-(1-methyl-1H-tetrazol-5-yl)-1,3-butadienyl]-tetrahydro-4-hydroxy-2H-pyran-2-one acetate, which is a hydroxymethyl glutaryl coenzyme A (HMG-CoA) reductase inhibitor and a potential anticholesterol drug candidate 3.1.1.3 triacylglycerol lipase synthesis construction of an overexpressing Aspergillus oryzae strain immobilized on supporting particles for enantioselective transesterification reaction 3.1.1.3 triacylglycerol lipase synthesis the enzyme can be used for hydrolysis and synthesis of various esters, mutagenic modification and optimization of Candida rugosa isozymes for enantioselective, substrate-specific biocatalysis, improvement of thermostability, enantioselectivity, and substrate specificity, possible reactions are hydrolysis, direct esterification, acidolysis, alcoholysis, ester-interchange, and glycerolysis, overview 3.1.1.3 triacylglycerol lipase synthesis application of the immobilized lipase in non-conventional biocatalysis for the synthesis of surfactants and biodiesel, overview 3.1.1.3 triacylglycerol lipase synthesis the enantioselective lipase shows potential to be used as a catalyst to prepare optically pure pharmaceuticals 3.1.1.3 triacylglycerol lipase synthesis the enzyme shows potential in the production of optically pure ibuprofen 3.1.1.3 triacylglycerol lipase synthesis lipases constitute one of the most important groups of biocatalysts due to their ability to catalyze three different kinds of reactions viz. hydrolysis, esterification, and transesterification with unique chemo-, regio-, and enantiospecific selectivity. Lipase-based processes are employed for oil/fat processing, synthesis of industrially important oleochemicals, enantiopure pharmaceuticals, agrochemicals, flavor esters, structured lipids, and biodiesel production 3.1.1.3 triacylglycerol lipase synthesis lipases maintain activity and selectivity in organic solvents, which has enabled their wide use as biocatalysts finding applications in food, dairy, detergent, and pharmaceutical industries 3.1.1.3 triacylglycerol lipase synthesis synthesis of arylaliphatic glycolipids, ethyl esterification of docosahexaenoic acid to ethyl docosahexaenoate, synthesis of citronellol laurate from citronellol and lauric acid, optically active ester synthesis, ester synthesis, desymmetrization and production of peracids, organic synthesis of chiral intermediates, synthesis of butyl caprylate in n-heptane, synthesis of butyl lactate by transesterification, synthesis of amide 3.1.1.3 triacylglycerol lipase synthesis the highly enantioselective lipase from Serratia marcescens ECU1010 is a robust biocatalyst for practical use in large-scale production of diltiazem intermediate 3.1.1.3 triacylglycerol lipase synthesis the lipase from Rhizomucor miehei may be used as a means to prepare fatty acid concentrates rich in docosahexaenoic acid 3.1.1.3 triacylglycerol lipase synthesis conversion of plant oil to 92-97% of biodiesel is feasible at 1% enzyme load (24 h, 35°C) using the feedstocks containing 2-20% of water, 0-10% of glycerol, 0-20% of free fatty acids. The enzyme can be collected in a narrow white layer settled between biodiesel and glycerol-water phases,containing also free fatty acids and monoglycerides. The lipase can be then reused after compensation for 5-10% loss of the enzyme. The main contaminants in the produced biodiesel are free fatty acids (2-6%) and monoglycerides (1-3%). Major amounts of free fatty acids and monoglycerides are removed after brief mixing of biodiesel with alkali (2-5% of 5 M NaOH) and centrifugation 3.1.1.3 triacylglycerol lipase synthesis ethanolysis of soybean oil in a solvent-free system for synthesis of biodiesel. The optimal conditions are: 31.5°C, 7 h reaction time, substrate molar ratio 7.5:1 ethanol:soybean oil, enzyme content 15% (g enzyme/g oil), 4% added water/g oil. The experimental yield conversion is 96% 3.1.1.3 triacylglycerol lipase synthesis immobilization and stabilization of lipase on aldehyde-Lewatit. Over 90% of lipase activity is recovered after the immobilization, the immobilized enzyme is 10fold more thermostable than the commercial preparation, Lipozyme TL-IM. The stabilized preparation catalyzes enzymatic transesterification of ethanol and soybean oil. With 7.5:1 molar ratio of ethanol:soybean oil, 15% immobilized enzyme and 4% water at 30°C in the presence of n-hexane, the transesterification reaches 100% conversion, while in solvent-free system the yield is 75%. At stoichiometric molar ratio, the yield is 70% conversion after 10 h of reaction in both systems. A two step ethanolysis produces 100% conversion after 10 h of reaction in both solvent and solvent-free systems 3.1.1.3 triacylglycerol lipase synthesis solubilization in sodium bis-(2-ethylhexyl)sulfosuccinate-stabilized water-in-oil microemulsions in n-heptane and analysis of hydrolysis and condensation activity. Condensation activity is essentially independent of temperature over the range 5 to 37°C. The stability over a 30-day period is very good at all pH levels (6.1, 7.2, 9.3) and R values studied (5, 7.5, 10, 20), except when high pHs and low R values are combined 3.1.1.3 triacylglycerol lipase synthesis solubilization in sodium bis-(2-ethylhexyl)sulfosuccinate-stabilized water-in-oil microemulsions in n-heptane and analysis of hydrolysis and condensation activity. Esterification activity shows only a slight dependence on temperature over the studied range and an apparent activation energy of 20 kJ/mol for octyl decanoate synthesis. The enzyme shows good stability over a 30-day period in R = 7.5 and R = 10 microemulsions, pH 6.1 3.1.1.3 triacylglycerol lipase synthesis synthesis of biodiesel by Rhizomucor miehei lipase immobilized on macroporous anion exchange resins using an ethanolysis process of sunflower oil. Quantitative conversions of triglycerides to fatty acid ethyl esters is obtained under mild reaction conditions that correspond to the transformation of triglycerides in a mixture of two moles of fatty acid ethyl esters and a mole of monoglyceride, thus avoiding the glycerol production. Reaction can be carried out under standard conditions with oil/ethanol volume ratio 12/3.5 ml at 40°C and 40 mg of immobilized enzyme 3.1.1.3 triacylglycerol lipase synthesis synthesis of biodiesel from canola oil. Adding tert-butanol to the reaction medium increases the conversion of oil to fatty acid methyl ester for the enzyme immobilized on epoxy-functionalized silica, 50 wt.% tert-butanol by substrate weight gives the best yield. Presence of water significantly increases fatty acid methyl ester yield. 85% residual activity after 16 reaction cycles 3.1.1.3 triacylglycerol lipase synthesis synthesis of biodiesel from canola oil. Complete conversion to fatty acid methyl esters is achieved using the enzyme immobilized on epoxy-functionalized silica, 30% (w/w) tert-butanol by substrate weight, reaction time of 96 h, 50°C and molar ratio of methanol to oil 3:1, which is added to the reaction mixture in three steps. Water suppresses the methanolysis reaction. 85% residual activity after 16 reaction cycles 3.1.1.3 triacylglycerol lipase synthesis synthesis of biodiesel from canola oil. Enzyme immobilized on epoxy-functionalized silica reaches to 100% yield at 10% tert-butanol. Presence of water significantly increases fatty acid methyl ester yield. 95% residual activity after 16 reaction cycles 3.1.1.3 triacylglycerol lipase synthesis synthesis of biodiesel from palm olein by ethanolysis using protein-coated microcrystals. Addition of tert-butanol markedly increases the biocatalyst activity and stability. Optimized reactions (20%, w/w protein-coated microcrystal-lipase to triacylglycerol and 1:4 fatty acid equivalence/ethanol molar ratio) lead to the production of alkyl esters from palm olein at 89.9% yield on molar basis after incubation at 45°C for 24 h in the presence of tert-butanol at a 1:1 molar ratio to triacylglycerol. Crude palm oil and palm fatty acid distillate are also converted to biodiesel with 82.1 and 75.5% yield, respectively 3.1.1.3 triacylglycerol lipase synthesis synthesis of eugenyl benzoate by esterification of eugenol and benzoic acid catalyzed by the chitosan-chitin nanowhiskers supported lipase. Under optimum conditions, a maximum conversion yield of 66% at 50°C in 5 h using 3 mg/ml of lipase, and a substrate molar ratio (eugenol: benzoic acid) of 3:1 is obtained. The lipase is reusable up to 8 esterification cycles and shows higher thermal stability than free enzyme 3.1.1.3 triacylglycerol lipase synthesis use of enzyme as biocatalyst to obtain a second generation biodiesel-like biofuel by the conversion of sunflower oil into a blend of fatty acid ethyl esters (FAEE), monoacylglycerols (MG) and diacylglycerols (DG). pH, molar ratio of ethanol to oil and water content influence the conversion in the systems. Low temperatures (20°C), high pH values (close to 12), and an oil/ethanol volume ratios of 3.4/1 provide conversions around 70% and kinematic viscosities about 8.5 mm2/s after 1 h reaction 3.1.1.3 triacylglycerol lipase synthesis use of enzyme immobilized on styrene-divinylbenzene beads for the synthesis of butyl butanoate using n-hexane as solvent. The enzyme presents high initial reaction rates up to 1.0 M butanoic acid, and allows a productivity of 14.5 mmol/g/h 3.1.1.4 phospholipase A2 synthesis protocol for the overexpression ofisoform PLA2III in Escherichia coli in the form of inclusion bodies, and for its purification and refolding using the Gateway system (Invitrogen) for the expression of a large quantity of the mature with an N-terminal His-tag, and Ni-affinity chromatography for purification. The purified recombinant PLA2III fusion protein is then refolded using a step-wise dialysis approach. About 40 mg purified and active protein is obtained from 1 l of cell culture 3.1.1.6 acetylesterase synthesis enzyme is useful for the commercial geraniol production from palmarosa oil 3.1.1.7 acetylcholinesterase synthesis production of human enzyme by expression in Nicotiana benthamiana with C-terminal SEKDEL endoplasmic retention signal, purified protein is kinetically undistinguishable from commercially available enzyme 3.1.1.7 acetylcholinesterase synthesis the enzyme immobilized on porous silicon offers a possibilities as a viable biocatalyst in bioprocessing for the chemical and pharmaceutical industries, and bioremediation to enhance productivity and robustness 3.1.1.14 chlorophyllase synthesis useful tool for preparing chlorophyllides and chlorophyll derivatives esterified with various alcohols 3.1.1.14 chlorophyllase synthesis recombinant enzyme CrCLH1 can be used as a biocatalyst to produce chlorophyllide derivatives 3.1.1.14 chlorophyllase synthesis the enzyme CyanoCLH can be used as a biocatalyst for the production of a bacteriochlorin a (BCA) precursor by degrading bacteriochlorophyll a (BChl a). Bacteriopheophorbide a (BPheid a) is used as a precursor for bacteriochlorin a (BCA), which can be used for photodynamic therapy in both in vitro and in vivo biochemical applications 3.1.1.17 gluconolactonase synthesis bioconversion of glucose and fructose to gluconic acid and sorbitol, respectively, by the enzymes glucose-fructose oxidoreductase and gluconolactonase, contained in untreated cells of Zymomonas mobilis 3.1.1.17 gluconolactonase synthesis enzyme is involved in the production of gluconic acid 3.1.1.20 tannase synthesis production of propyl gallate for the food industry and trimethoprim in the pharmaceutical industry 3.1.1.20 tannase synthesis gallate is used for synthesis of propyl gallate, a widely used food antioxidant 3.1.1.20 tannase synthesis the enzyme has wide applications in food, beverage, brewing, cosmetic, and chemical industry, overview 3.1.1.20 tannase synthesis synthesis of gallic esters from gallic acid and alcohols in organic solvents using enzyme microencapsulated with chitosan-alginate complex coacervate membrane. Highest yield is 44.3% in benzene, and 35.7% in hexane, best substrates are 1-propanol, 1-butanol, or 1-pentanol 3.1.1.20 tannase synthesis the enzyme immobilized on Amberlite IR retains about 85% of the initial catalytic activity even after ninth cycle of its use, immobilization method optimization, overview 3.1.1.20 tannase synthesis the mycelium-bound enzyme is useful as biocatalyst in a whole cell system at proper conditions to higher conversion of propyl gallate, the method could also reduce the cost and the time for the immobilization process 3.1.1.20 tannase synthesis tannase can be used in different industrial sectors such as in food (juices and wine) and pharmaceutical production (trimethoprim) because it catalyses the hydrolysis of hydrolysable tannins. The crude extract of Saccharomyces cerevisiae is an attractive enzyme for industrial applications, such as for beverage manufacturing and gallic acid production, due to its catalytic and thermodynamic properties (heat-stable and resistant to metal ions) 3.1.1.20 tannase synthesis tannin acyl hydrolase catalyzes the hydrolysis of hydrolyzable tannins. It is used in the manufacture of instant tea and in the production of gallic acid 3.1.1.23 acylglycerol lipase synthesis the highly stable Mortierella alliacea lipase may be useful for the synthesis of structured lipids, particularly acylglycerols containing functional unsaturated fatty acids at the sn-2 position 3.1.1.25 1,4-lactonase synthesis production of pantoic acid, which is used as a Vitamin B2-complex, production of D- and L-pantolactones, which are used as chiral intermediates in chemical synthesis 3.1.1.25 1,4-lactonase synthesis the recombinant enzyme is a highly efficient biocatalyst for asymmetric synthesis of chiral compounds 3.1.1.26 galactolipase synthesis pancreatic lipase-related protein 2 may be used to produce lipid and free fatty acid fractions enriched in either 16:3 n-3 or 18:3 n-3 fatty acids 3.1.1.32 phospholipase A1 synthesis PLA1 may be used in lysolecithin production 3.1.1.32 phospholipase A1 synthesis the enzyme mixture from Thermomyces lanuginosus/Fusarium oxysporum acts as biocatalyst in a solvent-free system containing PLA1 for the modification of phosphatidylcholine from fatty acid saponification of fish oil, overview 3.1.1.32 phospholipase A1 synthesis bio-imprinting of phospholipase A1 with 5% soybean lecithin at pH 5.0, increases its phospholipase activity in lecithin-hexane solution by 30.9fold and substantially enhances its substrate specificity. Incorporated of 105 mg diatomite/mg as an immobilization carrier further increases phospholipase activity from 562 U/g to1288 U/g 3.1.1.32 phospholipase A1 synthesis expression of phospholipase A1 in Escherichia coli results in extremely low productivity associated with inhibition of transformed cell growth. Poduction of Serratia sp. phospholipase A1 in a cell-free protein synthesis system results in an over 1000fold higher titer of functional phospholipase A1 3.1.1.32 phospholipase A1 synthesis immobilized Aspergillus oryzae overexpressing phospholipase A1 and cultivated in the presence of reticulated polyurethane foams, shows extracellular phospholipase A1 activities of 51.2-62.0 U/ml after 96-120 h. The extracellular phospholipase A1 activity of the immobilized cells at 0.5% polypeptone concentration is 34.8 U/ml. High growth rates of immobilized cells contribute to the enhanced phospholipase A1 production 3.1.1.32 phospholipase A1 synthesis maximum phospholipase A1 production of 51.55 U/ml can be achieved using defatted rice bran in solid state fermentation with moisture content of 1:1.5 and after 48h of incubation at 37°C 3.1.1.32 phospholipase A1 synthesis optimal reaction conditions migration for partial hydrolysis of soy phosphatidylcholine in hexane in order to maximize the lysophosphatidylcholine content while suppressing acyl are 60°C, 3 h reaction time, with a water content of 10% of phosphatidylcholine, and enzyme loading of 1% of phosphatidylcholine. Under these conditions, the reaction products contain 83.7 mol% lysophosphatidylcholine and are free of glycerylphosphorylcholine. Gycerylphosphorylcholine has a higher total unsaturated fatty acid content than original phosphatidylcholine has and is mainly composed of linoleic acid 3.1.1.32 phospholipase A1 synthesis preparation of L-alpha-glycerylphosphorylcholine via phospholipase A1-catalyzed hydrolysis of soy phosphatidylcholine. Optimal conditions are temperature 50°C, reaction time 30 h, water content 69 g/100 g of phosphatidylcholine weight, and enzyme loading 13 g/100 g of phosphatidylcholine weight. The optimal n-hexane-to-water ratio in the medium is 5.8:1 (v/v). L-alpha-glycerylphosphorylcholine with purity of 99.3 g/100 g is obtained 3.1.1.34 lipoprotein lipase synthesis enantiomerically enriched diarylmethanols are useful as the precursors or building blocks for the synthesis of pharmaceutically important compounds such as antihistaminic, antiarrhythmic, and anticholinergic agents. Successful application of this LPL preparation (LPL-D1) to the dynamic kinetic resolution (DKR) of diarylmethanols including aryl heteroarylmethanols, method, overview 3.1.1.40 orsellinate-depside hydrolase synthesis depsides and their constitutive untis are used in the perfume industry 3.1.1.41 cephalosporin-C deacetylase synthesis bioconversion to and from 7-aminocephalosporanic acid and deacetyl-7-aminocephalosporanic acid. Both are important intermediates in the industrial production of semisynthetic antibiotics 3.1.1.41 cephalosporin-C deacetylase synthesis production of 7-aminocephalosporanic acid, which is an intermediate for semi-synthetic penicillins and cephalosporins 3.1.1.41 cephalosporin-C deacetylase synthesis production of 7-aminocephalosporanic acid, which is an intermediate for semisynthetic penicillins and cephalosporins 3.1.1.41 cephalosporin-C deacetylase synthesis production of deacetyl 7-aminocephalosporanic acid as precursor for the synthesis of cefcapene pivoxil hydrochloride (Flomox) 3.1.1.41 cephalosporin-C deacetylase synthesis synthesis of deacetylcephalosporin C directly in a fermentation process 3.1.1.41 cephalosporin-C deacetylase synthesis enzyme reaction products deacetyl cephalosporins are a highly valuable starting material for producing semisynthetic beta-lactam antibiotics 3.1.1.41 cephalosporin-C deacetylase synthesis high-level production of cephalosporin C deacetylase by recombinant Escherichia coli. The CAH activity produced increases approximately 3 times after systematic optimization. Using glycerol feeding with pH control, an effective fermentation process for recombinant CAH production is established in a 7.0 L fermenter 3.1.1.41 cephalosporin-C deacetylase synthesis one-pot bi-enzyme catalyzed procedure for industrial conversion of cephalosporin C to deacetyl-7-aminocephalosporanic acid. Use of immobilized cephalosporin C acylase and immobilized cephalosporin C deacetylase give a yield of 78.39% deacetyl-7-aminocephalosporanic acid achieved in 30 min in a single reactor under the optimized conditions. The half-time of immobilized cephalosporin C deacetylase is approximately 18 h 3.1.1.42 chlorogenate hydrolase synthesis development of a convenient one-pot procedure for conversion of 5-caffeoylquinic acid, from coffee beans, to 3-cyclohexylpropyl caffeate, which exhibits an antiproliferative effect toward various human tumor cells. The procedure comprises of two consecutive reactions by chlorogenate hydrolase from Aspergillus japonicus and Candida antarctica lipase B, and is performed using an ionic liquid, 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide, as the reaction solvent, method optimization, overview. The system provides 12.8 mM 3-cyclohexylpropyl caffeate from 15 mM 5-caffeoylquinic acid with conversion yield of 85.3%. The reaction scarcely proceeds in 1-butyl-3-methylimidazolium tetrafluoroborate, evaluation of several ionic liquids, overview 3.1.1.43 alpha-amino-acid esterase synthesis the enzyme catalyzes amino-beta-lactam synthesis, conditions and substrates, overview 3.1.1.43 alpha-amino-acid esterase synthesis the enzyme is a promising biocatalyst for the production of semi-synthetic beta-lactam antibiotics, penicillins and cephalosporins 3.1.1.59 juvenile-hormone esterase synthesis the high tolerance of organic solvents may make JHE useful in future applications as a synthetic catalyst 3.1.1.60 bis(2-ethylhexyl)phthalate esterase synthesis optimum conditions for the production of the maximum (30.5 U) intracellular esterase, are 10 mM di(2-ethylhexyl)phthalate and 72 h incubation at pH 8.0 were found as optimum conditions 3.1.1.67 fatty-acyl-ethyl-ester synthase synthesis comparison of three different fatty acid ethyl ester-producing strains of Saccharomyces cerevisiae. Strain CB2I20, with high expression of a heterologous wax ester synthase gene (ws2) and strain BdJ15, containing disruptions of genes DGA1, LRO1, ARE1, ARE2 and POX1, which prevent the conversion of acyl-CoA to sterol esters, triacylglycerides and the degradation to acetyl-CoA, trigger oxidative stress that consequently influences cellular growth. In strain BdJ15 stress is possibly triggered by disabling the buffering capacity of lipid droplets in encapsulating toxic fatty acids such as oleic acid. There is increased demand for NADPH required for the reduction steps in fatty acid biosynthesis 3.1.1.68 xylono-1,4-lactonase synthesis an engineered Escherichia coli strain, with functional co-expression of a xylose dehydrogenase (gene xdh) and a xylonolactonase (gene xylC) from Caulobacter crescentus, has a promising perspective for large-scale production of xylonate 3.1.1.68 xylono-1,4-lactonase synthesis construction of a route for glycolate production in Escherichia coli by introducing NAD+-dependent xylose dehydrogenase xdh and xylonolactonase xylC from Caulobacter crescentus. The engineered strain produces 28.82 g/L glycolate from xylose with 0.60 g/L/h productivity and 0.38 g/g xylose yield. 27.18 g/L acetate accumulates after fermentation. An ackA knockout results in about 66% decrease in acetate formation. The final engineered strain produces 43.60 g/L glycolate, with 0.91 g/L/h productivity and 0.46 g/g xylose yield 3.1.1.72 acetylxylan esterase synthesis synthesis of peracetic acid using recombinant acetylxylan esterase and subsequent use of peracetic acid for the pretreatment of lignocellulosic biomass in situ, overview 3.1.1.72 acetylxylan esterase synthesis use of thermostable acetylxylan esterase from Thermobifida fusca and its synergistic action with xylanase for the production of xylooligosaccharides, overview. Cooperative enzymatic treatment of oat-spelt xylan by transformant xylanase and acetylxylan esterase 3.1.1.73 feruloyl esterase synthesis ferulic, p-coumaric, caffeic, and sinapinic acid have widespread industrial potential due to their antioxidant properties 3.1.1.73 feruloyl esterase synthesis isolation of phenolic acids from lignocellulosic wastes as precursors for the synthesis of chemicals 3.1.1.73 feruloyl esterase synthesis potential application in the bioconversion of lignocellulosic wastes yielding ferulic acid as precursor for chemical synthesis 3.1.1.73 feruloyl esterase synthesis potential for the production of ferulic acid as antioxidant or flavor precursor 3.1.1.73 feruloyl esterase synthesis potential for the production of ferulic acid as antioxidant, food preservative, anti-inflammatory agent, photoprotectant or flavor precursor 3.1.1.73 feruloyl esterase synthesis potential for the production of ferulic acid as antioxidant, food preservative, anti-inflammatory and antitumor agent, photoprotectant or flavor precursor 3.1.1.73 feruloyl esterase synthesis potential use for the production of ferulic acid as precursor for the synthesis of sun-blocker or food flavors 3.1.1.73 feruloyl esterase synthesis synthesis of pentylferuate as antioxidant and flavor precursor 3.1.1.73 feruloyl esterase synthesis feruloyl esterase has been identified as a key enzyme involved in microbial transformations of ferulic acid to vanillin. Ferulic acid can be produced using microencapsulated Lactobacillus fermentum (ATCC 11976) with significant levels of biological feruloyl esterase activity 3.1.1.73 feruloyl esterase synthesis synthesis of ferulic acid, an antioxidant and flavour presursor with a fusion protein of the Trichoderma reesei swollenin I (SWOI) and Aspergillus niger feruloyl esterase A (FAEA). The release of ferulic acid from wheat bran during a period of 24 h of enzymatic hydrolysis with the SWOI-FAEA improved the efficiency of ferulic acid release by 50% compared with the results obtained using the free FAEA and SWOI 3.1.1.73 feruloyl esterase synthesis feruloyl esterase, immobilized on mesoporous silica, is useful in the development of biocatalysts for customization of the antioxidant properties of hydroxycinnamic acids 3.1.1.73 feruloyl esterase synthesis the enzyme is used for preparation of ferulic acid, synergistic effect of feruloyl esterase and xylanase from Aspergillus usamii in preparation of ferulic acid by degradation of wheat bran 3.1.1.73 feruloyl esterase synthesis codon otimization for expression in Pichia pastoris increases enzyme yield about 6.4fold to 35.1 U/ml in shake flask cultivation 3.1.1.73 feruloyl esterase synthesis enzyme immobilized on mesoporous silica by physical adsorption shows more than 3fold variation in the transesterification/hydrolysis products molar ratio depending on the reaction buffer used and its pH. Hydrolysis is the dominant activity of the enzyme. Km is not affected by the immobilization but kcat is reduced 10fold. The immobilized enzyme retains more than 20% of its activity after ten cycles of transesterification reaction 3.1.1.73 feruloyl esterase synthesis use of enzyme for transesterification reactions based on vinyl ferulate. After optimization, competitive transesterification yields are obtained for prenyl ferulate (87.5-92.6%) and L-arabinose ferulate (56.2-61.7%) at reduced reaction times ( below 24 h) resulting in more than 1 g/l/h, and more than 300 kg product/kg feruloyl esterase. The enzyme can be recycled for six consecutive cycles retaining 66.6% of the synthetic activity and 100% of the selectivity 3.1.1.73 feruloyl esterase synthesis use of feruloyl esterases, immobilized in cross-linked enzyme aggregates for transesterification towards the production of prenyl ferulate and arabinose ferulate 3.1.1.73 feruloyl esterase synthesis use of feruloyl esterases, immobilized in cross-linked enzyme aggregates for transesterification towards the production of prenyl ferulate and arabinyl ferulate. with isoform Fae125, optimum product yields obtained are 83.7% for prenyl ferulate and 58.1% for arabinyl ferulate. FAE125 aggregates are stable in the optimum conditions of transesterification reactions, maintaining 70% residual activity after five consecutive reactions 3.1.1.74 cutinase synthesis a high enzyme production, high specific enzyme activity, and high enzyme yield are obtained upon expression with a 5% air saturation of oxygen. At low dissolved oxygen concentration, enzyme yield and specific activity increase with increase of culture pH-value from 5.25 to 6.25 3.1.1.74 cutinase synthesis application of enginered mutant T101A/Q132A/I218A in synthetic fiber biotransformation 3.1.1.74 cutinase synthesis recombinant cutinase from Fusarium solani pisi is used as a catalyst in enzymatic transesterification between a mixture of triglyceride oils and methanol for biodiesel production in a bis(2-ethylhexyl) sodium sulfosuccinate (AOT)/isooctane reversed micellar system, kinetics, overview 3.1.1.74 cutinase synthesis immobilized cutinase HiC from the ascomycete Humicola insolens is applied as a biocatalyst for the synthesis of functionalized acryclic esters by transesterification using transesterification of methyl acrylate with 6-mercapto-1-hexanol at a high molar ratio in a solvent free system as a model reaction 3.1.1.74 cutinase synthesis enzyme synthesizes butyl butanoate with a maximum esterification efficiency of 96.9% at 4 h 3.1.1.74 cutinase synthesis expression of enzyme in Saccharomyces cerevisiae. Approximately 28% of the cutinase localizes to the cell walls and/or between cell wall and cell membrane. Protoplasts entrapped in a membrane capsule with a low-viscous liquid-core of 1.92 % w/v calcium-alginate in a static culture secrete measurable amounts of cutinase into the broth. The entrapped protoplasts are cultivated in a shake flask at low osmotic pressure without disruption. During 60 h of cultivation, the extracellular cutinase activity of the free protoplasts at 29.3 atm and protoplasts entrapped in the capsule at 17.2 atm are 0.13 and 0.39 U/mL, respectively 3.1.1.74 cutinase synthesis growth of Fusarium oxysporum MTCC 2480 on agro industrial wastes as inducer of cutinase production. Cutin isolated from peels of multi green colored watermelon yield 6.77 U/ml as compared to 9.64 U/ml of cutinase using apple cutin. The ester linkages in water melon cutin are completely hydrolyzed during submerged fermentation 3.1.1.74 cutinase synthesis synthesis of tyrosyl esters of various aliphatic fatty acids by a recombinant cutinase. The reaction system consists of an aqueous phase saturated with the corresponding fatty-acid vinyl ester. The maximum yield of tyrosyl butanoate achieved is 60.7% after 4 h at 20°C, pH 7.0 3.1.1.74 cutinase synthesis the cutinase exhibits strong synthetic activity for butyl butanoate in non-aqueous environment,under the optimized reaction conditions esterification efficiency of 95% is observed 3.1.1.74 cutinase synthesis use of enzyme for polycondensation. Under thin film conditions the covalently immobilized enzyme catalyzes the synthesis of oligoesters of dimetil adipate with different polyols leading to Mw of about 1900 and Mn of about 1000. Immobilized Cut1 retains 37% of hydrolytic activity 3.1.1.74 cutinase synthesis use of enzyme for the synthesis of aliphatic esters. The maximum yield of ethyl caproate reaches 99.2% at a cutinase concentration of 50 U/ml, 40°C, and water content of 0.5%. The cutinase-catalyzed esterification displays strong tolerance for water content (up to 8%) and acid concentration (up to 0.8 M). Ester yields of more than 98% and 95% are achieved for acids of C3-C8 and alcohols of C1-C6, respectively 3.1.1.75 poly(3-hydroxybutyrate) depolymerase synthesis growth at 45°C, in minimal medium (pH7.0) containing glucose and 0.3% poly(2-hydroxybutanoate) as enzyme inducer, enhances enzyme production. Maximum activity is obtained after 24 h of incubation 3.1.1.75 poly(3-hydroxybutyrate) depolymerase synthesis highest extracellular depolymerase enzyme activity is achieved when 0.25% (w/v) of poly(3-hydroxybutanoate) and 1 g/L of urea are used as carbon and nitrogen source, respectively, in the culture media 3.1.1.75 poly(3-hydroxybutyrate) depolymerase synthesis maximum production of polyhydroxybutanoate depolymerase (6 U/mL) by 72 h when grown in mineral salt medium containing 0.2 % w/v polyhydroxybutanoate, pH 5.0, at 30 °C and 200 rpm shaking conditions 3.1.1.76 poly(3-hydroxyoctanoate) depolymerase synthesis the enzyme migt be used as biocatalyst for the production of enantiopure hydroxyalkanoic acids and oligomers as building blocks for the synthesis of biobased polymers 3.1.1.76 poly(3-hydroxyoctanoate) depolymerase synthesis construcution of a biosynthetic pathway for the production of (R)-3-hydroxyalkanoates through in vivo depolymerization of poly(3-hydroxyalkanoates) in an Escherichia coli fadA mutant WA101 by introducing the Pseudomonas sp. 61-3 polyhydroxyalkanoate synthase gene (phaC2) and the Pseudomonas aeruginosa intracellular polyhydroxyalkanoate depolymerase gene (phaZ). Upon culture in Luria-Bertani (LB) medium containing 2 g/l of sodium decanoate, 3-hydroxyalkanoats can be produced to the concentration of 0.49 g/l. The mole fractions are 7.5 mol% of 3-hydroxybutanoate, 31.6 mol% of 3-hydroxyhexanoate, 30 mol% of 3-hydroxyoctanoate, 29.4 mol% of 3-hydroxydecanoate, and 1.5 mol% of 3-hydroxydodecanoate 3.1.1.76 poly(3-hydroxyoctanoate) depolymerase synthesis engineering of Rhodospirillum rubrum to synthesize a heteropolymer of mainly 3-hydroxydecanoic acid and 3-hydroxyoctanoic acid from CO- and CO2-containing artificial synthesis gas (syngas) using Pseudomonas putida 3-hydroxyacyl acyl carrier protein thioesterase, a medium-chain-length fatty acid coenzyme A ligase, and an medium-chain-length polyhydroxyalkanoate synthase. A heteropolymer of 3-hydroxydecanoic acid and 3-hydroxyoctanoic acid accumulates to up to 7.1% (wt/wt) of the cell dry weight. The recombinant strains are able to partially degrade the polymer, and the deletion of isoform PhaZ2 does not reduce mobilization of the accumulated polymer significantly. Mutation C176A in the active site of PhaZ2 leads to a slight increase in poly-3-hydroxyalkanoate accumulation 3.1.1.79 hormone-sensitive lipase synthesis the enzyme catalyzes the hydrolysis of fatty acid esters at very low temperature and is therefore of great potential industrial and pharmaceutical interest 3.1.1.79 hormone-sensitive lipase synthesis the glutaraldehyde-medited crosslinked enzyme, showing increased stability, can be used as a biocatalyst in biotransformation processes 3.1.1.100 chlorophyllide a hydrolase synthesis the utilization of BciC provides mild conditions (45°C, 10 min) that may be useful for the in vitro preparation of various chemically (un)stable chlorophyllous pigments 3.1.1.110 xylono-1,5-lactonase synthesis construction of a pathway for glycolate production from xylose using xylose dehydrogenase (XdH) and xylonolactonase (xylC). The final engineered strain produces 43.60 g/l glycolate, with 0.91 g/l/h productivity and 0.46 g/g xylose yield 3.1.1.110 xylono-1,5-lactonase synthesis for synthesis of D-xylonate in Saccharomyces cerevisiae, coexpression of XylC with D-xylose dehydrogenase XylB facilitates rapid opening of the lactone and more D-xylonate is initially produced than in its absence. The lactone and linear forms of D-xylonic acid are produced, accumulate intracellularly, and are partially exported within 15-60 min of D-xylose provision 3.1.1.110 xylono-1,5-lactonase synthesis production of xylonate from xylose in Escerichia coli. Through the coexpression of a xylose dehydrogenase (XdH) and a xylonolactonase (XylC) from Caulobacter crescentus, the recombinant strain can convert 1 g/l xylose to 0.84 g/l xylonate and 0.10 g/l xylonolactone. After disruption of endogenous genes XylA and XylB encoding xylose isomerase and xylulose kinase, the finally engineered strain under fed-batch conditions, produces up to 27.3 g/l xylonate and 1.7 g/l xylonolactone from 30 g/l xylose, about 88% of the theoretical yield 3.1.1.110 xylono-1,5-lactonase synthesis when engineering Saccharomyces cerevisiae for production of D-xylonate from D-xylose, a strain expressing the D-xylonolactone lactonase xylC along with D-xylose dehydrogenase xylB excretes more D-xylonate earlier during the cultivation, both at pH 5.5 and at pH3, than the strains lacking the lactonase. In Sacchaormyces cerevisiae, the linear form may be more toxic than the lactone form and a more gradual hydrolysis of the lactone form may be advantageous to the cell 3.1.1.113 ethyl acetate hydrolase synthesis expression of EstZ in Escherichia coli KO11 reduces the concentration of ethyl acetate in fermentation broth (4.8xa0% ethanol) to less than 20 mg/l 3.1.1.117 (4-O-methyl)-D-glucuronate---lignin esterase synthesis due to its thermostability, enzyme CkXyn10C-GE15A is a promising candidate for industrial processes, with both catalytic domains exhibiting melting temperatures over 70°C. Of particular interest is the glucuronoyl esterase domain, as it represents the first studied thermostable enzyme displaying this activity 3.1.1.117 (4-O-methyl)-D-glucuronate---lignin esterase synthesis Thermothielavioides terrestris glucoronoyl esterases TtGEs can be used as promising accessory enzymes to improve the hydrolysis efficiency of commercial enzymes in saccharification of lignocellulosic materials due to their thermophilic characteristics 3.1.3.1 alkaline phosphatase synthesis overexpression of enzyme in Escherichia coli. After 1 day of growth, when the Escherichia coli culture is near the stationary phase, high amounts of enzyme are produced but the alkaline phosphatase activity in the cell-free extract is near the background level. Further incubation of bacterial culture leads to a tremendous increase in alkaline phosphatase activity which is maximal at the 3rd day of incubation and was 48–100 times higher than at the 1st day of growth. Typically, 90-140 mg of active protein is produced in 1 l of culture 3.1.3.2 acid phosphatase synthesis the enzyme is useful in preparative dephosphorylation of compounds, in preparative phosphorylation of glucose and related compounds, and in the phosphorylation of structurally different alcoholic substrates with diphosphate as cheap phosphate donor 3.1.3.2 acid phosphatase synthesis immobilization of enzyme onto glutaraldehyde activated chitosan beads. Chitosan beads activated with 2% of glutaraldehyde demonstrate maximum immobilization yield of about 83%. The immobilized enzyme shows optimum activity at pH 7.0, while the soluble form is maximally active at pH 5.0. Both soluble and immobilized acid phosphatase exhibit maximum activity at 60°C. The immobilization on chitosan beads enhances the shelf life of acid phosphatase. The immobilized enzyme retains its more than 50% hydrolytic activity for approximately two months and can be reused for more than 40 cycles of reaction 3.1.3.7 3'(2'),5'-bisphosphate nucleotidase synthesis construction of a protein expression vector using 3',5'-bisphosphate nucleotidase HAL2 in the presence of Ca2+ as an affinity tag. The tag can be used for procaryotic protein expression and purification by maintaining Ca2+ for efficient affinity binding and chelating Ca2+ for elution 3.1.3.8 3-phytase synthesis preparation of myo-inositol phosphates as tools for metabolic investigation, enzyme stabilizers, as enzyme inhibitors and therefore potential drugs 3.1.3.8 3-phytase synthesis expression of phytase in a Medicago truncatula cell suspension line. Recombinant phytase accumulates to at least 25 mg/l and remaines stable along the growth curve, and an enriched fraction with high enzymatic activity is easily obtained 3.1.3.8 3-phytase synthesis expression of phytase in Glycine max under control of a root-specific promoter. The phytase activity and phosphate levels in transgenic soybean root secretions are 4.7 U/mg protein and 439 microM, respectively, compared to 0.8 U/mg protein and 120 microM, respectively, in control soybeans 3.1.3.8 3-phytase synthesis expression of phytase in Lactobacillus casei. The highest phytase activities in the supernatant and cells are 22.12 and 4.49 U per ml at the fourth day of incubation 3.1.3.8 3-phytase synthesis immobilization of enzyme via a cross-linked enzyme aggregate. The immobilized enzyme incubated with vanadate shows a similar efficiency and asymmetric induction as the free enzyme and can be used at least three times without significant loss of activity. In presence of organic solvents, the activtiy is still limited. Vanadate is coordinatecd to oxygen functions at two different binding sites, and the alpha-helical content decreases upon coordination of vanadate 3.1.3.8 3-phytase synthesis immobilization of phytase on Sepabead EC-EP and use in the biodegradation of soymilk phytate. The immobilized enzyme exhibits an activity of 0.1 U per g of carrier and activity yield of 70.83%. Optimum temperature is 55°C, optimum pH for the immobilized enzyme is 5.5. Enzyme is stable between pH 3.0-8.0 and below 70°C. The immobilized enzyme hydrolyzes 65% of soymilk phytate in 8 h at 60°C, as compared with 56% hydrolysis observed for the native enzyme over the same period of time 3.1.3.8 3-phytase synthesis the optimal medium for phytase production contains oat 10.0 g/l, ammonium sulfate 15.0 g/l, glucose 30 g/l, and NaCl 20.0 g/l, while the optimal cultivation conditions for phytase production are pH 5.0, a temperature of 28°C, and a shaking speed of 170 rpm. Under the optimal conditions, over 557.9 mU/ml of phytase activity is produced within 72 h of fermentation at the shake flask level 3.1.3.8 3-phytase synthesis a maximal level of phytase of 113.7 U/g of dry substrate is obtained in wheat bran based medium containing 5% sucrose, 50% humidity, 7.5% of biomass at 33°C, pH 7.0 during 72 h and a 285% improvement in enzyme titre is achieved 3.1.3.11 fructose-bisphosphatase synthesis enzyme is a limiting factor for lysine production by Corynebacterium glutamicum. In strains overexpression enzyme lysine yields on glucose and/or fructose as carbon source remain unchanged, but lysine yield on sucrose increases twofold 3.1.3.11 fructose-bisphosphatase synthesis overexpression of enzyme increases the lysine yield by 40% for growth on glucose and by 30% for growth on fructose or sucrose. Overexpression causes a redirection of carbon flux from glycolysis toward the pentose pathway and thus leads to increased NADPH supply. Use of overexpression for starch- and molasses-based industrial lysine production 3.1.3.11 fructose-bisphosphatase synthesis the substrate for the sedoheptulose 1,7-bisphosphatase reaction, sedoheptulose 1,7-bisphosphate, can be synthesized in vitro by using both the chromosomal-encoded or the plasmid-encoded fructose 1,6-bisphosphate aldolase proteins from Bacillus methanolicus 3.1.3.11 fructose-bisphosphatase synthesis the substrate for the sedoheptulose 1,7-bisphosphatase reaction, sedoheptulose 1,7-bisphosphate, can be synthesized in vitro by using both the chromosomal-encoded or theplasmid-encoded fructose 1,6-bisphosphate aldolase proteins from Bacillus methanolicus 3.1.3.21 glycerol-1-phosphatase synthesis synthesis of glycerol by direct conversion of solar energy 3.1.3.22 mannitol-1-phosphatase synthesis heterologous expression of a fusion enzyme, comprising a mannitol-1-phosphate dehydrogenase and a mannitol-1-phosphatase, leads to synthesis of mannitol by Escherichia coli and by the cyanobacterium Synechococcus sp. PCC 7002 3.1.3.25 inositol-phosphate phosphatase synthesis construction of a pathway to produce inositol from glucose using polyphosphate glucokinase PPGK from Arthrobacter sp. OY3WO11, inositol 1-phosphate synthase IPS from Trypanosoma brucei TREU927 and inositol monophosphatase IMP Escherichia coli, all expressed in Escherichia coli. The conversion ratio from glucose to inositol reaches 90% 3.1.3.25 inositol-phosphate phosphatase synthesis production of myo-inositol from glucose by a novel trienzymatic cascade of polyphosphate glucokinase, inositol 1-phosphate synthase and inositol monophosphatase. myo-Inositol (inositol) is important in the cosmetics, pharmaceutical and functional food industries. The conversion ratio from glucose to inositol reaches 90%, which is promising for the enzymatic synthesis of inositol without ATP supplementation 3.1.3.26 4-phytase synthesis preparation of myo-inositol phosphates as tools for metabolic investigation, enzyme stabilizers, as enzyme inhibitors and therefore potential drugs 3.1.3.27 phosphatidylglycerophosphatase synthesis preparation of phosphatidylglycerophosphate 3.1.3.69 glucosylglycerol 3-phosphatase synthesis plasmid-bound expression of glucosylglycerol synthase and glucosylglycerol phosphatase from Synechocystis sp. PCC 6803 enables alpha-D-glucosylglycerol synthesis exclusively in osmotically stressed cells of Corynebacterium glutamicum. Upon deletion of both the glgA encoded glycogen synthase and the otsA encoded trehalose-6-phosphate synthase, the alpha-D-glucosylglycerol concentration in culture supernatants is increased from 0.5 mM in to 2.9 mM. Upon nitrogen limitation, transgenic Corynebacterium produces more than 10 mM alpha-D-glucosylglycerol (about 2 g/l) 3.1.3.72 5-phytase synthesis transgenic expression in Pichia pastoris. Increasing isoform Alp2 copy number is detrimental to heterologous expression, clones with one copy of wild-type Alp2 produce the highest activity, clones with two, four and seven or more copies produce 70%, 25% and 10% respectively, of enzyme activity. Use of a sequence-optimized Alp2 gene increases the yield of the active enzyme by 25-50% in one/two copy clones. Reducing the temperature during heterologous expression leads to increases of 1.2-20fold suggesting that protein folding and post-translational processes may be the dominant factors limiting ALP2 expression 3.1.4.1 phosphodiesterase I synthesis production of 5'-mononucleotides, degradation of 2',5'-dinucleotides in nuclease P1 digest of technical grade yeast RNA 3.1.4.1 phosphodiesterase I synthesis immobilized enzyme: production of 5'-ribonucleotides, starting substances for the preparation of food additives and drugs 3.1.4.1 phosphodiesterase I synthesis addition of autoclaved microbial cell suspensions to Catharanthus roseus cell cultures results in inhibition of growth with stimulation of enzyme production. A combination of Aleteromonas macleodii and 0.1% w/v alginate oligomers minimizes cell growth inhibition and enhances enzyme production 20times above control. Mixed alginate elecitors significantly promote phosphodiesterase production up to 120fold 3.1.4.4 phospholipase D synthesis heterologous high-yield expression of enzyme in Pichia pastoris and purification by single calcium-dependent octyl-sepharose chromatography step gives 8 mg of pure recombinant protein from 1 l of yeast culture without proteolytic degradation of the N-terminal calcium-dependent C2 lipid binding domain 3.1.4.4 phospholipase D synthesis production of enzyme in Escherichia coli is strongly influenced by the catalytic activity as cytotoxicity of protein for host is strictly coorelated to enzyme activity. Synthesis of inactive mutants gives recovery yields 5-10 times higher than for wild-type 3.1.4.4 phospholipase D synthesis creation of a high performance PLD with superior transphosphatidylation activity and low hydrolysis activity can be very useful for the synthesis of phospholipids used in pharmaceuticals, foods, cosmetics, and other industries 3.1.4.4 phospholipase D synthesis phospholipase D is a useful enzyme for its transphosphatidylation activity, which enables the enzymatic synthesis of various phospholipids, natural and unnatural phospholipids, and phospholipids with a functional head group, detailed overview 3.1.4.4 phospholipase D synthesis hyperactivation of PLD by bio-imprinting and immobilization by adsorption and precipitation, followed by cross-linking results in increased catalytic activity. The maximum activity of immobilized PLD reaches 166953 U/g protein, compared to 11922 U/mg protein for the free form. The selectivity of PLD is significantly enhanced after immobilization. The yield of phosphatidylglycerol and phosphatidic acid reaches 94.0% and 5.96%, respectively 3.1.6.1 arylsulfatase (type I) synthesis purification method for removing phytoestrogens and related compounds without affecting the enzyme activity. Use of solid phase extraction on a polymeric resin for purification 3.1.6.1 arylsulfatase (type I) synthesis the enzyme could be useful for producing a low sulfated agar and electrophoretic grade agarose 3.1.6.1 arylsulfatase (type I) synthesis immobilization of enzyme on carboxyl functioned magnetic nanoparticles using glutaraldehyde. Immobilization leads to increased tolerance to some metal ions, inhibitors and detergents. Immobilized arylsulfatase maintains 61.7% of its initial desulfuration rates after seven cycles. After the reaction of agar with immobilized arylsulfatase for 90 min at 50°C, 46% of the sulfate in the agar is removed 3.1.6.4 N-acetylgalactosamine-6-sulfatase synthesis production and characterization of an active recombinant N-acetylgalactosamine-6-sulfate sulfatase in Escherichia coli BL21. When native signal peptide is present, higher enzyme activity levels are observed in both soluble and inclusion bodies fractions, and signal peptide removal has a significant impact on enzyme activation. Enzyme activity in the culture media is only detected when signal peptide is presented and the culture is carried out under semi-continuous mode 3.1.6.4 N-acetylgalactosamine-6-sulfatase synthesis increase of recombinant GALNS activity produced in Escherichia coli is obtained with a promoter regulated under sigmas. Additional improvements are observed when osmotic shock is applied. Overexpression of chaperones has no effect on recombinant GALNS activity. High concentrations of sucrose in conjunction with the physiological-regulated promoter proUmod significantly increase the GALNS production and activity 3.1.6.4 N-acetylgalactosamine-6-sulfatase synthesis removal of the native signal peptide and coexpression with human formylglycine-generating enzyme SUMF1 in Pichia pastoris allow an improvement of 4.5fold in the specific GALNS activity. Recombinant GALNS shows a high stability at 4°C, while the activity is markedly reduced at 37 and 45°C. Recombinant GALNS is taken-up by HEK-293 cells and human skin fibroblasts in a dose-dependent manner, without any additional protein or host modification 3.1.6.13 iduronate-2-sulfatase synthesis expression and purification of enzyme from Escherichia coli, strategy for improving protein expression and purification 3.1.6.13 iduronate-2-sulfatase synthesis expression of protein in Pichia pastoris. The highest production of recombinant IDS is obtained at oxygen-limited conditions using a codon-optimized IDS cDNA.The purified enzyme shows a final activity of 12.45 nmol/mg/h. IDS shows high stability in human serum and is taken up by HEK-293 cells in a dose-dependent manner through mannose receptors 3.1.6.19 (R)-specific secondary-alkylsulfatase (type III) synthesis stereoselctive organic synthesis, stereoselective and enantioselective biohydrolysis of sulfate esters of sec-alcohols 3.1.6.19 (R)-specific secondary-alkylsulfatase (type III) synthesis stereoselective organic synthesis, enantioselective hydrolysis of rac-secondary-alkyl sulphate esters 3.1.7.11 geranyl diphosphate diphosphatase synthesis coexpression of geranyl diphosphate synthase and geraniol synthase in Escherichia coli. Gene copy number optimization leads to a 1.6fold increase of geraniol production when four copies of geranyl diphosphate synthase and one copy of geraniol synthase are used. The additional fermentation conditions optimization, including removal of organic layers and addition of n-decane, leads to a geraniol production of 74.6 mg/l 3.1.7.11 geranyl diphosphate diphosphatase synthesis overexpression of a mutant GES with a 5'-untranslated sequence designed for high translational efficiency, along with the additional expression of isopentenyl diphosphate isomerase, and geranyl diphosphate synthase, yields 300 mg/l/12 h geraniol and its derivatives in a shaking flask 3.1.7.11 geranyl diphosphate diphosphatase synthesis significant increase in geraniol synthesis is achieved by coexpression of geraniol synthase, mutant S80F of farnesyl diphosphate synthase, and the encoding genes involved in the whole mevalonic acid biosynthetic pathway in Escherichia coli. The additional optimization of medium composition, fermentation time, and addition of metal ions leads to the geraniol production of 48.5 mg/l 3.1.7.11 geranyl diphosphate diphosphatase synthesis synthesis of geraniol by expression of GES truncated at residue S43 in Saccharomyces cerevisiae. Coexpression of the reverse fusion of farnesyl diphosphate synthase Erg20 mutant F96W-N127W and truncated GES and another copy of farnesyl diphosphate synthase Erg20 mutant F96W-N127W promotes the geraniol titer to 523.96 mg/l at shakes flask level. A highest reported titer of 1.68 g/l geraniol in eukaryote cells is achieved in 2.0 l fed-batch fermentation under carbon restriction strategy 3.1.7.13 neryl diphosphate diphosphatase synthesis biosynthesis of nerol from glucose in the metabolic engineered Escherichia coli. The truncated neryl diphosphate synthase and nerol synthase are co expressed in Escherichia coli, the engineered strain LZ001 accumulates 0.053 mg/l of nerol. Then the heterologous mevalonate pathway is constructed in the engineered strain. Finally, the overexpression of acetyl-CoA acetyltransferase from Saccharomaces cerevisiae (ERG10) increases the yield to 1.564 mg/l in recombinant strain LZ005 3.1.8.1 aryldialkylphosphatase synthesis efficient production of enzyme in Escherichia coli, by designing high cell density cultivations and a membrane-based downstream process. In fed batches, enzyme production is increased by 69fold up to 4660 U/l, using galactose as inducer. the process is scalable from 2.5 up to 150 l. An enzyme recovery of 77% with a purity grade of 80% can be reached 3.1.11.2 exodeoxyribonuclease III synthesis enzyme is used for purification of eucaryotic extrachromosomal circular DNAs using exonuclease III 3.1.11.3 exodeoxyribonuclease (lambda-induced) synthesis solid-phase digestion of dsDNAs using immobilization of lambda exonuclease onto poly(methylmethacrylate) micropillars populated within a microfluidic device for the on-chip digestion. The efficiency for the catalysis of dsDNA digestion using lamba-exonuclease, including its processivity and reaction rate, are higher when the enzyme is attached to a solid support compared to the free solution digestion. A clipping rate of 1000 nucleotides per s can be obtained for the digestion of lambda-DNA (48.5 kbp) by lambda-exonuclease 3.1.26.5 ribonuclease P synthesis the bacterial RNase P ribozyme might be used to release RNAs of interest with homogeneous 3'-OH ends from primary transcripts via site-specific cleavage, overview. Also, T7 transcription of mature tRNAs with clustered G residues at the 5'-end may result in 5'-end heterogeneities, which can be avoided by first transcribing the 5'-precursor tRNA (ptRNA) followed by P RNA-catalyzed processing to release the mature tRNA carrying a homogeneous 5'-monophosphate end 3.1.30.1 Aspergillus nuclease S1 synthesis removal of DNA loops during cDNA synthesis 3.1.30.1 Aspergillus nuclease S1 synthesis industrial production of 5'-phosphomononucleotides from yeast RNA 3.2.1.1 alpha-amylase synthesis industrial-scale starch liquefaction 3.2.1.1 alpha-amylase synthesis oligomer units are an intermediate in the high-fructose syrup production 3.2.1.1 alpha-amylase synthesis immobilization of enzyme in calcium alginate beads through entrapment technique. Activity of immobilized enzyme is 81% of free enzyme, its optimum acivity at pH 4.5-6.0 and 40°C, compared to pH 5.5 and 30°C for free enzyme. Immobilized enzyme retains its activity longer than free enzyme 3.2.1.1 alpha-amylase synthesis optimization of enzyme production parameters results in growth temperature 70°C, pH 7.75 and 84 h in nutrient medium 3.2.1.1 alpha-amylase synthesis use of probiotic Bacillus spores as a matrix for enzyme immobilization by covalent and adsorption methods. The maximum concentration of the alpha-amylase immobilized is 360 microg/1.2 10EE11spores. Maximum activity is achieved at an enzyme concentration of approximately 60 microg/0.4 10EE10 spores, corresponding to an estimated activity of 8000 IU per mg and 1.2 10EE11 spores for covalent immobilization and 85300 IU for the adsorption method. Enzyme immobilization yield is estimated to be 77% and 20.07% for the covalent and adsorption methods, respectively. The alpha-amylase immobilized by both methods, displays improved activity in the basic pH range. The optimum pH for the free enzyme is 5 while it shifts to 8 for the immobilized enzyme. The optimum temperatures for the free and immobilized enzymes are 0C and 0C, respectively. The covalently immobilized alpha-amylase retains 65% of its initial activity, even after 10 times of recycling 3.2.1.B1 extracellular agarase synthesis use of enzyme for preparation of neoagarooctaose and neoagarodecaose and separation of neoagaro-oligosaccharides by consecutive column chromatography 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis the immobilized enzyme with high operational stability can be used for continuous production of glucose from soluble dextrin 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis production of glucose, which is a feed stock for high fructose syrup 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis the enzyme of the M115 mutant strain is useful for enhanced ethanol production by Saccharomyces cerevisiae, strain ATTC26602, using raw starch as substrate in solid state fermentation 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis entrapment of amyloglucosidase into dipalmitoylphosphatidylcholine multilamellar vesicles and large unilamellar vesicles for biocatalysis inside liposomes and bioanalytical applications, overview 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis enzyme immobilization on polyacrylamide gel results in an enzyme with increases thermostability for use in biocatalysis 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis glycosylation of the phenolic hydroxyl group of the phenyl propanoid systems, eugenol and curcumin, using an amyloglucosidase from Rhizopus sp. and a beta-glucosidase from sweet almonds together with carbohydrates D-glucose, D-mannose, maltose, sucrose,and D-mannitol in di-isopropyl ether produce glycosides at 7-52% yields in 72 h, method optimization, overview, two compounds are glycosylated in order to enhance their water solubility and pharmacological activities 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis preparations of glucoamylase are widely used in many branches of industry for hydrolyzing starch-containing raw materials 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis the enzyme is industrially an important biocatalyst that decomposes starch into glucose by tearing-off alpha-1,4-linked glucose residue from the non-reduced end of the polysaccharide chain 3.2.1.3 glucan 1,4-alpha-glucosidase synthesis enzyme may be used for raw corn starch hydrolysis and subsequent bioethanol production using Saccharomyces cerevisiae. The yield in terms of grams of ethanol produced per gram of sugar consumed is 0.365 g/g, with 71.6% of theoretical yield from raw corn starch 3.2.1.4 cellulase synthesis synthesis of glyceroyl beta-N-acetyllactosaminide and derivatives, that could be used as starting material for the synthesis of neoglycolipid and new kinds of detergents and as acceptors for glycosidase and glycosyltransferase 3.2.1.4 cellulase synthesis when the enzyme is used in combination withbeta-glucosidase, cellulose is completely hydrolyzed to glucose at high temperature, suggesting great potential for EGPh in bioethanol industrial applications 3.2.1.4 cellulase synthesis glucose production from cellulose material using beta-glucosidase from Pyrococcus furioses and endocellulase from Pyrococcus horikoshii. The combination reaction can produce only glucose without the other oligosaccharides from phosphoric acid swollen Avicel 3.2.1.4 cellulase synthesis production of enzyme in parallel-operated shake flasks and, alternatively, in parallel-operated stirred-tank bioreactors on a 10-m. scale. Reaction conditions with 53.3 g/l microcrystalline cellulose in the initial medium, no lactose feeding and 3.3 g/l and day intermittent ammonium sulfate addition are optimal. The optimum substrate supply on a liter-scale results in the production of 4.88 filter paper units of enzyme per ml with after 96 h 3.2.1.4 cellulase synthesis use of a cellulase blend to evaluate its application in a simultaneous saccharification and fermentation process for second generation ethanol production from sugar cane bagasse. After enzyme production in a bioreactor and tangential ultrafiltration in hollow fiber membranes, the cellulolytic preparation is stable for at least 300 h at both 37°C and 50°C. The ethanol production is carried out by sugar cane bagasse partially delignified cellulignin fed-batch simultaneous saccharification and fermantation process, using the onsite cellulase blend. The method applied results in 100 g/l ethanol concentration at the end of the process, which corresponds to a fermentation efficiency of 78% of the maximum obtainable theoretically. The experimental results lead to the ratio of 380 l of ethanolper ton of sugar cane bagasse partially delignified cellulignin 3.2.1.4 cellulase synthesis 40% higher cellulase activity on filter paper in 72 h is observed with the addition of 1 mM of nickel-cobaltite (NiCo2O4) nanoparticles in the growth medium. Maximum production of endoglucanase (211 IU/gds), beta-glucosidase (301 IU/gds), and xylanase (803 IU/gds) is achieved after 72 h without nanoparticles, while in the presence of 1 mM of nanoparticles, endoglucanase, beta-glucosidase, and xylanase activity increase by about 49, 53, and 19.8%, respectively, after 48 h of incubation 3.2.1.4 cellulase synthesis a medium based on starch casein minerals containing carboxymethyl cellulose and beef extract supports enhanced cellulase production. Carboxymethyl cellulose, beef extract , NaCl, temperature and pH are significant for cellulase production. Optimization of cellulase production results in an enhancement of endoglucanase activity to 27 IU per ml 3.2.1.4 cellulase synthesis expression of enzyme in Escherichia coli and Thermotoga sp. after fusion to the signal peptides of TM1840 (amyA) or TM0070 (xynB). Expressed in Escherichia coli and Thermotoga sp. renders the hosts with increased endo- and exoglucanase activities. In Escherichia coli, the recombinant enzymes are mainly bound to the bacterial cells, whereas in Thermotoga sp., about half of the enzyme activities are observed in the culture supernatants. However, the cellulase activities are lost in Thermotoga sp. after three consecutive transfers 3.2.1.4 cellulase synthesis expression of enzyme in Escherichia coli and Thermotoga sp. after fusion to the signal peptides of TM1840 (amyA) or TM0070 (xynB). Expressed in Escherichia coli and Thermotoga sp. renders the hosts with increased endoglucanase activities. In Escherichia coli, the recombinant enzymes are mainly bound to the bacterial cells, whereas in Thermotoga sp., about half of the enzyme activities are observed in the culture supernatants. However, the cellulase activities are lost in Thermotoga sp. after three consecutive transfers 3.2.1.4 cellulase synthesis heterologous expression in Bacillus subtilis combined with customized signal peptides for secretion from a random libraries with 173 different signal peptides originating from the Bacillus subtilis genome. The customized signal peptide does not affect enzyme performance when assayed on carboxymethyl cellulose, phosphoric acid swollen cellulose, and microcrystalline cellulose 3.2.1.4 cellulase synthesis in regulator cre1-silenced strain C88, the filter paper hydrolyzing activity and beta-1,4-endoglucanase activity are 3.76-, and 1.31fold higher, respectively, than those in the parental strain when the strains are cultured in inducing medium for 6 days. The activities of beta-1,4-exoglucanase and cellobiase are 2.64-, and 5.59fold higher, respectively, than those in the parental strain when the strains are cultured for 5 days 3.2.1.4 cellulase synthesis optimization of cultural conditions for enhanced cellulase production. Under solid-state fermentation, yields of carboxymethylcellulase are 463.9 U/g, filter paper cellulase 101.1 U/g and beta-glucosidase 99 U/g 3.2.1.4 cellulase synthesis alkali-pretreated roots of Taraxacum kok-saghyz (rubber dandelion), incubated with crude enzyme extracts from Thermomyces lanuginosus STm yield more natural rubber (90 mg/g dry root) than the protocols, Eskew process (24 mg/g) and commercial-enzyme-combination process (45 mg/g). The crude enzyme treatment at 91.6% rubber purity approaches the purity of the commercial-enzyme-combination process at 94.1% purity 3.2.1.4 cellulase synthesis scale-up systems for cellulase production and enzymatic hydrolysis of pretreated rice straw at highsolid loadings and by Aspergillus terreus. In a horizontal rotary drum reactor at 50°C with 25 % (w/v) solid loading and 9 FPU/g substrate enzyme load up to 20 % highly concentrated fermentable sugars are obtained at 40 h with an increased saccharification efficiency of 76 % compared to laboratory findings (69.2 %). Nearly 79-84% of the cellulases and more than 90% of the sugars are recovered from the saccharification mixture 3.2.1.4 cellulase synthesis under optimised conditions of growth on wheat bran, 420.8 and 22.73 units/g substrate of endo-beta-1,4-glucanase and filter paper cellulase are produced, respectively. Both endo-beta-1,4-glucanase and filter paper activity production show significant dependence on ammonium sulfate concentration and pH 3.2.1.4 cellulase synthesis enhanced production of enzyme in Escherichia coli. High-cell-density and optimal CenC expression are obtained in ZYBM9 medium induced either with 0.5 mM IPTG/150 mM lactose, after 6 h induction at 37°C. Before induction, bacterial cells are given heat shock (42°C) for 1 h when culture density (OD600 nm) reached at 0.6. Intracellular enzyme activity is enhanced by 6.67- and 3.20fold in ZYBM9 (yeast extract 0.5% (w/v), NaCl 0.5% (w/v), tryptone 1.0% (w/v), NH4Cl 0.1% (w/v), KH2PO4 0.3% (w/v), Na2HPO4 0.6% (w/v), MgSO4.7H2O 1 mM, and Glucose 0.4% (w/v)) and 3×ZYBM9 medium, respectively, under optimal conditions 3.2.1.4 cellulase synthesis the cold active butanol-tolerant endoglucanase is valuable for biobutanol production by a simultaneous saccharification and fermentation process 3.2.1.6 endo-1,3(4)-beta-glucanase synthesis the major products of water-soluble beta-glucan hydrolyzed by over-produced endo-beta-(1-3),(1-4)-glucanase are trioligosaccharides and tetrasaccharides, which can be developed as useful products such as antihypercholesterolemic, anti-hypertriglyceridemic, and anti-hyperglycemic agents 3.2.1.7 inulinase synthesis high-level expression in Pichia pastoris leads to production of enzyme at 286.8 U/ml and 8873 U/mg 3.2.1.7 inulinase synthesis isolation of mutant M-30 with enhanced inulinase production, mutant is stable after cultivation for 20 generations. Inulin, yeast extract, NaCl, temperature, pH for maximum inulinase production by the mutant M-30 are 20.0 g/l, 5.0 g/l, 20.0 g/l, at 28°C and pH 6.5, respectively. Under the optimized conditions, 127.7 U/ml of inulinase activity is reached in the liquid culture 3.2.1.7 inulinase synthesis proposed kinetic model for fructose production defined within temperature and substrate concentration ranges of industrial interest such as 40-60°C and 3-60 g/l, respectively. Model is based on a minimum number of parameters. The hypotheses are always specified and assumed only on the basis of convenience and rational consideration. The kinetic model was successfully validated by comparison with a vast set of experimental results 3.2.1.7 inulinase synthesis application of a bi-enzymatic system based on the combined use of levansucrase from Bacillus amyloliquefaciens and endo-inulinase from Aspergillus niger in a one-step reaction for the synthesis of fructooligosaccharides and oligolevans using sucrose as the sole substrate. The optimal conditions leading to a high yield of short chain fructooligosaccharides, i.e.1:1 ratio, 0.5 h, 0.6 M, are different from those resulting in a high yield of medium chain fructooligosaccharides and oligolevans, i.e. 1.85:1 ratio, 1.77 h, 0.6 M. The production of fructooligosaccharides and oligolevans at a large scale gives a yield of 57-65%, w/w and produces 65.8-266.8 g/l and h, and uses of low temperature of 35°C and low concentrations of sucrose 3.2.1.7 inulinase synthesis gene expression in Pichia pastoris using codon optimization results in the secretion of recombinant endoinulinase activity that reaches 1349 U/ml. Inulooligosaccharides production from inulin using the recombinant enzyme, after 8 h under opti­mal conditions, which include 400 g/l inulin, an enzyme concentration of 40 U/g substrate, 50°C and pH 6.0gives a yield of 91% 3.2.1.7 inulinase synthesis growth of Aspergillus niger AUMC 9375 on the mixture of a 6:1 w/w ratio of sun flower tuber:lettuce roots, yields the highest levels of inulinase at 50% moisture, 30°C, pH 5.0, with seven days of incubation, and with yeast extract as the best nitrogen source. Purified inulinase is successfully immobilized with an immobilization yield of 71.28%. After incubation for 2 h at 60°C, the free enzyme activity decreases markedly to 10%, whereas that of the immobilized form decreases only to 87%. The immobilized inulinase can be used for 10 cycles and in addition, can be stored for 32 days at 4°C 3.2.1.7 inulinase synthesis immobilization of endoinulinase results in higher stability than the free endoinulinase under various temperature levels. A residual activity of 81.2% can be still obtained after ten reaction cycles 3.2.1.7 inulinase synthesis optimal grwoth conditions for expression of enzyme are 1% inulin,1% yeast extract, and 0.05% KH2PO4. Under optimum conditions, endoinulinase production reaches 28.67 IU/ml and biomass yield 0.162 OD600/15, in excellence correlation with predicted values. Endoinulinase production from a simple and cost-effective medium using raw Dahlia inulin is comparable with pure inulin 3.2.1.7 inulinase synthesis endoinulinase is an inulolytic enzyme which is used for the production of fructooligosaccharides from inulin 3.2.1.7 inulinase synthesis endoinulinases are an industrial tool critical for the production of inulooligosaccharides (IOS) 3.2.1.7 inulinase synthesis enzymatic hydrolyzation of inulin by endo-inulinase to produce oligofructoses, a type of food additive and health product, a promising green and environmentally friendly technique 3.2.1.7 inulinase synthesis enzymatic synthesis of fructooligosaccharides (FOS) from sucrose by endo-inulinase-catalyzed transfructosylation reaction in biphasic systems, production of FOS from sucrose by commercial inulinase from Aspergillus niger 3.2.1.7 inulinase synthesis inulooligosaccharides (IOS) represent an important class of oligosaccharides at industrial scale. Efficient conversion of inulin to IOS through endoinulinase from Aspergillus niger 3.2.1.8 endo-1,4-beta-xylanase synthesis optimum levels of wheat bran (15-20 g/l), lactose (1.0-1.5 g/l), tryptone (2-2.5 g/l) and NaCl (7.0-8.0 g/l) support a 6.75fold increase in xylanase production 3.2.1.8 endo-1,4-beta-xylanase synthesis production of enzyme in Pichia pastoris with codon optimization. The activity of dual-copy enzyme is maximized at 15158 U/ml after 120 h of shaking 3.2.1.8 endo-1,4-beta-xylanase synthesis yield of enzyme is enhanced more than four fold in the presence of 1% corn husk and 0.5% peptone or feather hydrolysate at pH 11 and 37°C 3.2.1.8 endo-1,4-beta-xylanase synthesis recombinant expression of endo-beta-1,4-xylanase in Pichia pastoris. Codon optimization leads to 59% increase in activity. Coexpression of the Vitreoscilla hemoglobin (VHb) gene leads to higher biomass, cell viability, and xylanase activity. The maximum xylanase activity reaches 58792 U/ml when the induction temperature is 22°C 3.2.1.8 endo-1,4-beta-xylanase synthesis alkali-pretreated roots of Taraxacum kok-saghyz (rubber dandelion), incubated with crude enzyme extracts from Thermomyces lanuginosus STm yield more natural rubber (90 mg/g dry root) than the protocols, Eskew process (24 mg/g) and commercial-enzyme-combination process (45 mg/g). The crude enzyme treatment at 91.6% rubber purity approaches the purity of the commercial-enzyme-combination process at 94.1% purity 3.2.1.8 endo-1,4-beta-xylanase synthesis optimization of recombinant enzyme production in 1-liter flasks. Initial cell density is the most important parameter. Under optimized conditions, 1498 mg xylanase per liter can be achieved 3.2.1.8 endo-1,4-beta-xylanase synthesis Bacillus licheniformis strain DM5 is attributed for the production of prebiotic and anti-inflammatory XOS from agrowaste 3.2.1.11 dextranase synthesis - 3.2.1.11 dextranase synthesis manufacture of dentifrices 3.2.1.11 dextranase synthesis application in sugar cane mills 3.2.1.11 dextranase synthesis investigation on various adding times of dextranase to the dextransucrase system to reveal the synergistic processes of dextransucrase and dextranase. Dextranase added into the dextransucrase-sucrose system at different times gives rise to different main dextran products. Dextranase added into sucrose system at the same time with dextransucrase synthesizes low molecular weight dextran targeted to 5 kDa, while dextranase added during the reaction process of dextransucrase directionally prepares dextran with medium Mw of 10 kDa and 20 kDa. The synthesized oligodextrans are mainly composed of alpha-1,6-glycosidic linkages and low alpha-1,3-glycosidic branches 3.2.1.11 dextranase synthesis maximum enzyme production is obtained in a self designed medium (pH 6.0) containing 1% dextran 5000 Da, after 24 h culture incubation at 37°C.High dextranase production is achieved when medium is supplemented with dextran as only carbon source and no enzyme production is found in medium having glucose, sucrose or starch. Dextranase production is inversely proportional to dextran polymer length and extent of branching 3.2.1.11 dextranase synthesis optimal conditions for synthesis of enzyme are 28 h, 8.0, 30°C, and 25% volume of liquid in 100-ml Erlenmeyer flasks, respectively 3.2.1.11 dextranase synthesis optimized culture conditions for dextranase productions are 37°C, pH 10, 32 h, and 20% (v/w) moisture content. The addition of 0.175 mM CrCl3 increases the enzyme production by about 4.5fold 3.2.1.11 dextranase synthesis immobilization of enzyme in Fe3+-cross-linked alginate/carboxymethyl cellulose beads. The immobilization process improves the optimum temperature from 35°C to 45°C. The immobilized enzyme shows its optimum activity for synthesis of dextran in pH range 4.5-5.4 compared to pH 5.4 in case of free form. The immobilization process improve the thermal and pH enzyme stability to great extent. The enzyme retains 60% activity after 15 batch reactions 3.2.1.11 dextranase synthesis alpha-dextranase activity reaches 242.8 U/ml when fermentation conditions are 29°C, pH 6.0 and 220 rpm. Addition of glass beads to fermentation medium after 12 h improves enzyme activity by 135.5% 3.2.1.11 dextranase synthesis culturing Chaetomium gracile in medium containing crude dextran and use of high molecular weight of dextrans results in higher dextranase production. Cells incubated in medium containing glucose as sole carbon source exhibit a high growth rate but do not produce dextranase. Using fed-batch and two-step fermentation strategies, production of 159.5 and 187.0 U/ml, respectively, can be achieved 3.2.1.14 chitinase synthesis expression of chitinase in Bacillus thuringiensis under the control of a strong promoter and a 5'-mRNA stabilizing sequence leads to markedly elevated chinolytic activity and a diminution of 10-20% in size of the crystals produced. Strains overexpressing chitinase produce fewer viable spores. No change in protease activity is observed depsite teh overproduction of chitinase 3.2.1.14 chitinase synthesis recombinant enzyme produced in Escherichia coli can efficiently convert colloidal chitin to N-acetyl glucosamine and chitobiose at pH 4.0, 6.0 and 9.0 at 50°C and retains its activity up to 3 days under these conditions 3.2.1.14 chitinase synthesis the thermophilic chitinase may prove useful for industrial applications in chitooligosaccharide production from chitin 3.2.1.17 lysozyme synthesis the extracellular pH-sensitive glycosylation system can be used to obtain bioactive and surface functional neoglycoproteins 3.2.1.17 lysozyme synthesis production of secreted recombinant human lysozyme by use of overexpression vector pPIC3.5k, carrying the strong promoter AOX1 of aldehyde oxidase 1, the HSA signal peptide, the enterokinase recognition motif, and the lysozyme gene. Mature protein is identical with native human lysozyme. It exhibits in vitro bacteriolytic activity against the Gram-positive bacterium Micrococcus lysodeikticus and the Gram-negative bacterium Escherichia coli 3.2.1.20 alpha-glucosidase synthesis the enzyme can be potentially useful for starch hydrolysis as well for the novel synthesis of oligosaccharides in industry 3.2.1.20 alpha-glucosidase synthesis glucose production from maltodextrins employing a thermophilic immobilized cell biocatalyst in a packed-bed reactor, biotransformation of a commercial dextrin mixture at industrial concentrations (30–40%, w/v) into glucose at 75°C, achieving up to 98% conversion 3.2.1.20 alpha-glucosidase synthesis immobilization of enzyme within agar-agar support via entrapment. The maximum immobilization efficiency of 82.77% is achieved using 4.0% agar-agar keeping the diameter of beads up to 3.0 mm. Km value of immobilized enzyme increases from 1.717 to 2.117 mM whereas Vmax decreases from 8,411 to 7,450 U/min as compared to free enzyme. The immobilization significantly increases the stability of maltase against various temperatures and immobilized maltase retains 100% of its original activity after 2 h at 50°C, whereas the free maltase only shows 60% residual activity under the same conditions. Entrapped maltase can be reused for up to 12 cycles and retains 50% of activity even after the 5th cycle. Agar entrapped maltase retains 73% of its initial activity even after 2 months when stored at 30°C 3.2.1.20 alpha-glucosidase synthesis the transglycosylation activity of the mutant glycosynthase produced from AglA can be used to synthesize arbutin alpha-glucosides. The glycosynthase reaction is very specific and produces a single transglycosylation compound. Therefore, it can be used to generate regiospecific glycosidic bonds 3.2.1.20 alpha-glucosidase synthesis the alpha-transglucosidase-producing Geobacillus stearothermophilus as a potential application technique can be successfully used to prepare industrial isomaltooligosaccharides (IMOs) 3.2.1.21 beta-glucosidase synthesis synthesis of glycoconjugates and oligosaccharides 3.2.1.21 beta-glucosidase synthesis increased hydrolysis of cellobiose by immobilized enzyme preparation 3.2.1.21 beta-glucosidase synthesis co-expression of beta-glucosidase and endoglucanase in Saccharomyces cerevisiae. Ethanol fermentation from 20 g per l barley beta-glucan with the co-displaying strain reaches 7.94 g per l ethanol after 24 h of fermentation. The conversion rate of ethanol is 69.6% of the theoretical ethanol concentration 3.2.1.21 beta-glucosidase synthesis construction of a lactic-acid producing Saccharomyces cerevisiae strain expressing isoform Bgl1 on cell surface by fusing the mature protein to the C-terminal half region of alpha-agglutinin. Strain is able to grow on cellobiose and glucose minimal medium at the same rate. The maximum rate of L-lactate production on cellobiose is 2.8 g per l and similar to that on glucose 3.2.1.21 beta-glucosidase synthesis enzyme is able to hydrolyze rice straw into simple sugars 3.2.1.21 beta-glucosidase synthesis optimization of culture condition for production of beta-glucosidase from Aspergillus niger and subsequent use for hydrolysis of ginsenosides. Presence of wheat bran and KH2PO4 and stirring speed have significant effect on enzyme activity 3.2.1.21 beta-glucosidase synthesis use of enzyme for syntheses of pyridoxine glycosides in di-isopropylether. Synthesis of 7-O-(alpha-D-glucopyranosyl)pyridoxine, 7-O-(beta-D-glucopyranosyl)pyridoxine, 6-O-(alpha-D-glucopyranosyl)pyridoxine, 7-O-(alpha-D-galactopyranosyl)pyridoxine, 7-O-(beta-D-galactopyranosyl)pyridoxine, 6-O-(alpha-D-galactopyranosyl)pyridoxine, 7-O-(alpha-D-mannopyranosyl)pyridoxine, 7-O-(beta-D-mannopyranosyl)pyridoxine, 6-O-(alpha-D-mannopyranosyl)pyridoxine in yields ranging from 23% to 40% 3.2.1.21 beta-glucosidase synthesis the enzyme is useful in synthetic biology to produce complex bioactive glycosides and to avoid chemical hazards 3.2.1.21 beta-glucosidase synthesis glucose production from cellulose material using beta-glucosidase from Pyrococcus furioses and endocellulase from Pyrococcus horikoshii. The combination reaction can produce only glucose without the other oligosaccharides from phosphoric acid swollen Avicel 3.2.1.21 beta-glucosidase synthesis preparation of lactose-free pasteurized milk with a recombinant thermostable beta-glucosidase 3.2.1.21 beta-glucosidase synthesis immobilization of enzyme on macroporous resin NKA-9 modified with polyethylenimine and glutaraldehyde. The optimal conditions of immobilized enzyme are the same as that of the free enzyme, the highest activity with cellobiose as the substrate approaches 1.7 U/g. Immobilization improves the thermostability, pH stability and glucose tolerance, the residual activity is 68% of the initial activity at the end of 10 repeated cycles. 2 mM Zn2+ increases the relative activity of the immobilized enzyme to 192% and 199% with cellobiose and 4-nitrophenyl-beta-D-glucopyranoside as substrates, respectively and improves the reusability, high-temperature stability, and glucose tolerance 3.2.1.21 beta-glucosidase synthesis optimized medium composition for beta-glucosidase expression is corn cob (51.8 g/l), beef extract (23.8 g/l), salicin (0.5 g/l), MnSO4·H2O (0.363 g/l), MgSO4·7H2O (0.4 g/l), and NaCl (5 g/l). Under the optimal conditions, the activity of beta-glucosidase is up to 4.71 U/ml 3.2.1.21 beta-glucosidase synthesis the optimal medium composition for beta-glucosidase production is 2.99% (w/v) bagasse, 0.33% (w/v) yeast extract, 0.38% (w/v) Triton X-100, 0.39% (w/v) NaNO3, and pH 8.0 at 30°C. Large-scale production in 7-l stirred tank bioreactor results in beta-glucosidase production of up to 23.29 IU/g within 80 h of incubation 3.2.1.21 beta-glucosidase synthesis the enzyme can constitute a valuable biocatalyst for the synthesis of disaccharides involving beta(1-3) linked disaccharide structures as, for example, Bifidus factors 3.2.1.22 alpha-galactosidase synthesis production of extracellular alpha-galactosidase by solid-state fermentation. Soybean flour is the best solid substrate, and packed-bed bioreactors perform well giving a yield of 197 U/gds, with a forced aeration of 2 vvm. Highest yield is obtained after 96 h of incubation 3.2.1.22 alpha-galactosidase synthesis the mutant is an efficient alpha-galactosynthase producing different galactosylated disaccharides from beta-galactosyl-azide donors and 4-nitrophenyl-alpha-and beta-glycosides as acceptors 3.2.1.23 beta-galactosidase synthesis simple and inexpensive method for synthesizing (2R)-glycerol-O-D-beta-galactopyranoside by utilization of the transgalalactosylating properties of beta-galactosidase and the chloroform solubility of a derivative of (2R)-glycerol-O-D-beta-galactopyranoside that is formed by the transfer of galactose onto isopropylidene glycerol 3.2.1.23 beta-galactosidase synthesis enzyme mutant E184A is a valuable catalyst for the synthesis of metabolically stable analogues of the important glycosidic linkages to the 3 and 4 positions of glucosides and galactosides 3.2.1.23 beta-galactosidase synthesis expression in Escherichia coli under control of araBD promoter. The addition of D-fucose causes an improvement in specific beta-galactosidase production, although beta-galactosidase is produced as an inclusion body. The addition of D-fucose after induction leads to an increase in the specific activity of beta-galactosidase inclusion bodies and causes a changes in the structure of beta-galactosidase inclusion bodies, with higher enzyme activity 3.2.1.23 beta-galactosidase synthesis production of beta-galactosidase by expression of the genes encoding the large and the small subunit in Lactobacillus plantarum WCFS1. Cultivations yield about 23000 U of enzyme per l 3.2.1.23 beta-galactosidase synthesis the enzyme catalyzes the production of the synthetic disaccharide lactulose (4-O-beta-D-galactopyranosyl-D-fructose) via a transgalactosylation using lactose as a galactose donor and fructose as an acceptor. Lactulose is used in treatment of hyperammonemia and as a gentle laxative. It is also applied to commercial infant formulas and various milk products because it specifically promotes the intestinal proliferation of Bifidobacterium, which creates an acid medium that inhibits the growth of undesirable bacteria 3.2.1.23 beta-galactosidase synthesis the F441Y mutant enzyme has potential application in the industrial preparation of galactooligosaccharides 3.2.1.23 beta-galactosidase synthesis immobilization by covalent attachment onto Eupergit C with a binding efficiency of 95%. Immobilization increases both activity and stability at higher pH values and temperature but does not significantly change kinetic parameters for the substrate lactose. The immobilized enzyme shows a strong transgalactosylation reaction, resulting in the formation of galactooligosaccharides. The maximum yield of 34% galactooligosaccharides is obtained when the degree of lactose conversion is roughly 80% 3.2.1.23 beta-galactosidase synthesis fermentation parameters for the maximum production of cold active beta-galactosidase are pH 7.3, 82% (v/v) cheese whey, 3.84% tryptone. An overall 3.6fold increase in cold active beta-galactosidase production (34.37 U/ml) is achieved in optimized medium 3.2.1.23 beta-galactosidase synthesis immobilization and stabilization of beta-galactosidase on Duolite A568 using a combination of physical adsorption, incubation at pH 9.0 and cross-linking with glutaraldehyde leads to a 44% increase in enzymatic activity as compared with a two-step immobilization process (adsorption and cross-linking). The immobilized enzyme presents a good thermal stability at temperatures around 50°C, and very good pH stability in the range from 1.5 to 9.0 3.2.1.23 beta-galactosidase synthesis immobilization of enzyme on aminovinylsulfone. The enzyme is immobilized at moderate ion strength at pH values from 5.0 to 9.0 via ion exchange on aminovinylsulfone support. 50-80% of the initial activity and a stabilization factor of around 8-15 can be obtained 3.2.1.23 beta-galactosidase synthesis immobilization of enzyme on functionalized multi-walled carbon nanotubes. Acid functionalization using H2SO4/HNO3 is the most effivcient method. Enzyme maintains 51% of initial activity after 90 days at 4°C and more than 90% of initial activity up to the 4th recycle 3.2.1.23 beta-galactosidase synthesis Optimal extracellular beta-galactosidase activity in recombinant Escherichia coli is observed when induction is initiated when the optical density at 600 nm reaches 40, when expression is induced at 37°C; and lactose is added at a constant feeding rate of 1.0 g/l/h. The extracellular activity reaches 220.0 U/ml, which represents 65.0% of the total beta-galactosidase activity expressed 3.2.1.23 beta-galactosidase synthesis synthesis of propyl-beta-galactoside using immobilized beta-galactosidase in glyoxyl-agarose. Reaction yield increases twofold with the glyoxyl-agarose derivative, and after ten sequential batches, the efficiency is 115% higher than obtained with the free enzyme. Enzyme immobilization favors product recovery, and avoids browning reactions. Propyl-beta-galactoside can be recovered with a purity above 99% 3.2.1.26 beta-fructofuranosidase synthesis immobilization using anion-exchange resin WA-30 and cross-linking with glutaraldehyde depresses the hydrolysis reaction of isoform F2 3.2.1.26 beta-fructofuranosidase synthesis improved enzyme production by exposure of cells to 0.06 mg/ml N-methyl-N-nitro-N-nitrosoguanidine 0.06 mg/ml for 20 min. The resulting strain NG-5 offers improved extracellular beta-fructofuranidose production of 34 U/ml/min compared to the wild-type strain's 1.15 U/ml/min. A 40fold increase of beta-fructofuranidose activity can be achieved with the process parameters incubation period 48 h, sucrose concentration 5.0 g/l, initial pH of 6.0, inoculum size 2.0% v/v, 16 h old, and urea concentration of 0.2%, w/v 3.2.1.26 beta-fructofuranosidase synthesis immobilization by sodium alginate increases enzyme stability 3.2.1.26 beta-fructofuranosidase synthesis immobilization of invertase on a hydrogel comprised of methacrylic acid and N-vinyl pyrrolidone and ethyleneglycol dimethacrylate, converted to nanogel by an emulsification method and further functionalized by Curtius azide reaction. The values of Vmax, maximum reaction rate, of 0.123 unit/mg, Michaelis constant of 7.429 mol/L and energy of activation of 3.511 kJ/mol for the immobilized invertase are comparable with those of the free invertase at optimum conditions. The covalent immobilization enhances the pH and thermal stability of invertase 3.2.1.26 beta-fructofuranosidase synthesis immobilization of invertase on a porous silicon layer with appropriate catalytic behavior for the sucrose hydrolysis. The procedure is based on support surface chemical oxidation, silanization, activation with glutaraldehyde and finally covalent bonding of the free enzyme to the functionalized surface. Vmax undergoes a substantial increase of about 30% upon immobilization. The value of Km increases by a factor of 1.53 upon immobilization. The initial activity is still preserved up to 28 days while the free enzyme undergoes a 26% loss of activity after the same period 3.2.1.26 beta-fructofuranosidase synthesis immobilization of invertase on polyurethane rigid adhesive foam for application in an enzymatic bioreactor. The kinetic parameters are Km 46.5 mM for immobilized invertase versus 61.2 mM for free invertase. The immobilized invertase derivative maintains 50.1% of initial activity, i.e. 69.17 U/g support, for 8 months. The bioreactor shows the best production of inverted sugar syrup using up-flow rate of 0.48 l/h with average conversion of 10.64%/h at a feeding rate of 104 /h 3.2.1.26 beta-fructofuranosidase synthesis production of high levels of cell extract and extracellular invertases when grown under submerged fermentation and solid-state fermentation, using agroindustrial products or residues as substrates, mainly soy bran and wheat bran, at 40°C for 72 h and 96 h, respectively. Addition of glucose or fructose in submerged fermentation inhibits enzyme production, while the addition of 1% (w/v) peptone as organic nitrogen source enhances the production by 3.7fold. 1% (w/v) (NH4)2HPO4 inhibits enzyme production around 80% 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis a continuous stirred-tank reactor charged with the enzyme and operated at steady-state conditions could be a useful reaction system for the production of galacto-oligosaccharides in which composition is narrower and more easily programmable, in terms of the individual components contained, as compared to the batchwise reaction 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis development of a continuous cellobiose hydrolysis system for glucose production with a high degree of conversion by the enzyme immobilized on chitosan activated with glutaraldehyde 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis enzymatic synthesis of 2-beta-D-galactopyranosyloxyethyl methacrylate starting from 2-hydroxyethyl methacrylate and 4-nitrophenyl-beta-D-galactopyranoside 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis enzymatic synthesis of polyol- and masked polyol-glycosides. The enzyme performs the synthesis of glycosides in a competitive high yield compared with that obtained with mesophilic enzymes. It permits the use of a wide variety of acceptors including tetritol compounds and their masked or protected derivatives. The synthesis of glucosides of 2,3-isopropylidene protected pure enantiomers of threitol shows the possibility of obtaining reasonable yields of products in the presence of a scarce amount of the acceptor by adding aliquots of donor at time intervals, in such a way that a molar excess of acceptor was maintained, thus avoiding undesirable formation of glycosides of the donor or enzymatic hydrolysis of the product 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis formation of mannosides and xylosides by transglycosylation 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis high yield production of hydroxytyrosol from a commercially available oleuropein by using the immobilised recombinant EcSbgly from the hyperthermophilic archaeon Sulfolobus solfataricus on chitosan support 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis industrial production of galactooligosaccharides, very efficient enzyme for the synthesis of beta-galactooligosaccharides because of its high product yields, transfer rates, substrate specificity, and thermostability 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis optimization of hexyl-beta-D-glycoside synthesis from lactose in hexanol at low water activity and high temperature. Compared to other beta-glycosidases in lactose conversion into alkyl glycoside, the enzyme shows high activity in a hexanol one-phase system and synthesized high yields of both hexyl-beta-D-galactoside and hexyl-beta-D-glucoside. Using 32 g/l lactose (93 mM), the enzyme synthesizes yields of 41% galactoside (38.1 mM) and 29% glucoside (27.0 mM) 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis synthesis of 2-deoxyglycosides and 2-deoxygalactosides from glucal or galactal as donors. The yields observed with alkyl acceptors confirms that the robustness of the biocatalyst is of great help in designing practical syntheses of pure beta-anomers of 2-deoxy derivatives of 4-penten-1-ol (obtained in 80% yield at 20 fold molar excess) and 3,4-dimethoxybenzyl alcohol (obtained in 19% yield at 3.3 fold molar excess). The attachment of 2-deoxyglyco units is performed on various pyranosidic acceptors (4-nitrophenyl alpha-D-glucopyranoside, 2-nitrophenyl 2-deoxy-N-acetyl-alpha-D-glucosamine and 4-nitrophenyl 2-deoxy-N-acetyl-beta-D-glucosamine). At low molecular excesses of the acceptors, satisfactory yields (20-40%) of chromophoric 2-deoxy di- and trisaccharides are obtained 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis synthesis of ginsenoside K (20-O-beta-D-glucopyranosyl-20(S)-protopanaxadiol) by a recombinant enzyme. 20-O-beta-D-Glucopyranosyl-20(S)-protopanaxadiol has anti-tumor, anti-inflammatory, anti-allergic and hepatoprotective effects 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis the immobilized enzyme is useful for the production of galactooligosaccharides by using a packed-bed enzyme reactor operated at 70°C 3.2.1.B26 Sulfolobus solfataricus beta-glycosidase synthesis application of the alpha-L-arabinofuranosidase from Caldicellulosiruptor saccharolyticus along with the beta-glycosidase from Sulfolobus solfataricus to yield ginsenoside compound K from the protopanaxadiol-type ginsenosides in red-ginseng extracts. The optimal reaction conditions are pH 6.0, 80°C, 2 U/ml beta-glucosidase, 3 U/ml alpha-L-arabinofuranosidase, and 7.5 g/l ginsenosides in red-ginseng extract. Under these optimized conditions 4.2 g/l ginsenoside compound K from 7.5 g/l protopanaxadiol-type ginsenosides is obtained in 12 h without other ginsenosides, with a molar yield of 100% and a productivity of 348 mg/l/h 3.2.1.28 alpha,alpha-trehalase synthesis expression as a fusion protein with an N-terminal or C-terminal hexahistidine tag in a baculovirus-silkworm expression system. Only N-terminally tagged trehalase shows a high activity 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis a continuous stirred-tank reactor charged with the enzyme and operated at steady-state conditions could be a useful reaction system for the production of galacto-oligosaccharides in which composition is narrower and more easily programmable, in terms of the individual components contained, as compared to the batchwise reaction 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis optimization of hexyl-beta-glycoside synthesis from lactose in hexanol at low water activity and high temperature. Compared to other beta-glycosidases in lactose conversion into alkyl glycoside, the enzyme shows high activity in a hexanol one-phase system and synthesized high yields of both hexyl-beta-galactoside and hexyl-beta-glucoside. The enzyme synthesizes yields of 63% galactoside (58.6mM) and 28% glucoside (26.1 mM) 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis the immobilized enzyme is useful for the production of galactooligosaccharides by using a packed-bed enzyme reactor operated at 70°C 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis beta-glycosidase converts ginsenosides Rb1, Rb2, Rc, and Rd to protopanaxadiol aglycone via compound K. With increases in the enzyme activity, the productivities increase. The substrate concentration is optimal at ginsenoside Rd or 10% (w/v) ginseng root extract. 4 mM of ginsenoside Rd is converted to 3.3 mM compound K with a yield of 82.5% (mol/mol) and a productivity of 2010 mg per l and h at 1 h and is hydrolyzed completely to the aglycone with 364 mg per l and h after 5 h 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis continuous enzymatic process for the production of the prebiotic disaccharide lactulose through transgalactosylation by CelB. CelB is immobilized onto anion-exchange resin Amberlite IRA-93 or onto Eupergit C with immobilization yields of 72% and 83%, respectively, giving specific activities of 920 nkat/g dry carrier and 1500 nkat/g dry carrier at 75°C with 4-nitrophenyl-beta-D-galactopyranoside as substrate. Maximum lactulose yields of 43% related to the initial lactose concentration are reached. The corresponding productivities are 52 g lactulose per l and h using Amberlite IRA-93 and 15 g lactulose per l and h unsing Eupergit C, respectively. While both carrier-bound CelB preparations are 100% stable for at least 14 days, the half-life of the free CelB in the enzyme membrane reactor is only about 1.5 days 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis development of a microstructured immobilized enzyme reactor for production of beta-glucosylglycerol, transglycosylation reaction, under conditions of continuous flow at 70°C. CelB is covalently attached onto coated microchannel walls to give an effective enzyme activity of 30 U per total reactor working volume of 25 ml. Glycerol causes a concentration-dependent decrease in the conversion of the glucosyl donors 2-nitrophenyl beta-D-glucoside and cellobiose via hydrolysis and strongly suppresses participation of the substrate in the reaction as glucosyl acceptor. The yields of beta-glucosylglycerol are about 80% and 60% based on 2-nitrophenyl beta-D-glucoside and cellobiose converted, respectively, and maintain up to near exhaustion of substrate, giving about 120 mM (30 g/l) of beta-glucosylglycerol from the reaction of cellobiose and 1 M glycerol. The structure of the transglucosylation products is 1-O-beta-D-glucopyranosyl-rac-glycerol (79%) and 2-O-beta-D-glucopyranosyl-sn-glycerol (21%) 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis expression of CleB gene in Escherichia coli gives approx 100000 U of enzyme activity/l of culture medium after 8 h of growth 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis expression of enzyme using a baculovirus expression vector system in silkworm, Bombyx mori and purification to about 81% homogeneity in a single heat-treatment step. The expressed beta-glucosidase accounts for more than 10% of silkworm total haemolymph proteins. The expression level reaches 10199.5 U per ml hemolymph and 19797.4 U per silkworm larva, and the specific activity of the one-step purified crude enzyme is 885 U per mg 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis hydrolysis of lactose in UHT skim milk at 70°C using CelB covalently attached onto Eupergit C in yields of 80%, and in a packed-bed immobilized enzyme reactor. The packed-bed reactor is about 10fold more stable and gives about the same productivity at 80% substrate conversion as the hollow-fiber reactor at 60% substrate conversion. The marked difference in the stability of free and immobilized CelB seems to reflect mainly binding of the soluble enzyme to the membrane surface of the hollow-fiber module. Microbial contamination of the reactors did not occur during reaction times of up to 39 d 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis improvement of the stability of recombinant CelB by entrapment into Escherichia coli cells under conditions promoting strong inactivation. Glutardialdehyde-mediated protein cross-linking or rigidification of the cell membrane by adding magnesium ions is required to prevent release of CelB from within the cell into the bulk solution. In the presence of 1M glucose or when applying recirculation rates of 2.6per min, the entrapped enzyme is around 2fold more stable at 80°C than free CelB. The significance of the stabilisation is attenuated by the decrease in CelB initial activity which wis due to cross-linking and glutardialdehyde concentration-dependent 3.2.1.B28 Pyrococcus furiosus beta-glycosidase synthesis use of CelB for oligosaccharide production from lactose in a kinetically controlled reaction. At reaction temperatures of 80°C and higher, the inactivation rate of the enzyme in the presence of sugars is increased by a factor of 2, caused by the occurrence of Maillard reactions between the sugar and the enzyme 3.2.1.31 beta-glucuronidase synthesis the enzyme appears suitable for use in enzymatic oligosaccharide synthesis in either the transglycosylation mode or by use of glycosynthase and thioglycoligase approaches 3.2.1.31 beta-glucuronidase synthesis mutant enzyme A365H/R563E has great potential in the industrial production of glycyrrhetinic acid 3-O-mono-beta-D-glucuronide 3.2.1.32 endo-1,3-beta-xylanase synthesis the enzyme may be an essential component for the preparation of protoplasts from some groups of seaweed 3.2.1.32 endo-1,3-beta-xylanase synthesis preparation of a large number of protoplasts are isolated from Porphyra yezoensis, Phorphyra tenera, and Bangia atropurpurea 3.2.1.32 endo-1,3-beta-xylanase synthesis production of D-ylulose from beta-1,3-xylan, of the killer alga Caulerpa taxifolia. The synergistic action of beta-1,3-xylanase TxyA and beta-1,3-xylosidase XloA from Vibrio sp. strain XY-214 enables efficient saccharification of beta-1,3-xylan to D-xylose. D-Xylose is then converted to D-xylulose by using XylA 3.2.1.33 amylo-alpha-1,6-glucosidase synthesis use of enzyme for industrial production of cycloamylose 3.2.1.B33 Sulfolobus shibatae beta-glycosidase synthesis synthesis of beta-glucosylglycerol and its derivatives through the transglycosylation by Sulfolobus shibatae beta-glycosidase SSG and Deinococcus geothermalis amylosucrase DGAS. SSG catalyzes a transglycosylation reaction with glycerol as an acceptor and cellobiose as a donor to produce 56% of beta-D-glucopyranosyl-(1->1/3)-D-glycerol and beta-D-glucopyranosyl-(1->4)-D-glycerol. As a result, 61% of alpha-D-glucopyranosyl-(1->4)-beta-D-glucopyranosyl-(1->1/3)-D-glycerol and 28% of alpha-D-maltopyranosyl-(1->4)-beta-D-glucopyranosyl-(1->1/3)-D-glycerol are synthesized as unnatural glucosylglycerols 3.2.1.B34 Sulfolobus acidocaldarius beta-glycosidase synthesis enzyme is a potential producer of the rare ginsenosides compound K, compound Y, and compound Mc from the major ginsenosides Rb1, Rb2, Rc, and Rd 3.2.1.B34 Sulfolobus acidocaldarius beta-glycosidase synthesis use of alpha-L-arabinofuranosidase from Caldicellulosiruptor saccharolyticus, EC 3.2.1.55, along with beta-glycosidase from Sulfolobus solfataricus to produce ginsenoside compound K from the protopanaxadiol-type ginsenosides in red-ginseng extract. The optimal reaction conditions are as follows: pH 6.0, 80°C, 2 U/ml Sulfolobus solfataricus enzyme, 3 U/ml Caldicellulosiruptor saccharolyticus enzyme, and 7.5 g/l protopanaxadiol-type ginsenosides. The enzymes produce 4.2 g/l ginsenoside compound K from 7.5 g/l ginsenosides in 12 h without other ginsenosides, with a molar yield of 100% and a productivity of 348 mg/l/h 3.2.1.37 xylan 1,4-beta-xylosidase synthesis synthesis of a group of uncommon xylosides via the transxylosylation activity 3.2.1.37 xylan 1,4-beta-xylosidase synthesis immobilization of enzyme on chitosan as best support material gives immobilization and activity yields of 94% and 87%, respectively, of initial activity, and also provides the highest stability, retaining 94% of its initial activity even after being recycled 25times. Maximal activity of immobilized enzyme is achieved at pH 8.0 and 53°C, whereas that for the free enzyme is obtained at pH 7.0 and 50°C. The immobilized enzyme is more thermostable than the free beta-xylosidase. Km values of the free enzyme increases from 2.37 mM to 3.42 mM at the immobilized state. Immobilized enzyme catalyzes the reverse hydrolysis reaction, forming xylooligosaccharides in the presence of a high concentration of xylose. Co-immobilized of beta-xylosidase and xylanase on chitosan leads to a continuous hydrolysis of 3% oat spelt xylan at 50°C and better hydrolysis yields and higher amount of xylose are obtained 3.2.1.37 xylan 1,4-beta-xylosidase synthesis the enzyme immobilized by entrapment into alginate can be used for a continuous production of xylose from xylooligosaccharides at high temperature 3.2.1.37 xylan 1,4-beta-xylosidase synthesis formation of xylooligosaccharides from alpha-D-xylopyranosyl fluoride in a conjugated reaction between beta-xylosidase E335G mutant enzyme and endo-1,4-beta-xylanase E265G mutant enzyme 3.2.1.37 xylan 1,4-beta-xylosidase synthesis upon codon optimization for expression in Pichia pastoris, the expression level increases to 5.7 g/l in a fermenter system 3.2.1.38 beta-D-fucosidase synthesis synthesis of fucosylsugars using the transglycosylation activity 3.2.1.39 glucan endo-1,3-beta-D-glucosidase synthesis heterologous 1,3-beta-glucanase production in Escherichia coli is favoured with moderate culture aeration (0.7-0.9 vvm) and moderate stirring (125–150 rev/min) 3.2.1.39 glucan endo-1,3-beta-D-glucosidase synthesis the enzyme in Schizophyllum commune strain GIM5-43 is used for partial schizophyllan cleavage to achieve a moderate molecular weight and better solubility of schizophyllan 3.2.1.40 alpha-L-rhamnosidase synthesis synthesis of aglycones from glycosides 3.2.1.40 alpha-L-rhamnosidase synthesis immobilization of recombinant enzyme on Ca2+ alginate beads. Immobilization enables its reutilization up to 9 hydrolysis batches without an appreciable loss in activity 3.2.1.40 alpha-L-rhamnosidase synthesis a wild-type alpha-L-rhamnosidase from Alternaria sp. L1 can synthesize rhamnose-containing chemicals through reverse hydrolysis reaction with inexpensive rhamnose as glycosyl donor 3.2.1.40 alpha-L-rhamnosidase synthesis adding sorbitol has great potential to promote enzymatic conversion of rutin to isoquercitrin production. Isoquercitrin has several biological activities, including anti-mutagenesis, anti-virus, anti-hypertensive, anti-proliferative effects, lipid peroxidation, oxidative-stress protection as well as other pharmacological effects 3.2.1.B40 Pyrococcus horikoshii beta-glycosidase synthesis in the presence of sodium formate buffer pH 4.0 at 75°C the E324G mutant acts as a hyperthermophilic glycosynthase. Though the yield of the reaction does not exceed 10%, it is demonstrated that this could be a general strategy for the preparation of hyperthermophilic glycosynthase. The peculiar specificity of the enzyme for alkyl-glycosides makes the resulting glycosynthase a promising tool for biocatalysis 3.2.1.41 pullulanase synthesis use of enzyme for synthesis of novel heterobranche beta-cyclodextrins 3.2.1.41 pullulanase synthesis preparation of maltotriose by hydrolyzing of pullulan with pullulanase, optimum conditions are: time 6 h, pH 5.0, temperature 45°C, amount of pullulanase 10 ASPU/g, concentration of pullulan 3% w/v, method, overview 3.2.1.41 pullulanase synthesis recombinant expression of PulB in Bacillus subtilis is increased by stronger constitutive promoter P43 and use of strain WB600 instread of WB800. Extracellular pullulanase activity reaches up to 24.5 U/ml 3.2.1.B44 glycyrrhizin glucuronohydrolase (glycyrrhetinic acid 3-O-beta-D-glucuronide-forming) synthesis the enzyme is potentially a powerful biocatalyst for environmentally friendly and cost-effective production of glycyrrhetinic acid 3-O-mono-beta-D-glucuronide 3.2.1.51 alpha-L-fucosidase synthesis the transfucosylation ability of one of the fucosidase isoform might be of interest for the synthesis of fucosides 3.2.1.54 cyclomaltodextrinase synthesis CDase I-5 is applied to modify the starch structure to produce low-amylose starch products by incubating rice starch with this enzyme. The amylose content of rice starch decreases from 28.5 to 9% while the amylopectin content remains almost constant with no significant change in side chain length distribution 3.2.1.54 cyclomaltodextrinase synthesis the G415E mutant is an excellent candidate for the industrial production of specific-length maltooligosaccharides from cyclodextrins 3.2.1.55 non-reducing end alpha-L-arabinofuranosidase synthesis application of the recombinant enzyme in the production of xylobiose 3.2.1.55 non-reducing end alpha-L-arabinofuranosidase synthesis alpha-L-arabinanases are essential glycosyl hydrolases participating in the complete hydrolysis of hemicellulose, a natural resource for various industrial processes, such as bioethanol or pharmaceuticals production 3.2.1.55 non-reducing end alpha-L-arabinofuranosidase synthesis the enzyme might be useful for enzymatic synthesis of galactosylfuranoside-containing pharmacophores to be used in mammals 3.2.1.55 non-reducing end alpha-L-arabinofuranosidase synthesis the thermostable enzyme is used for production of ginsenoside Rd from ginsenoside Rc, optimal reaction conditions are pH 5.5, 80°C, 227 U enzyme/ml, and 8.0 g/l ginsenoside Rc in the presence of 30% v/v n-hexane 3.2.1.55 non-reducing end alpha-L-arabinofuranosidase synthesis use of alpha-L-arabinofuranosidase from Caldicellulosiruptor saccharolyticus along with beta-glycosidase from Sulfolobus solfataricus to produce ginsenoside compound K from the protopanaxadiol-type ginsenosides in red-ginseng extract. The optimal reaction conditions are as follows: pH 6.0, 80°C, 2 U/ml Sulfolobus solfataricus enzyme, 3 U/ml Caldicellulosiruptor saccharolyticus enzyme, and 7.5 g/l protopanaxadiol-type ginsenosides. The enzymes produce 4.2 g/l ginsenoside compound K from 7.5 g/l ginsenosides in 12 h without other ginsenosides, with a molar yield of 100% and a productivity of 348 mg/l/h 3.2.1.61 mycodextranase synthesis the enzyme seems to be a useful tool for the preparation of expensive nigerose and nigerooligosaccharides 3.2.1.65 levanase synthesis protection from aging 3.2.1.65 levanase synthesis paper industry 3.2.1.65 levanase synthesis synthesis of various products such as ethanol or aceton-butanol 3.2.1.65 levanase synthesis commercial production of ultra-high-fructose syrups 3.2.1.65 levanase synthesis the enzyme is used for simultaneous saccharification and fermentation of inulin to 2,3-butanediol. A fed-batch simultaneous saccharification and fermentation yields 103.0 g/liter 2,3-butanediol in 30 h, with a high productivity of 3.4 g/liter/h 3.2.1.65 levanase synthesis the enzyme is uzilized for production of beta2-6 fructose oligosaccharides (levan-type FOS) through a sequential reactionwith levan produced from sucrose by bacterial levansucrases, method development, overview 3.2.1.67 galacturonan 1,4-alpha-galacturonidase synthesis pectic enzymes used to generate protoplasts in cell cultures 3.2.1.67 galacturonan 1,4-alpha-galacturonidase synthesis pectic enzymes industrially important in processing of agricultural products, production of pectinolytic enzymes, specifical enzyme for modification of pectins 3.2.1.67 galacturonan 1,4-alpha-galacturonidase synthesis preparing methylated pectins 3.2.1.67 galacturonan 1,4-alpha-galacturonidase synthesis as adjunct to cellulases and hemicellulases in cellulosic biomass treatment 3.2.1.67 galacturonan 1,4-alpha-galacturonidase synthesis mutant strain M3 can be used for production of exopolygalacturonase 3.2.1.68 isoamylase synthesis the enzyme is used for debranching of amylomaize in the production of cycloamylose, natural amylopectin containing starch with enhanced conversion yield after debranching, use of Thermus aquaticus 4-alpha-glucanotransferase for conversion of debranched amylomaize amylose and amylomaize amylopectin into cycloamylose, overview 3.2.1.73 licheninase synthesis the unusually resistance against inactivation by heat, ethanol or ionic detergents makes the enzyme highly suitable for industrial application in the mashing process of beer brewing 3.2.1.73 licheninase synthesis construction of a fusion gene, encoding beta-1,3-1,4-glucanase both from Bacillus amyloliquefaciens and Clostridium thermocellum, via end-to-end fusion and expression in Escherichia coli. The catalytic efficiency of the fusion enzyme for oat beta-glucan is 2.7- and 20fold higher than that of the parental Bacillus amyloliquefaciens and Clostridium thermocellum enzymes, respectively, and the fusion enzyme can retain more than 50% of activity following incubation at 80°C for 30 min, whereas the residual activities of Bacillus amyloliquefaciens and Clostridium thermocellum enzymes are both less than 30% 3.2.1.73 licheninase synthesis over-expression in Pichia pastoris, with a yield of about 1000 U/ml in a 3.7 l fermentor 3.2.1.73 licheninase synthesis upon expression in Pichia pastoirs as active extracellular beta-1,3-1,4-glucanase, the recombinant protein is secreted predominantly into the medium and comprises up to 85% of the total extracellular proteins and reaches a protein concentration of 9.1 g/l with an activity of 55,300 U/ml in 5-l fermentor culture 3.2.1.73 licheninase synthesis analysis of fermentation conditions for beta-1,3-1,4-glucanase production under solid-state fermentation. Under the optimized fermentation conditions, viz. oatmeal as sole carbon source, 5% (w/w) peptone as sole nitrogen source, initial moisture of 80% (w/w), initial culture pH of 5.0, incubation temperature of 50°C and incubation time of 6 days, the highest beta-1,3-1,4-glucanase activity of 20025 U/g dry substrate is achieved. The addition of the purified beta-1,3-1,4-glucanase in mash obviously reduces its filtration time (24.6%) and viscosity (2.61%) 3.2.1.73 licheninase synthesis expression of mutant K20S/N31C/S40E/S43E/E46P/P102C/K117S/N125C/K165S/T187C/H205P in Bacillus subtilis to maximal extracellular activity of 4840.4 U/ml 3.2.1.73 licheninase synthesis immobilization of enzyme on porous silica using glutaraldehyde. Enzyme activity decreases sharply at high concentrations of glutaraldehyde. Immobilized protein is stable over a wide range of pH and can be stored long term at 4°C. After 10 cycles, the enzyme retains 42% of its initial catalytic activity 3.2.1.73 licheninase synthesis the enzyme can be used in the production of anti-hypercholesterolemic agents 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis production of enzyme by expression in Aspergillus niger under control of the Aspergillus niger glyceraldehyde-3-phosphate dehydrogenase promoter gpdP and the Aspergillus awamori glucoamylase terminator glaAT. The glucose concentration and the organic nitrogen source have an effect on both the volumetric enzyme activity and the specific enzyme activity. The highest mannanase activity levels of 16596 nkat ml-1 and 574 nkat mg-1 dcw are obtained for Aspergillus niger when cultivated in a process-viable medium containing corn steep liquor as the organic nitrogen source and high glucose concentrations 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis expression in Yarrowia lipolytica using beta-mannosidase's own secretion signal. Fed batch fermentations result in a 3.9fold increase in volumetric enzyme activity compared with batch fermentation, and a maximum titre of 26,139 nkat/ml 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis production of beta-mannanase using palm kernel cake as substrate in solid substrate fermentation. Optimal conditions are incubation temperature of 32°C, initial moisture level of 59% and aeration rate of 0.5 l/min, resulting in a beta-mannanase yield of 2231.26 U/g 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis development of a fed-batch strategy for engineered enzyme, using high cell-density fermentation. Mannanase activity reaches 5069 U/ml after cultivation for 56 h in 50 l fermenter 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis expression in Escherichia coli based on T7 RNA polymerase promoter and tac promoter systems. Both Escherichia coli OmpA signal peptide and native Bacillus signal peptide can be used efficiently for secretion of recombinant protein. Enzyme can be harvested from whole cell lysate, periplasmic extract or culture broth 4-20 h after induction by IPTG 3.2.1.78 mannan endo-1,4-beta-mannosidase synthesis production of beta-mannanase from palm kernel cake as a substrate in solid substrate fermentation. A statistical model suggests that the optimal conditions for attaining the highest level of beta-mannanase are incubation temperature of 32°C, initial moisture level of 59% and aeration rate of 0.5 l/min. A beta-mannanase yield of 2231.26 U/g is obtained under these optimal conditions 3.2.1.80 fructan beta-fructosidase synthesis microbial production of high fructose syrup, alcohol, acetone and butanol 3.2.1.80 fructan beta-fructosidase synthesis the enzyme preparation is used to hydrolyze pure inulin and raw inulin from Asparagus racemosus for the preparation of a high-fructose syrup 3.2.1.80 fructan beta-fructosidase synthesis expression of bacterial levanase in yeast enables simultaneous saccharification and fermentation of grass juice to bioethanol 3.2.1.81 beta-agarase synthesis use of enzyme for preparation of neoagarohexaose and neoagarotetraose and separation of neoagaro-oligosaccharides by consecutive column chromatography 3.2.1.91 cellulose 1,4-beta-cellobiosidase (non-reducing end) synthesis expression in Pichia pastoris after codon optimization and using the strong methanol-inducible promoter AOX1. 5.84 U CBH II per ml can be obtained at 96 h 3.2.1.91 cellulose 1,4-beta-cellobiosidase (non-reducing end) synthesis expression of enzyme in Escherichia coli and Thermotoga sp. after fusion to the signal peptides of TM1840 (amyA) or TM0070 (xynB). Expressed in Escherichia coli and Thermotoga sp. renders the hosts with increased endo- and exoglucanase activities. In Escherichia coli, the recombinant enzymes are mainly bound to the bacterial cells, whereas in Thermotoga sp., about half of the enzyme activities are observed in the culture supernatants. However, the cellulase activities are lost in Thermotoga sp. after three consecutive transfers 3.2.1.91 cellulose 1,4-beta-cellobiosidase (non-reducing end) synthesis heterologous expression in Bacillus subtilis combined with customized signal peptides for secretion from a random libraries with 173 different signal peptides originating from the Bacillus subtilis genome. The customized signal peptide might influence substrate specificity by affecting the local structure of the CelK-specific N-terminal region containing an immunoglobulin-like domain 3.2.1.91 cellulose 1,4-beta-cellobiosidase (non-reducing end) synthesis SCHEMA structure-guided recombination of fungal class II cellobiohydrolases (CBH II cellulases) from Humicola insolens, Hypocrea jecorina and Chaetomium thermophiulum and mathematical modeling yields a collection of highly thermostable CBH II chimeras with more activity than Humicola insolens CBH II after incubation at 63 °C. The total of 15 validated thermostable CBH II enzymes have high sequence diversity, differing from their closest natural homologs at up to 63 amino acid positions. Selected purified thermostable chimeras hydrolyze phosphoric acid swollen cellulose at temperatures 7 to 15°C higher than the parent enzymes. These chimeras also hydrolyze as much or more cellulose than the parent CBH II enzymes in long-time cellulose hydrolysis assays and have pH/activity profiles as broad, or broader than, the parent enzymes. The best chimera with buildung blocks from all three organisms exhibits both relatively high specific activity and high thermostability 3.2.1.91 cellulose 1,4-beta-cellobiosidase (non-reducing end) synthesis SCHEMA structure-guided recombination of fungal class II cellobiohydrolases (CBH II cellulases) from Humicola insolens, Hypocrea jecorina and Chaetomium thermophiulum and mathematical modeling yields a collection of highly thermostable CBH II chimeras. The total of 15 validated thermostable CBH II enzymes have high sequence diversity, differing from their closest natural homologs at up to 63 amino acid positions. Selected purified thermostable chimeras hydrolyze phosphoric acid swollen cellulose at temperatures 7 to 15°C higher than the parent enzymes. These chimeras also hydrolyze as much or more cellulose than the parent CBH II enzymes in long-time cellulose hydrolysis assays and have pH/activity profiles as broad, or broader than, the parent enzymes. The best chimera with buildung blocks from all three organisms exhibits both relatively high specific activity and high thermostability 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis fluorescence-based assay for the transglycosylation activity of endo-beta-N-acetylglucosaminidases, highly sensitive, easy and quantitative method for screening endo-beta-N-acetylglucosaminidases with transglycosylation activity useful for glycoconjugate synthesis 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis highly efficient chemoenzymatic synthesis of N-glycopeptides. The use of the synthetic oligosaccharide oxazolines as the donor substrates for the transglycosylation expands the substrate availability and results in substantial enhancement of the synthetic efficiency 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis construction of natural or selectively modified glycopeptides by endoglycosidase-catalyzed transglycosylation 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis synthesis of oxazoline mono-, di-, tri- and hexasaccharides as potential glycosyl donors for enzyme catalyzed glycosylation of glycopeptides and glycoprotein remodelling 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis use of enzyme for synthesis of a human immunodeficiency virus type 1 glycopeptide with potent anti-HIV activity 3.2.1.96 mannosyl-glycoprotein endo-beta-N-acetylglucosaminidase synthesis use of enzyme for the in vitro synthesis of glycoproteins containing complex type oligosaccharides from glycoproteins produced by yeast. Transglycosylation activity of enzyme can change high-mannose type oligosaccharides on glycoproteins to complex type ones 3.2.1.97 endo-alpha-N-acetylgalactosaminidase synthesis production of Galbeta(1-3)GalNAc from asialofetuin 3.2.1.97 endo-alpha-N-acetylgalactosaminidase synthesis synthesis of a wide variety of O-linked glycopeptides 3.2.1.98 glucan 1,4-alpha-maltohexaosidase synthesis very advantageous for obtaining pure maltohexaose 3.2.1.98 glucan 1,4-alpha-maltohexaosidase synthesis continuous production of maltohexaose on large scale using the immobilized exomaltohexaohydrolase 3.2.1.98 glucan 1,4-alpha-maltohexaosidase synthesis enzymatic reaction, transglycosylation, provides practical technique for industrial production of p-nitrophenyl alpha-maltoheptaoside, useful substrate for assay of human alpha-amylase in serum and urine 3.2.1.99 arabinan endo-1,5-alpha-L-arabinanase synthesis applied to the refinement of cotton fiber, enzyme is able to release the cotton fiber coating, yielding product of high quality but with lower amounts of wastes 3.2.1.130 glycoprotein endo-alpha-1,2-mannosidase synthesis the mutant has excellent transglycosylation activity and extremely low hydrolytic activity. The minimum motif required for glycosyl acceptor is Man-alpha-(1->2)Man. The synthetic utility of the enzyme is demonstrated by generation of a high-mannose-type undecasaccharide (Glc1Man9GlcNAc2) 3.2.1.131 xylan alpha-1,2-glucuronosidase synthesis application in enzymatic production of xylooligosaccharides from xylan. The high level of the thermostable alpha-glucuronidase from Thermotoga maritima, combined purification by a simple heat treament, has a considerable potential in the production of xylooligosaccharide, especially xylobiose 3.2.1.132 chitosanase synthesis valuable enzyme for the commercial production of chitosan oligosaccharides and other chitosan hydrolysates 3.2.1.132 chitosanase synthesis enhancement of enzyme production from 3.6 U/ml to 118 U/ml by substrate induction, statistical optimization of medium composition and culture conditions with colloidal chitosan being the best inducer and carbon source for chitosanase production 3.2.1.132 chitosanase synthesis enzyme is able to catalyze the synthesis of small amounts of chitooctaose from a mixture of chitobiose to chitoheptaose oligomers, possible through transglycosylation. Carrying out this process in reversed micellar microreactors formed by sodium bis-2(ethylhexyl) sulfosuccinate in isooctane significantly increases formation of high degree polymerized chitooligosaccharides. Pentamer and hexamer oligosaccharides are the main glycosyl acceptors 3.2.1.132 chitosanase synthesis expression in yeast cells as a whole-cell biocatalyst. Protein is localized to cell surface 3.2.1.132 chitosanase synthesis optimization of production conditions. In the optimized medium, strain JG produces 0.8 mmol per min and l of enzymic activity in 72 h 3.2.1.132 chitosanase synthesis chitosanases can be used to produce partially acetylated chitosan oligosaccharides for different applications 3.2.1.132 chitosanase synthesis the enzyme can be a competitive candidate for chitosan oligosaccharide-manufacturing industry 3.2.1.132 chitosanase synthesis the enzyme is a good candidate for production of beta-D-GlcN-(1->4)-beta-D-GlcN 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis thermostable maltogenic amylase with industrial potential, suitable for producing high maltose syrups from liquefied starch 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis industrial processes use heat-stable alpha-amylase for degrading starch 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis heterologous protein expression in Escherichia coli may contribute to better industrial production of maltogenic amylase 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis formation of maltosyl-tagatose from D-tagatose and maltotriose with maltogenic amylase. Glucosyl-tagatose is produced from maltosyl-tagatose by removal of a glucosyl moiety by glucoamylase. Glucosyl-tagatose has potential as a low-calorie sweetener and cryostabilizer 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis production of branched maltooligosaccharide 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis production of highly branched amylopectin and amylose from enzymatically modified rice starch 3.2.1.133 glucan 1,4-alpha-maltohydrolase synthesis the purified enzyme is employed to catalyze genistin glycosylation using gamma-cyclodextrin as both glucosyl donor and solubilizer 3.2.1.135 neopullulanase synthesis panose migth be used as an anticariogenic sweetener in foods, development of a system for continuous production of extremely high panose syrup from pullulan by employing the enzyme 3.2.1.139 alpha-glucuronidase synthesis hydrolysis of amylouronate to glucuronate by AUH-I 3.2.1.140 lacto-N-biosidase synthesis synthesis of Galbeta(1-3)GlcNAcbeta(1-3)Galbeta(1-4)Glc, i.e. lacto-neotetraose 3.2.1.141 4-alpha-D-{(1->4)-alpha-D-glucano}trehalose trehalohydrolase synthesis production of trehalose from starch 3.2.1.141 4-alpha-D-{(1->4)-alpha-D-glucano}trehalose trehalohydrolase synthesis production of trehalose from starch. Trehalose is utilized as a stabilizer for dried or frozen food, in cosmetics and in medicines as a drug additive 3.2.1.141 4-alpha-D-{(1->4)-alpha-D-glucano}trehalose trehalohydrolase synthesis conditions for the production of trehalose from starch by thermostable maltooligosyl trehalose synthase and maltooligosyl trehalose trehalohydrolase from Sulfolubus acidocaldarius DSM 639 3.2.1.141 4-alpha-D-{(1->4)-alpha-D-glucano}trehalose trehalohydrolase synthesis trehalose production from starch 3.2.1.142 limit dextrinase synthesis expression of the limit dextrinase encoding gene fragment without signal peptide, with the Saccharomyces cerevisiae alpha-factor secretion signal under control of the alcohol oxidase 1 promoter, in Pichia pastoris leads to highly active barley limit dextrinase secreted during high cell-density fermentation. Optimization of a fedbatch fermentation procedure enables efficient production in a 5-l bioreactor, yielding 34 mg homogenous enzyme with 84% recovery 3.2.1.151 xyloglucan-specific endo-beta-1,4-glucanase synthesis mutant engineered xyloglucanases acting as glycosynthases are emerging as useful tools for the synthesis of large, complex polysaccharides, method development for robust and versatile method for the preparative synthesis of homogeneous xyloglucans with regular substitution patterns not available in nature, overview 3.2.1.152 mannosylglycoprotein endo-beta-mannosidase synthesis possible application in the synthesis of oligosaccharides containing mannosyl-beta-1,4-structures 3.2.1.155 xyloglucan-specific exo-beta-1,4-glucanase synthesis both the cane molasses medium and lactose-based conventional medium can serve as excellent growth media for Trichoderma reesei. The most abundant cellulolytic enzymes identified in both media are cellobiohydrolases (Cel7A/Cel6A) and endoglucanases (Cel7A/Cel5A) and are more abundant in CMM. Both media can serve as an inducer of xylanolytic enzymes. The main xylanases (XYNI/XYNIV) and xyloglucanase (Cel74A) are found at higher concentrations in the the cane molasses medium than lactose-based conventional medium 3.2.1.157 iota-carrageenase synthesis the enzyme can be utilized as a potential biocatalyst for producing iota-carrageenan oligosaccharides with different polymerization degrees 3.2.1.163 1,6-alpha-D-mannosidase synthesis regioselective synthesis of mannobiose and mannotriose by reverse hydrolysis using the 1,6-alpha-D-mannosidase from Aspergillus phoenicis, method optimization, overview 3.2.1.163 1,6-alpha-D-mannosidase synthesis synthesis of FimH receptor-active manno-oligosaccharides by reverse hydrolysis using alpha-mannosidases from Penicillium citrinum, Aspergillus phoenicis and almond in a sequential reaction process, method development and optimization, overview 3.2.1.163 1,6-alpha-D-mannosidase synthesis the organism is employed in regioselective synthesis of manno-oligosaccharides involving the enzyme 3.2.1.165 exo-1,4-beta-D-glucosaminidase synthesis efficient tool for industrial production of glucosamine monosaccharide 3.2.1.165 exo-1,4-beta-D-glucosaminidase synthesis enzymatic formation of chitooligosaccharides by transglycosylation 3.2.1.165 exo-1,4-beta-D-glucosaminidase synthesis production of N-acetylglucosamine from chitosan by enzymatic degradation 3.2.1.167 baicalin-beta-D-glucuronidase synthesis synthesis of baicalein (i.e. 5,6,7-trihydroxy-2-phenyl-4H-chromen-4-one), a main active ingredient of Scutellaria sp. used in traditional Chinese medicine. It is difficult to obtain baicalein directly from skullaps because of its low content 3.2.1.167 baicalin-beta-D-glucuronidase synthesis the enzyme can be used for biotransformation of glycone baicalin to more pharmacologically active aglycone baicalein in extracts from Scutellaria baicalensis Georgi plant roots. Development of a chemically defined medium-based baicalein bioproduction process with a comparable yield compared to complex medium-based ones, overview 3.2.1.176 cellulose 1,4-beta-cellobiosidase (reducing end) synthesis heterologous expression in Bacillus subtilis combined with customized signal peptides for secretion from a random libraries with 173 different signal peptides originating from the Bacillus subtilis genome. The customized signal peptide does not affect enzyme performance when assayed on carboxymethyl cellulose, phosphoric acid swollen cellulose, and microcrystalline cellulose 3.2.1.176 cellulose 1,4-beta-cellobiosidase (reducing end) synthesis recombinant enzyme expressed in Zea mays is glycosylated and 6 kDa smaller than the native fungal protein. The cellobiohydrolase performs as well as or better than its fungal counterpart in releasing sugars from complex substrates such as pretreated corn stover or wood 3.2.1.177 alpha-D-xyloside xylohydrolase synthesis utility of enzyme AxlA from Aspergillus niger in supplementation of CTec2/HTec2 from Trichoderma reesei for enhancing release of free Glc and Xyl in combination with commercial enzyme cocktails from dicotyledonous and monocotyledonous plants, overview. AxlA supplementation also improves Glc yields from corn stover treated with the commercial cellulase Accellerase 1000 3.2.1.191 ginsenosidase type III synthesis the beta-galactosidase from Aspergillus sp. can be useful for the mass production of rare ginsenosides 3.2.1.191 ginsenosidase type III synthesis the enzyme is applied for the production of gypenoside LXXV from gypenoside XVII at the gram-scale. The pure GypLXXV can be used for the study of anti-cancer effects against three kinds of cancer cells in vitro 3.2.1.192 ginsenoside Rb1 beta-glucosidase synthesis industrial applications of Lactobacillus rhamnosus strain GG to the biocatalysis of ginsenosides and/or other phytochemicals 3.2.1.192 ginsenoside Rb1 beta-glucosidase synthesis the beta-galactosidase from Aspergillus sp. can be useful for the mass production of rare ginsenosides 3.2.1.192 ginsenoside Rb1 beta-glucosidase synthesis the beta-glucosidase activity of Paenibacillus sp. MBT213 strain may be utilized in development of variety of health foods, dairy foods and pharmaceutical products 3.2.1.193 ginsenosidase type I synthesis in pharmaceutical and commercial industries, this recombinant Bgy2 can be suitable for producting ginsenoside Rd and compound K 3.2.1.193 ginsenosidase type I synthesis production of the pharmacologically active minor ginsenoside F2 from the major ginsenosides Rb1 and Rd by using the recombinant Lactococcus lactis strain expressing heterologous the beta-glucosidase gene 3.2.1.194 ginsenosidase type IV synthesis the enzyme is applied for the production of gypenoside LXXV from gypenoside XVII at the gram-scale. The pure GypLXXV can be used for the study of anti-cancer effects against three kinds of cancer cells in vitro 3.2.1.195 20-O-multi-glycoside ginsenosidase synthesis in a reaction at 85°C and pH 5.0, 25 g/l of ginsenoside Rc is transformed into 21.8 g/l of ginsenoside Rd within 60 min, with a corresponding molar conversion of 99.4% and a high ginsenoside Rd productivity of 21800 mg/l/h 3.2.1.206 oleuropein beta-glucosidase synthesis high yield production of hydroxytyrosol from a commercially available oleuropein by using the immobilised recombinant EcSbgly from the hyperthermophilic archaeon Sulfolobus solfataricus on chitosan support 3.2.1.211 endo-(1->3)-fucoidanase synthesis the enzyme can be used for the manufacture of biologically active fucooligosaccharides from the fucoidans of Chorda filum 3.2.1.211 endo-(1->3)-fucoidanase synthesis the enzyme can be used for the manufacture of biologically active fucooligosaccharides from the fucoidans of Fucus evanescens 3.2.1.212 endo-(1->4)-fucoidanase synthesis the enzyme can be used for the modification of natural fucoidans to obtain more regular and easier characterized derivatives useful for research and practical applications 3.2.2.4 AMP nucleosidase synthesis stabilization of the adenylate energy charge 3.2.2.4 AMP nucleosidase synthesis purine nucleotide synthesis in procaryotes 3.3.2.1 isochorismatase synthesis synthesis of homochiral cis-cyclohexa-3,5-diene-1,2-diols 3.3.2.8 limonene-1,2-epoxide hydrolase synthesis application of directed evolution using iterative saturation mutagenesis as a means to engineer LEH mutants showing broad substrate scope with high stereoselectivity. Mutants are obtained which catalyze the desymmetrization of cyclopentene-oxide with stereoselective formation of either the (R,R)- or the (S,S)-diol on an optional basis. The mutants prove to be excellent catalysts for the desymmetrization of other meso-epoxides and for the hydrolytic kinetic resolution of racemic substrates 3.3.2.9 microsomal epoxide hydrolase synthesis the enantioselective enzyme is useful in production of chiral substances, e.g. production of (2R,3S)-ethyl 3-phenylglycidate with 95% enantiomeric excess and 26% yield in 12 h from 0.2% (w/v) of the racemat by whole cells of Pseudomonas sp. strain BZS21, maximal activity with dimethyl formamide as co-solvent 3.3.2.9 microsomal epoxide hydrolase synthesis the purified recombinant enzyme can be used as biocatalyst for kinetic resolution of racemic styrene oxide with the result of over 99% enantiopure (S)-styrene oxide in 23,5% yield 3.3.2.9 microsomal epoxide hydrolase synthesis enzyme prefers (R)-styrene oxide. Production of enantiopure (S)-styrene oxide by use of enzyme in batch kinetic resolution of racemic styrene oxide 3.3.2.9 microsomal epoxide hydrolase synthesis highly enantioselective synthesis of chiral 1,2 diols from epoxides in ionic liquid [bmim][PF6] or [bmim][Tf2N] in presence of 10% water 3.3.2.10 soluble epoxide hydrolase synthesis the enzyme may be a good biocatalyst for the preparation of enantiopure epoxides or diols 3.3.2.10 soluble epoxide hydrolase synthesis potential as biocatalyst for the preparation of enantiopure epoxides 3.3.2.10 soluble epoxide hydrolase synthesis the enantioselective enzyme is useful in production of chiral substances, e.g. production of (2R,3S)-ethyl 3-phenylglycidate with 95% enantiomeric excess and 26% yield in 12 h from 0.2% (w/v) of the racemat by whole cells of Pseudomonas sp. strain BZS21, maximal activity with dimethyl formamide as co-solvent 3.3.2.10 soluble epoxide hydrolase synthesis the enzyme is useful for enantioselective bio-organic synthesis of chiral substances 3.3.2.10 soluble epoxide hydrolase synthesis enantioselective hydrolysis of racemic styrene derivative via attack of the benzylic position results in formation of the correspponding (R)-diol with enantiomeric excess of up to 96% and more than 90% conversion 3.3.2.10 soluble epoxide hydrolase synthesis highly enantioselective synthesis of chiral 1,2 diols from epoxides in ionic liquid [bmim][PF6] or [bmim][Tf2N] in presence of 10% water 3.4.11.1 leucyl aminopeptidase synthesis production of polyketide antibiotics 3.4.11.1 leucyl aminopeptidase synthesis production of alpha-amino acids, which are intermediates in the synthesis of antibiotics, injectables, food and feed additives 3.4.11.7 glutamyl aminopeptidase synthesis the enzyme is interesting for an industrial application, because of the high specificity for N-terminal Asp and Glu 3.4.11.10 bacterial leucyl aminopeptidase synthesis LAP is an important enzyme for the industrial production of enantiomerically pure amino acids 3.4.11.10 bacterial leucyl aminopeptidase synthesis leucine aminopeptidase from Vibrio proteolyticus is a broad specificity N-terminal aminopeptidase that is widely used in pharmaceutical processes where the removal of N-terminal residues in recombinant proteins is required 3.4.11.17 tryptophanyl aminopeptidase synthesis part of method for Trp-production 3.4.11.17 tryptophanyl aminopeptidase synthesis the enzyme is useful for synthesis of L-Trp because it is not inhibited by high levels of the product 3.4.11.18 methionyl aminopeptidase synthesis recombinant human interferon alpha-2b is produced in Escherichia coli in two types of molecules, one type, in majority, having N-terminal methionine intact, whereas the other type, in minority, having the N-terminal methionine cleaved by methionine aminopeptidase of the host. The N-terminal methionine of the remaining molecules can be removed by utilizing methionine aminopeptidase from Pyrococcus furiosus 3.4.11.18 methionyl aminopeptidase synthesis the covalently immobilized enzyme bound to iminodiacetic acid-agarose or chloroacetamido-hexyl-agarose shows long-term stability and allows a continuous, heterogenous processing of N-terminal methionines, for example, in recombinant proteins. Activation by zinc avoids the introduction of heavy metals with toxicological liabilities and oxidative potential into biotechnological processes 3.4.11.19 D-stereospecific aminopeptidase synthesis production of D-amino acids from L-amino acid amides by combination of alpha-amino-epsilon-caprolactam racemase, EC 5.1.1.15, and enzyme. Yield of conversion of L-alanine amid is more than 99% 3.4.11.19 D-stereospecific aminopeptidase synthesis enantioselective hydrolysis of 2-aminobutanamide by 10 g/l recombinant cells harboring Dap results in 51.8% conversion in 10 min and 99.8% e.e, of (S)-2-aminobutanamide 3.4.11.21 aspartyl aminopeptidase synthesis dipeptide sweeteners aspartame and alitame 3.4.11.23 PepB aminopeptidase synthesis enzyme can be used for synthesis of alkoxy-serines from DL-beta-alkoxy-alpha-amino propionamides 3.4.11.25 beta-peptidyl aminopeptidase synthesis attachment of a beta-amino acid to the N-terminus of a natural alpha-peptide. N-terminal beta-amino acid residues may be considered as protective groups against proteolytic enzymes in vitro and in vivo 3.4.11.25 beta-peptidyl aminopeptidase synthesis BapA catalyzes reversible protein acylation at the N-terminus. The selective modification can also be applied for protein labeling and tagging and should be generally useful, also to protect peptides and proteins from attack by common aminopeptidase 3.4.13.20 beta-Ala-His dipeptidase synthesis attachment of a beta-amino acid to the N-terminus of a natural alpha-peptide. N-terminal beta-amino acid residues may be considered as protective groups against proteolytic enzymes in vitro and in vivo 3.4.13.22 D-Ala-D-Ala dipeptidase synthesis novel cell breakage method based on VanX. The D-Ala-D-Ala dipeptidase encoded in a vancomycin-resistant VanA gene cluster, exhibits a strong cell lysis activity when expressed in isolation in Escherichia coli. Coexpression of VanX with the target protein causes cell autolysis and release of the cellular content into the culture medium. Application of this strategy for two model proteins, a green fluorescent protein variant and Gaussia luciferase, and optimization of the autolysis conditions and coexpression vectors shows that the fluorescence activity of green fluorescent protein variant collected from the medium is identical to that of green fluorescent protein variant purified by conventional methods. Cell breakage by VanX-mediated autolysis is very simple to implement and will efficiently complement traditional methods 3.4.13.22 D-Ala-D-Ala dipeptidase synthesis strong bacteriolysis occurrs when isolated VanX is expressed in Escherichia coli at temperatures lower than 30°C. No cell lysis is observed when VanX is expressed, even in large quantities, in the cell inclusion bodies at 37°C, suggesting that a natively folded VanX is required for lysis. In addition, VanX mutants with suppressed dipeptidase activity do not lyse Escherichia coli cells, confirming that bacteriolysis originates from the dipeptidase activity of VanX. There are also shape changes in Escherichia coli cells undergoing VanX-mediated lysis, these changes may be classified into three classes: bursting, deformation, and leaking fluid 3.4.14.1 dipeptidyl-peptidase I synthesis specific and efficient method for complete removal of polyhistidine purification tags from the N-termini of target proteins 3.4.14.11 Xaa-Pro dipeptidyl-peptidase synthesis comparison of the effects of cell disruption methods on the activity of PepX. The optimized values of high-pressure homogenization are one cycle at 130 MPa providing activity of 114.47 mU/ml, while sonication gives an activity of 145.09 mU/ml at 28 min with 91% power and three cycles 3.4.14.12 Xaa-Xaa-Pro tripeptidyl-peptidase synthesis strategy for expression and onestep-purification of the prolyl tripeptidyl peptidase 3.4.16.4 serine-type D-Ala-D-Ala carboxypeptidase synthesis highest yield of protein expression and purification from 12 mg of protein per liter of culture at 3 l bioreactor-scale is achieved in Streptomyces venezuelae ATCC 10595, a fast growing Streptomyces susceptible to glycopeptides. The addition of His6-tag at the N-terminus of the protein abolishes its biological activity either in vitro or in vivo assays. The addition of His6-tag at the C-terminus of the protein leads to a functional protein 3.4.16.5 carboxypeptidase C synthesis synthesis of CPY in Pichia pastoris as procarboxypeptidase Y with a yield of about 605 mg/l in shake-flasks after 168 h induction with 1 % (v/v) methanol. This precursor is cleaved by endogenous proteinases of Pichia pastoris and released into the fermentation broth as active carboxypeptidase Y within 2 weeks at 10°C. The recombinant enzyme is optimally active at 30°C and pH 6.0, with an optimal activity of about 305 U/mg 3.4.17.1 carboxypeptidase A synthesis enzymatic modification of human haemoglobin, useful for functional studies 3.4.17.1 carboxypeptidase A synthesis immobilization for synthetic use 3.4.17.1 carboxypeptidase A synthesis useful tools for peptide biosynthesis in non-conventional media, considered 3.4.17.1 carboxypeptidase A synthesis procedure for the production of CPA in the cytosol of Escherichia coli that yields approximately 0.5 mg of pure enzyme per liter of cell culture. The expression strategy maintains the proCPA zymogen in a soluble state by fusing it to the C-terminus of maltose-binding protein while simultaneously overproducing the protein disulfide isomerase DsbC in the cytosol from a separate plasmid. The yield of active and properly oxidized CPA is highest when coexpressed with DsbC in BL21(DE3) cells that do not also contain mutations in the trxB and gor genes. Most of the active CPA is generated after cell lysis and amylose affinity purification of the MBP-proCPA fusion protein, during the time that the partially purified protein is held overnight at 4°C prior to activation with thermolysin 3.4.17.B1 Sulfolobus solfataricus carboxypeptidase synthesis nanobioconjugate of the enzyme immobilized on silica-coated magnetic nanoparticles exhibits enhanced stability in aqueous media at room temperature as well as in different organic solvents. The improved stability in ethanol paves the way to possible applications of the immobilized enzyme, in particular as a biocatalyst for the synthesis of N-blocked amino acids. Another potential application might be amino acid racemate resolution, a critical and expensive step in chemical synthesis 3.4.17.2 carboxypeptidase B synthesis production of human insulin 3.4.17.19 Carboxypeptidase Taq synthesis expression of enzyme gene optimized for the Pichia pastoris codon usage and secretory expression. System is applicable for large-scale preparation of enzyme 3.4.19.1 acylaminoacyl-peptidase synthesis the enzyme is a very useful tool for the synthesis and modification of peptides. The stabilized Sepharose-coupled form of the enzyme is used to couple a carboxy-methylated N-formyl amino acid or N-acetyl amino acid to a short pre-existing peptide 3.4.19.12 ubiquitinyl hydrolase 1 synthesis used as molecular scissors for releasing a peptide or protein product 3.4.19.16 glucosinolate gamma-glutamyl hydrolase synthesis transient expression in Nicotiana benthamiana results in increased benzylglucosinolate levels that are accompanied by a high accumulation of the last intermediate in benzylglucosinolate synthesis, desulfobenzylglucosinolate, and a derivative thereof. Coexpression of adenosine 5'-phosphosulfate kinase APK2 reduces the accumulation of desulfobenzylglucosinolate and its derivative by more than 98% and increases benzylglucosinolate accumulation 16fold 3.4.21.1 chymotrypsin synthesis reaction catalyzer in apolar organic solvents 3.4.21.1 chymotrypsin synthesis enzymes are often used in organic solvents for catalyzing organic synthesis. Two enzyme preparations, EPRP (enzyme precipitated and rinsed with n-propanol) and PCMC (protein coated microcrystals) show much higher activities than lyophilized powders in such systems. Both preparations involve precipitation by an organic solvent 3.4.21.1 chymotrypsin synthesis construction of three different molecular weight multimodal temperature-responsive chymotrypsin-poly(sulfobetaine methacrylamide)-block-poly(N-isopropylacrylamide) protein-polymer conjugates that respond structurally to both low and high temperature. In the block copolymer grown from the surface of the enzyme, upper critical solution temperature phase transition is dependent on the chain length of the polymers in the conjugates, whereas lower critical solution temperature phase transition is independent of molecular weight. Each protein conjugate shows temperature dependent changes in substrate affinity and productivity when assayed from 0 to 40°C. The conjugates show higher stability to harsh conditions, including temperature, low pH, and protease degradation. The modified enzyme is active for over 8 h in the presence of a stomach protease at pH 1.0 3.4.21.1 chymotrypsin synthesis use of immobilized bovine alpha-chymotrypsin in a biotransformation process applying an aqueous micellar two-phase system for separation and recycling of the enzyme immobilisates. The thermoresponsive surfactant, Eumulgin ES, has no influence on the activity up to a concentration of 10%, while surfactants Tween 20, Triton X 114, Triton X 100 lead to an activity reduction. For Triton X-114, the activity rises to its previous value after washing or a buffer exchange influence. Optimization of the aqueous micellar two-phase system composition via a Design of Experiments approach allows for reuse of the immobilisates for hemoglobin digestion over eleven cycles in an Eumulgin ES aqueous micellar two-phase system 3.4.21.4 trypsin synthesis synthesis of benzoyl-Arg leucinamide by stabilized trypsin 3.4.21.4 trypsin synthesis production of human insulin-threonine-ester, which can be simply converted into human insulin by hydrolysis with subsequent purification steps 3.4.21.4 trypsin synthesis production of mono/di-arg insulin 3.4.21.4 trypsin synthesis attachment of O-carboxymethyl-poly-beta-cyclodextrin with molecular weight of 13000 Da to surface of enzyme. resulting neoglycoenzyme retains high proteolytic and esterolytic activity, optimum temperature is increased by 10°C, enzyme is more resistant to thermal inactivation and to denaturation by sodiumdodecyl sulfate 3.4.21.4 trypsin synthesis covalent immobilization of enzyme onto poly(methyl methacrylate)-co-(ethyl acrylate)-co-(acrylic acid) latex particles. Immobilized enzyme shows higher optimal temperature and pH-value than free form. Immobilized enzyme exhibits higher KM-value than free form and better chemical and thermal stability, it maintains 63% of initial activity after reusing ten times 3.4.21.4 trypsin synthesis immobilisation of enzyme on Fe3O4 nanoparticles. The immobilized enzyme is slightly morestable than the free enzyme at 45°C. 55% of activity of immobilized trypsin remains at 44°C after 2 h and 90% after 120 days storage. 40% of its initial activity remains after eight times of successive reuse 3.4.21.5 thrombin synthesis production of recombinant human prethrombin-2 in mouse myeloma cells, activation by recombinant ecarin and purification by affinity chromatography. Yield is about 70%, product is indistinguishable from plasma-derived enzyme 3.4.21.5 thrombin synthesis preparation of thrombin from human plasma. Isolation of prothrombin is followed by activation to thrombin and further purification, process is suitable for large-scale production with a high degree of virus safety 3.4.21.5 thrombin synthesis use of thrombin as enhancer in polymerase chain reaction. Presence of bovine thrombin is exceptionally effective at preventing the formation of primer dimers and enhancing the formation of the desired polymerase chain reaction products. The PCR enhancement effects of thrombin apply to low-copy synthetic single-stranded DNAs, synthetic ssDNA pools, human genomic DNA, or hepatitis B virus genomic DNA. Thrombin is also able to effectively relieve PCR inhibition by nanomaterial inhibitors such as gold nanoparticles and graphene oxide. Compared with bovine serum albumin, thrombin is more effective and requires concentrations 18-178 times less than that of serum albumin to achieve a similar level of PCR enhancement 3.4.21.6 coagulation factor Xa synthesis enzyme is a target for design of specific inhibitors 3.4.21.9 enteropeptidase synthesis the enzyme may be useful in amino acid sequence studies for the production of large fragents. The enzyme may also be useful in DNA-recombinant studies in releasing the desired polypeptide chain from neighboring sequences 3.4.21.9 enteropeptidase synthesis the cleavage immediately after the carboxyl-terminal residue of the (Asp)4-Lys recognition sequence allows regeneration of native amino-terminal residues of recombinant proteins, e.g. removal of the thioredoxin and polyhistidine fusion partners from proteins of intrest 3.4.21.9 enteropeptidase synthesis the enzyme can be used for cleavage of fusion proteins due to its high specific activity 3.4.21.9 enteropeptidase synthesis useful tool for in vitro cleavage of fusion proteins 3.4.21.9 enteropeptidase synthesis gene engineering studies on processing fusion proteins 3.4.21.9 enteropeptidase synthesis tool protease in the research and production of gene engineering 3.4.21.9 enteropeptidase synthesis a huge number of therapeutic proteins such as antibodies, coagulation factors, growth hormones or vaccines are produced as fusion proteins. To obtain the therapeutic protein in its monomeric, active form, the fusion partner has to be removed either by chemical or enzymatic cleavage. Enterokinase is a very attractive tool for the in vitro cleavage of fusion proteins 3.4.21.19 glutamyl endopeptidase synthesis the enzyme could be useful for commercial preparation of casein phosphopeptides 3.4.21.19 glutamyl endopeptidase synthesis enzyme can be used as catalyst for peptide synthesis in hydrophilic organic solvents with low water content, e.g. acetonitrile, overview 3.4.21.19 glutamyl endopeptidase synthesis increased accumulation of recombinant enzyme in Bacillus subtilis by growth in presence of casein or gelatin. During stationary growth phase, enzyme production is stimulated by Ca2+, Mn2+, and Co2+, and inhibited by Zn2+, Fe2+, and Cu2+ 3.4.21.19 glutamyl endopeptidase synthesis increased accumulation of recombinant enzyme in Bacillus subtilis during stationary growth phase by growth in presence of phosphate or ammonium ions, and in presence of gelatin or casein. During sporulation, enzyme production is stimulated by Ca2+, Mn2+, and Co2+, and inhibited by Zn2+, Fe2+, and Cu2+. Glucose is not inhibitory to enzyme production during stationary growth 3.4.21.19 glutamyl endopeptidase synthesis recombinant expression of enzyme as insoluble inclusion bodies, solubilization in 6 M guanidine-HCl in presence of reducing agent and renaturation by fast frequent dilution method. Highest yield of refolded protein at pH 8.4, 4°C. Renaturation is accompanied by gradual splitting of K12-E13 and T47-E48 bonds resulting in a 26 kDa protein that is converted to 25 kDa mature protein by limited proteolysis trypsin or subtilisin. Complete cleavage of N-terminal pro-peptide is necessary for final packing and activation of enzyme 3.4.21.38 coagulation factor XIIa synthesis the enzyme is used for preparation of factor Xia and plasma kallikrein 3.4.21.48 cerevisin synthesis high-level production of human mechano-growth factor in Saccharomyces cerevisiae is interfered by proteinase B. Expression of mechano-growth factor in a proteinase B deletion mutant results in a fivefold increase of yield 3.4.21.48 cerevisin synthesis much stricter experimental protocols than those routinely used are necessary to prevent the artefactual interpretation of protein levels in strains or conditions that increase proteinase B activity. Stability of some proteins, such as Slt2p or Chs4p, but not others, is severely compromised in the rim101 mutant due to the upregulation of the PRB1 gene. Proteolytic degradation during protein processing can be almost completely prevented by an overdose of subtilisin-like protease inhibitors, such as PMSF, or by avoiding cell freezing 3.4.21.50 lysyl endopeptidase synthesis use of the immobilized enzyme in the enzyme-assisted semisynthesis of human insulin 3.4.21.62 Subtilisin synthesis C221 subtilisin is catalytically wounded to the point that will barely hydrolyze peptide bonds but turn to be quite reactive with certain activated ester substrates, therefore a useful tool for catalyzing synthetic reactions 3.4.21.62 Subtilisin synthesis subtilisin BPN' is an important industrial enzyme 3.4.21.62 Subtilisin synthesis production of (R)-(2-methylpropyl)butanedioic acid 4-ethyl ester 3.4.21.62 Subtilisin synthesis production of (R)-2-benzyl-3-[[1-methyl-1-((morpholin-4-yl)-carbonyl)ehtyl]sulfonyl]propionic acid-ethyl ester, which was prepared as an intermediate for the renin inhibitors ciprokiren and remikiren 3.4.21.62 Subtilisin synthesis production of (S)-2-benzyl-3-(tert-butylsulfonyl)propionic acid, which was prepared as an intermediate for the renin inhibitor remikiren 3.4.21.62 Subtilisin synthesis production of phenylalanine, which is used as an intermediate for the synthesis of aspartame 3.4.21.62 Subtilisin synthesis conjugation of enzyme with comb-shaped poly-(ethylen glycol) and solubilization in ionic liquids without adding water. Enzyme exhibits higher transesterification activity in solution of [Eminm][Tf2N] than in toluene. No enzymic activity in DMSO, THF, or acetonitrile 3.4.21.62 Subtilisin synthesis preparation of liposomes containing the enzyme and modulation of membrane permeability by cholate results in a nano-bioreactor system with higher apparent substrate selectivity than free enzyme due to different permeation of membrane by the substrates 3.4.21.62 Subtilisin synthesis synthesis of benzyloxycarbonyl-L-Asp-L-Ser-NH2 with benzyloxycarbonyl-L-Asp methyl ester as acyl donor and serine amide as the nucleophile. Optimum conditions are pH 10.0, 35°C, in acetonitrile/Na2CO3-NaHCO3 buffer system, 85:15, with a dipeptide yield of 75.5% 3.4.21.62 Subtilisin synthesis the engineered enzyme displays synthetically useful enantioselectivity for most of the secondary alcohols tested. The enantioselectivity of ISCBLS is in particular good to high for m- or p-substituted 1-phenylethanols. The dynamic kinetic resolutions of these secondary alcohols by the combination of ISCBLS and a ruthenium-based racemization catalyst provide the products of (S)-configuration with good results (80-94% yield, 90-99% enantiomeric excess). Enzyme CBLS is of great use as the enantiocomplementary counterpart of (R)-selective lipase for the dynamic kinetic resolution of secondary alcohols 3.4.21.62 Subtilisin synthesis the enzyme subtilisin Carlsberg (SC) is industrially applied or synthetic organic chemistry applications, it has been used to enrich the S enantiomer of a racemic alcohol mixture by dynamic kinetic resolution (DKR) systems by combining SC with a ruthenium complex in organic solvent 3.4.21.64 peptidase K synthesis chemoenzymatic synthesis of oligo(L-phenylalanine) by the enzyme as a green and clean chemical reaction compared to organic synthesis 3.4.21.79 granzyme B synthesis expression system for the production of high yields of enzymatic and biologically active human grB by transfection of HEK-293 with grB. The HEK-293 host cells are protected from apoptotic cell death by fusing an inactivation site coupled to a (His)6 tag to the gene sequence of GrB. Inactive grB which is actively released from HEK-293 cells by insertion of a Igkappa leader sequence is purified on a nickel column utilizing the (His)6 tag. After enterokinase digestion and heparin affinity chromatography, high yields of enzymatic and biologically active human grB are obtained 3.4.21.82 Glutamyl endopeptidase II synthesis peptide coupling catalyzed by proteasesis an alternative to chemical solution and solid phase peptide synthesis 3.4.21.91 Flavivirin synthesis expression and purification of a natural form of DENV protease containing the full-length NS2B protein and the protease domain of NS3. The protease is expressed in Escherichia coli and purified in detergent micelles necessary for its folding. This purified protein is active in detergent micelles such as lyso-myristoyl phosphatidylcholine 3.4.21.96 Lactocepin synthesis optimisation of enzymatic hydrolysis of beta-casein to produce the angiotensin-I-converting enzyme (ACE) inhibitory peptides. Under optimal conditions (enzyme-to-substrate ([E]/[S]) ratio (w/w) of 0.132 and pH of 8.00 at 38.8°C), the ACE inhibitory activity of hydrolysates is 72.06% and the total peptides is 11.75 mg/ml. The resulting hydrolysates have higher thermal stability than beta-casein and show an increase in the free sulfhydryl content compared with raw beta-casein 3.4.22.2 papain synthesis use as catalyst in asymetric synthesis of acyl derivatives and in peptide synthesis 3.4.22.2 papain synthesis chemical modification of papain using different anhydrides of 1,2,4-benzenetricarboxylic and pyromellitic acids and immobilization on cotton fabric results in immobilized papain with optimum pH shifted from 6.0 to 9.0. Compared with immobilized native papain, the thermal stability and the resistance to alkali and washing detergent of immobilized modified enzyme are improved considerably. When the concentration of detergent is 20 mg/ml, the activity of the immobilized pyromellitic papain retains about 40% of its original activity, whereas the native papain is almost inhibited 3.4.22.2 papain synthesis enzymatic hydrolysis of casein to produce free amino acids by papain in a two-phase system, which is composed of n-propanol, NaCl and water. In this system, the top phase contains more n-propanol and the bottom phase contains more NaCl and water. Papain and casein are mainly distributed in bottom phase, and free aromatic amino acids tyrosine, tryptophan and phenylalanine produced by enzymatic hydrolysis aere mainly in top phase. When the two-phase system consists of 44% n-propanol, 60 g/l NaCl, 0.15 g/l papain and 13 g/l casein at 55°C and pH 5.6, the transformation yield is 99.5% 3.4.22.2 papain synthesis Fab antibody fragment production and purification by papain digestion of an intact monoclonal antibody. After digestion, separation of the Fab fragment from the Fc fragment and residual intact antibody is achieved using protein A affinity chromatography. The Fab fragments are of high quality suitable to produce diffraction quality crystals suitable for X-ray crystallographic analysis 3.4.22.2 papain synthesis immobilization of papain on Sepharose 6B in the presence of different concentrations of cysteine affect the enzyme activity depending on cysteine concentration. The maximum specific activity is observed when papain was immobilized with 200 mM cysteine. The immobilization process results in significant enhancement of stability to temperature and extreme pH. After immobilization, the optimum temperature of papain activity increases by 20 degrees from 60 to 80°C and its optimum pH activity shifts from 6.5 to 8.0. Catalytic efficiency and specific activity of the immobilized enzyme do not significantly change after immobilization 3.4.22.3 ficain synthesis kinetically controlled formation of peptide bonds by coupling the ester substrates benzyloxycarbonyl-Ala methyl ester and benzyloxycarbonyl-Gly methyl ester with L-Ala, D-Ala, L-Gln, D-Gln and L-Cys(acetamidomethyl) respectively 3.4.22.3 ficain synthesis digestion of murine monoclonal IgG to obtain immunoreactive bivalent antibody fragments 3.4.22.8 clostripain synthesis the enzyme can be used for the high yielding synthesis of a variety of peptide bond using L-Arg and, to a lesser degree, L-Lys as substrates with L-amino acid amides or D-amino acid amides 3.4.22.8 clostripain synthesis efficient biocatalyst for the synthesis of peptide isosteres 3.4.22.8 clostripain synthesis recombinant clostripain might prove useful in the production of insulin from the proinsulin fusion protein 3.4.22.14 actinidain synthesis the enzyme can be used efficiently for hydrolysis of collagen and isolation of different cell populations from various solid tissues 3.4.22.68 Ulp1 peptidase synthesis comparison of Ulp1 protease in active upon expression as inclusion bodies and soluble protein. Fusion of the N-terminal selfassembling peptide GFIL8 to Ulp1 increases production of active inclusion bodies in Escherichia coli. Attachment of the N-terminal cellulose-binding module facilitates the immobilization on regenerated amorphous cellulose with a binding capacity up to about 235 mg protein per gram of cellulose. The immobilized soluble Ulp1 maintains about 42% initial cleavage activity with repetitive use, whereas the aggregated Ulp1 loses its cleavage capacity after cleaving the protein substrate once. Crosslinking of inclusion bodies using glutaraldehyde inactivates Ulp1 3.4.22.68 Ulp1 peptidase synthesis immobilization of Ulp1 as a tool for cleavage of the SUMO tag of recombinant proteins. Ulp1 immobilized on N-hydroxysuccinimide-activated Sepharose maintains 95% substrate-cleavage ability and significantly enhanced pH and thermal stability. The immobilized Ulp1 can tolerate 15% (v/v) DMSO and 20% (v/v) ethanol. It can be reused for more than 15 batch reactions with 90% activity retention 3.4.22.68 Ulp1 peptidase synthesis optimization of expression of the catalytic domain. Optimization of cultivation conditions at shake flask results in Ulp1 expression of 195 mg/l in TB medium. Ni-NTA affinity purification of Ulp1 using 0.1% Triton X-100, 0.01mM DTT, 0.02mM EDTA and 1% glycerol leads to a purity of about 95% with a recovery yield of 80% and specific activity of 398600 U/mg. The protease cleaves the SUMO tag even at 1:10,000 enzyme to substrate ratio The in vivo cleavage of SUMO tag via coexpression strategy also results in more than 80% cleavage of SUMO fusion protein 3.4.22.70 sortase A synthesis in vitro applications of sortase A to protein conjugation. Application of recombinant Staphylococcus aureus sortase A to attach a tagged model protein substrate (green fluorescent protein) to polystyrene beads chemically modified with either alkylamine or the in vivo sortase A ligand, Gly-Gly-Gly, on their surfaces. Sortase A can be used to sequence-specifically ligate eGFP to amino-terminated poly(ethylene glycol) and to generate protein oligomers and cyclized monomers using suitably tagged eGFP An alkylamine can substitute for the natural Gly3 substrate, which suggests the possibility of using the enzyme in materials applications. The highly specific and mild sortase A-catalyzed reaction, based on small recognition tags unlikely to interfere with protein expression represents a useful addition to the protein immobilization and modification tool kit 3.4.22.70 sortase A synthesis semienzymatic cyclization of disulfide-rich peptides using sortase A, overview 3.4.22.70 sortase A synthesis SrtA works as a versatile tool in protein engineering 3.4.22.70 sortase A synthesis the bacterial transpeptidase sortase A is a well-established tool in protein chemistry and catalyzes the chemoselective ligation of peptides and proteins 3.4.23.1 pepsin A synthesis comparison of hydrolysis kinetics of hemoglobin after immobilization of enzyme on aluminium oxide and on 2-ethanolamine-O-phosphate-modified acidic alumina. Modified alumina results in comparatively less adsorption of peptides and complete adsorption of heme 3.4.23.4 chymosin synthesis use of chymosin to cleave a pro-chymosin derived fusion tag releasing native target proteins. After modification of the pro-chymosin fusion tag chymosin can remove this tag at more neutral pH 6.2, less prone to compromise the integrity of target proteins. Chymosin produces intact native target protein both at the level of small and large-scale preparations 3.4.23.41 yapsin 1 synthesis single disruption of either PEP4 gene or yapsin family member YPS1 gene leads to reduced degradation of recombinant human serum albumin and human parathyroid hormone upon expression in Pichia pastoris. In a PEP4YPS1 double disruptant, more than 80% of the human serum albumin and human parathyroid hormone secreted by the remains intact, after 72 h of incubation, in comparison to only 30% with the wild-type strain 3.4.23.43 prepilin peptidase synthesis scaled-up synthesis of PilD, followed by solubilization in dodecyl-beta-D-maltoside and chromatography, leads to a pure enzyme that retains its known biochemical activities 3.4.23.52 preflagellin peptidase synthesis scaled-up synthesis of PilD, followed by solubilization in dodecyl-beta-D-maltoside and chromatography, leads to a pure enzyme that retains its known biochemical activities 3.4.24.7 interstitial collagenase synthesis improved method for high-level expression of soluble human MMP-1 catalytic domain in Escherichia coli 3.4.24.B7 matrix metalloproteinase-26 synthesis use of Brevibacillus choshinensis as the host system for the soluble and active expression of MMP26. The enzyme is secreted in soluble form in the supernatant of cell culture medium. The yields of purified proform of MMP26 and catalytic form of MMP-26 are 12 and 18 mg/L, respectively, with high purity and homogeneity. Both pro- and catalytic form show gelatin zymography activity and the purified catalytic form has high enzymatic activity against DQ-gelatin substrate. Expression using several widely used expression vectors in Escheriochia coli cells results in insoluble expressions or soluble expressions with little catalytic activity 3.4.24.20 peptidyl-Lys metalloendopeptidase synthesis isolation of blocked N-terminal peptides 3.4.24.24 gelatinase A synthesis application of MMP-2 from sea cucumber (Stichopus japonicas) to prepare bioactive collagen hydrolysate 3.4.24.26 pseudolysin synthesis the Pseudomonas aeruginosa elastase, produced by Pichia pastoris, is a promising biocatalyst for peptide synthesis in organic solvents 3.4.24.26 pseudolysin synthesis the enzyme can be applied to obtain bioactive soluble peptides from eggshell-membrane. Potential applicability of its peptides as functional food and cosmetic additives 3.4.24.B26 myroilysin synthesis optimization of the culture conditions for protease production, best conditions are (w/v) 1% corn powder, 4% wheat bran, 2% bean powder, 0.4% Na2HPO4, 0.03% KH2PO4, 0.1% CaCl2, and artificial seawater, with the initial pH of 8.0 and 1% inoculation amount, 15°C culture temperature. Under the optimized conditions, the protease activity reaches 1137 U/ml, i.e., 174% of that before optimization. The protease activity in small scale fermentations reaches 1546.4 U/ml after parameter optimization 3.4.24.27 thermolysin synthesis production of L-alpha-aspartame, which is used as a low-calorie sweetener in food, including soft drinks, table-top sweeteners, dairy products, instant mixes, dressings, jams, confectionary, toppings and in pharmaceuticals 3.4.24.27 thermolysin synthesis hydrolysis and condensation reactions of peptides catalyzed by enzyme can be reversibly controlled by on/off ultrasound irradiation depending on its frequency region 3.4.24.27 thermolysin synthesis immobilized enzyme catalyzes the formation of beta-cyclodextrin esters using vinyl esters of butyrate, decanoate and laurate, as acyl donors in dimethylsulfoxide. Esterification occurs exclusively at the glucose C-2 position. Enzyme also catalyzes the synthesis of alpha-, beta-, gamma- and maltosyl-beta-cyclodextrin esters with vinyl laurate as the acyl donor in dimethylsulfoxide and dimethylformamide 3.4.24.27 thermolysin synthesis immobilized enzyme catalyzes the transesterification of vinyl laurate to several sucrose-containing tri- and tetrasaccharides. Preferred position of acylation is the 2-OH group of the alpha-D-glucopyranose moiety linked 1 to 2 to the beta-D-fructofuranose unit 3.4.24.27 thermolysin synthesis introduction of ionizing residues into the active site of enzyme as a means of modifying its pH-activity profile 3.4.24.27 thermolysin synthesis the enzyme is used for synthesis of N-carbobenzyloxy L-Asp-L-Phe methyl ester, a precursor of the artificial sweetener aspartam 3.4.24.27 thermolysin synthesis thermolysin is industrially used for the synthesis of N-carbobenzoxy-L-aspartyl-L-phenylalanine methyl ester, a precursor of an artificial sweetener aspartame, from N-carbobenzoxy-L-aspartic acid and L-phenylalanine methyl ester 3.4.24.27 thermolysin synthesis the enzyme can be used for production of caseicin A, an antimicrobial active peptide, from alpha-casein, for potential improvement of the safety of infant milk formula using milk-derived bioactive peptides 3.4.24.27 thermolysin synthesis using the enzyme to catalyze the condensation of the chiral aspartame-precursor, carbobenzoxy-L-aspartyl-L-phenylalanine methyl ester, from the protected amino acid substrates carbobenzoxy-L-aspartic acid and L-phenylalanine methyl ester in large scale production. Analysis of the protease mediated peptide synthesisof a precursor of the artificial sweetener aspartame, a multiton peptide synthesis catalyzed by the enzyme thermolysin. X-ray structures of thermolysin in complex with aspartame substrates separately, and after protease mediated peptide synthesis in a crystal, rationalize the reaction's substrate preferences and reveal an unexpected form of substrate inhibition that explains its sluggishness. Structure guided optimization of this and other PMPS reactions could expand the economic viability of commercial peptides beyond current high-potency, low-volume therapeutics, with substantial green chemistry advantages 3.4.24.27 thermolysin synthesis sequential hydrolysis trypsin-thermolysin of commercial hydrolysate (prepared from solubilized milk proteins) has been performed to produce bioactive peptides, mainly casein phosphopeptides. Thermolysin was the last enzyme to be used, generating the final product, with a low content of aromatic amino acids and shorter peptides 3.4.24.28 bacillolysin synthesis production of enzyme as an active mature enzyme in Escherichia coli in the absence of its prosequence. Expression of enzyme with N-terminal His-tag, which accumulates as inclusion bodies and renaturation after solubilization using guanidine hydrochloride 3.4.24.28 bacillolysin synthesis protease N covalently immobilized on agarose can be used for the preparation of acetylated ribonucleosides with only one free hydroxyl group in position C-5', as useful building blocks for the synthesis of monophosphate nucleotides or 5'-deoxyribonucleosides such as Capecitabine 3.4.24.78 gpr endopeptidase synthesis the enzyme is synthesized only during sporulation within the developing spore, and as a zymogen, ca. 2 h after its synthesis the zymogen autoprocesses to the active enzyme by intramolecular removal of 15 N-terminal residues 3.4.24.78 gpr endopeptidase synthesis the enzyme is synthesized only during sporulation within the developing spore, and as a zymogen, ca. 2 h after its synthesis the zymogen autoprocesses to the active enzyme by intramolecular removal of 16 N-terminal residues 3.4.24.83 anthrax lethal factor endopeptidase synthesis preparation of semisynthetic protective antigen-binding domain of anthrax lethal factor, LFN, by native chemical ligation of synthetic LFN residues 14-28 thioester with recombinant N29C-LFN residues 29-263 and comparison with two variants containing alterations in residues 14-28 of the N-terminal region. The properties of the variants in blocking ion conductance through the protective antigen pore and translocating across planar phospholipid bilayers in response to a pH gradient are consistent with current concepts of the mechanism of polypeptide translocation through the pore. The semisynthesis platform allows for investigation of the interaction of the pore with its substrates 3.4.24.85 S2P endopeptidase synthesis in absence of cholesterol, SREBP double cleavage leads to activation of transcription of genes encoding multiple enzymes of the cholesterol biosynthetic pathway 3.4.25.1 proteasome endopeptidase complex synthesis the incorporation of parus subunits into Arabidopsis holoprotease raises the intriguing possibility that plants synthesize multiple 26S proteasome types with unique properties and/or target specificities 3.5.1.1 asparaginase synthesis production of L-asparaginase from Serratia marcescens SK-07 in a batch bioreactor. The optimal levels of L-asparagine, glucose, yeast extract and peptone are 0.93, 3.81, 3.65 and 1.47 g/l, respectively, and maximal L-asparaginase production of 25.02 U mg/1 can be obtained. L-asparagine is the most favourable carbon source for enhanced production of L-asparaginase 3.5.1.1 asparaginase synthesis conjugation of enzyme to PEG-chitosan and glycol-chitosan imoproves catalytic efficiency 3- to 6fold under physiological conditions and enhances its resistance to thermal inactivation 3.5.1.1 asparaginase synthesis optimization of production and process conditions for recombinant human enzyme. The maximum biomass yield of 6.7 g/l of enzyme is achieved with fed-batch fermentation. The refolding efficiency is optimal at pH 8.5 (84%) and temperature 25°C (86%) 3.5.1.1 asparaginase synthesis production and optimization of growth conditions results in submerged fermentation in modified M9 medium with yeast extract and fructose as carbon and nitrogen sources, respectively, at pH 8.0, incubated for 120 h at 30°C 3.5.1.1 asparaginase synthesis recombinant asparaginase isozyme AnsB fused with the pelB signal sequence and a five aspartate tag is secreted efficiently into culture medium at 34.6 U/mg cell of specific activity. By batch fermentation, recombinant Escherichia coli produces 40.8 U/ml asparaginase isozyme II in the medium. Deletion of the gspDE gene reduces extracellular production of asparaginase isozyme II 3.5.1.1 asparaginase synthesis strain is potent extracellular producer of L-asparaginase (347.42 IU) and shows 15fold enhancement of L-asparaginase production (5,205 U/gds) under submerged fermentation condition in 5 days in presence of inducers and activators. Red gram husk is the best substrate in solid state fermentation supporting maximum enzyme activity of 246.32 IU after 5 days of incubation 3.5.1.2 glutaminase synthesis optimal conditions for native enzyme production are pH 7.0,37°C and 0.3 % L-glutamine. Maltose (0.4 %, w/v) is best carbon source supplement 3.5.1.2 glutaminase synthesis the enzyme production is optimal with glucose as carbon source (33 U/ml), L-glutamine as nitrogen source (33.1 U/ml), at pH 7, 30°C and 0.1% NaCl 3.5.1.4 amidase synthesis - 3.5.1.4 amidase synthesis acrylamide production 3.5.1.4 amidase synthesis producing particular organic acids from nitriles and detoxifying waste waters containing nitrile compounds 3.5.1.4 amidase synthesis detoxification of acrylamide and as biocatalysts for the industrial production of ammonium acrylate and acrylic acid 3.5.1.4 amidase synthesis degradation of acrylamide 3.5.1.4 amidase synthesis synthesis of pharmacologically important peptides 3.5.1.4 amidase synthesis production of (R)-2,2-dimethylcyclopropanecarboxylic acid, which is an intermediate for the synthesis of the dehydropeptidase-inhibitor cilastatin 3.5.1.4 amidase synthesis production of (S)-piperazine-2-carboxylic acid, which is used as an intermediate for pharmaceuticals, e.g. the orally active HIV protease inhibitor crixican 3.5.1.4 amidase synthesis application in an one-pot preparation of amides from aldehydes together with nitrile hydratase, EC 4.2.1.84 3.5.1.4 amidase synthesis amidase-catalyzed production of nicotinic acid in batch and continuous stirred membrane reactors 3.5.1.4 amidase synthesis the amidase has potential for application under high temperature conditions as a biocatalyst for D-selective amide hydrolysis producing enantiomerically pure carboxylic acids and for production of novel amides by acyl transfer 3.5.1.4 amidase synthesis the enzyme is useful for production of optically pure S-arylpropionic acids 3.5.1.4 amidase synthesis the acyl transfer activity of the enzyme can be used for the synthesis of variety of hydroxamic acids. Optimization of physiochemical parameters result into 30fold increase in the acyl transfer activity of amidase. The acyl transfer activity of amidase of Alcaligenes sp. MTCC 10674 under high temperature condition makes of potential application in developing a bioprocess for the production of variety of aliphatic and aromatic hydroxamic acid 3.5.1.4 amidase synthesis the enzyme enzyme has very high potential for biotransformation of N-substituted aromatic amides and for the production of a variety of hydroxamic acids 3.5.1.5 urease synthesis production of ammonia 3.5.1.11 penicillin amidase synthesis synthesis of 6-aminopenicillanate 3.5.1.11 penicillin amidase synthesis synthesis of penicillin G 3.5.1.11 penicillin amidase synthesis despite the relatively weak reactivity of the exocyclic amino group of 2'-deoxyadenosine or 2'-deoxyguanosine under the conditions for nucleotide bond formation, its selective protection is an unavoidable step in oligonucleotide synthesis. The phenylacetyl group can be successfully used as an enzyme-cleavable aminoprotecting group of 2'-deoxyadenosine and 2'-deoxyguanosine. Penicillin amidase would be effective in multiple deprotection of oligonucleotides containing N-phenylacetylated purine nucleobases under mild conditions 3.5.1.11 penicillin amidase synthesis production of (2R,3S)-beta-lactam intermediate 3.5.1.11 penicillin amidase synthesis production of 6-aminopenicillanic acid, which is an intermediate for the manufacture of semi-synthetic penicillins 3.5.1.11 penicillin amidase synthesis production of cefadroxil, cephalexin and cefaclor, which are beta-lactam antibiotics 3.5.1.11 penicillin amidase synthesis enantioselective hydrolysis of N-phenylacetyl-DL-tert-leucine to yield L-tert-leucine. Rate of enzyme-catalyzed reaction is significantly affected by the presence of 2% organic cosolvent. Initial rate falls with increasing logP of the cosolvent, but for logP values less than -0.24, the rate is faster than that in aqueous medium. Yield of L-tert-leucine is above 95% with the enantiomeric excess of more than 99% 3.5.1.11 penicillin amidase synthesis enzyme-catalyzed synthesis of beta-lactam antibiotics in supersaturated solutions of substrates. Due to nucleophile supersaturation, ratio of synthesis/hydrolysis be be increased up to threefold 3.5.1.11 penicillin amidase synthesis hydrolysis of phenylacetic hydrazide compounds syntesized on PEGA1900 and PEGA+ copolymers of acrylamide and ethylene glycol by enzyme diffusing into the polymer. Use in specific cleavage of linkers 3.5.1.11 penicillin amidase synthesis improved synthesis of 3’-functionalized cephalosporins by enzyme in polyethylene glycol-ammonium sulfate aqueous two-phase systems. Best results are achieved using PEG 600, 80% in water, equilibrated with 4 M ammonium sulfate. Acylation product is completely partitioned in the PEG phase, whereas the substrates maintain a suitable concentration in the enzyme environment. Use for synthesis of cefamandole and cefonicid 3.5.1.11 penicillin amidase synthesis Markovnikov addition of allopurinol to vinyl ester by enzyme immobilized on acrylic beads. With vinyl acetate in dimethylsulfoxide, yield is about 60%. Decrease in activity in the order vinyl acetate, vinyl pentanoate, vinyl decanoate. Divinyl dicarboxylate reacts faster than the comparable mono-acid vinyl ester 3.5.1.11 penicillin amidase synthesis optimization of culture conditions for overexpression of Kluyvera citrophila enzyme in Escherichia coli. Highest enzyme activity is obtained at 28°C in TB medium only when culture pH value is close to its original pH of about 7.3 3.5.1.11 penicillin amidase synthesis overexpression of enzyme, intracellular proteolysis can be prevented by coexpression of GroEL/ES. Coexpression of trigger factor facilitates folding of soluble pro-enzymeand improves enzyme maturation. DnaK/J-GrpE have an effect for solubilization and also improve maturation 3.5.1.11 penicillin amidase synthesis purification of His-tagged enzyme secreted into periplasmic space by immobilized metal-ion affnity chromatography. Maximum specific activity of enzyme is 38.5 U/mg, yield is 70% 3.5.1.11 penicillin amidase synthesis synthesis of ampicillin. High ratio of synthesis to hydrolysis at up to 200 mM 6-aminopenicillic acid and 500 mM D-phenylglycine methyl ester at 25°C and pH 6.5. When concentration of 6-aminopenicillic acid reaches saturation, rate of hydrolysis tends toward zero 3.5.1.11 penicillin amidase synthesis production of 6-aminopenicillanate and 7-amino-desacetoxycephalosporanic acid as precursor for the production of semi-synthetic beta-lactam derivatives 3.5.1.11 penicillin amidase synthesis production of 6-aminopenicillanate as precursor for the production of semi-synthetic beta-lactam derivatives 3.5.1.11 penicillin amidase synthesis AuAAC should be considered an industrial biocatalyst with high potential in the production of semisynthetic penicillins 3.5.1.11 penicillin amidase synthesis D-Gln and D-Glu can be obtained in one step in high enantiomeric excess (97% and 90%, respectively) in an enzymatic kinetic resolution of their racemates. This is achieved by enantioselective conversion of the L-enantiomers to the N-phenylacetyl derivatives with phenylacetic acid methylester as an acyl donor in aqueous solution by using an F24A mutant of PGA from Escherichia coli. The high enantiomeric excess values are mainly due to the significantly suppressed hydrolysis rate of N-phenylacetyl-L-Gln and N-phenylacetyl-L-Glu, respectively, compared to wild-type PGA 3.5.1.11 penicillin amidase synthesis immobilized penicillin G acylase derivatives are biocatalysts that are industrially used for the hydrolysis of Penicillin G by fermentation and for the kinetically controlled synthesis of semi-synthetic beta-lactam antibiotics. The desired orientation of immobilized enzyme with the active site freely accessible can be obtained by increasing the density of Lys residues on a predetermined region of the enzyme. The designed biocatalysts display improved synthetic performances and are able to maintain a similar activity to the free enzymes. The activity of the immobilized enzyme proportionally improves with the number of introduced Lys 3.5.1.11 penicillin amidase synthesis optimization of conversion of penicillin G into 6-aminopenicillanic acid by cross-linked enzyme aggregates of Bacillus badius penicillin G acylase. The faster conversion of penicillin G to 6-aminopenicillanic acid by cross-linked enzyme aggregates of Bacillus badius penicillin G acylase and efficient reusability holds a strong potential for the industrial application 3.5.1.11 penicillin amidase synthesis penicillin G acylase-catalyzed acylation of beta-phenylalanine with phenylacetamide as an acyl donor is efficient with a high preferential enantioselectivity for (R)-beta-phenylalanine. Both (S)-beta-phenylalanine, at 98% ee, and (R)-beta-phenylalanine, at 99% ee, can be produced using simple separation procedures 3.5.1.11 penicillin amidase synthesis cross-linked enzyme aggregates are useful in the biocatalysis in reactions of organic synthesis, e.g. of cephalexin. Optimization of cross-linking and agent with best results from glutaraldehyde in a 0.15 ratio with enzyme giving 87% conversion and 5.4 mM cephalexin/g cross-linked enzyme aggregates, 90% of the cross-linked enzyme aggregates enzyme is active after 170 h under operating conditions, overview 3.5.1.11 penicillin amidase synthesis penicillin acylase is one of the key pharmaceutical enzymes in the production of polysynthetic beta-lactam antibiotics 3.5.1.11 penicillin amidase synthesis penicillin amidase from Alacaligenes faecalis is an attractive biocatalyst for hydrolysis of penicillin G for production of 6-aminopenicillanic acid, which is used in the synthesis of semi-synthetic beta-lactam antibiotics 3.5.1.11 penicillin amidase synthesis PVA is a pharmaceutically important enzyme to produce 6-aminopenicillanic acid 3.5.1.11 penicillin amidase synthesis the enzyme might be useful in the preparation of a wide range of semi-synthetic beta-lactam antibiotics from acyl side-chain precursors and beta-lactam nucleus 3.5.1.11 penicillin amidase synthesis the enzyme is applied for large scale synthesis of 6-aminopenicillanic acid and 7-amino-3-deacetoxycephalosporanic acid 3.5.1.11 penicillin amidase synthesis the enzyme KcPGA is quite resilient to harsh conditions and is easier to immobilize for the industrial hydrolysis of natural penicillins to generate the 6-aminopenicillin nucleus, which is the starting material for semisynthetic antibiotic production 3.5.1.11 penicillin amidase synthesis enzyme immobilized on alginate/chitosan hybrid beads can be used on large scale for the synthesis of beta-lactam 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis - 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis enzyme used for production of L-amino acids 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis industrial applications 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis thermostable enzyme useful for industrial production, optical resolution of DL-amino acids on an industrial scale, utilization in industrial production of stereoisomers from racemates 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis production of L-3-(4-thiazolyl)alanine and D-N-acetyl-D-3-(4-thiazolyl)alanine, which are used as a component of antihypertensive inhibitors of the enzyme renin 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis production of L-amino acids, which are used for parental nutrition, feed and food additives, cosmetics, pesticides, and as intermediates for pharmaceuticals as well as chiral synthons for organic synthesis 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis aminoacylase-catalyzed enantioselective synthesis of aromatic beta-amino acids 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis biocatalytic production of optically active amino acids and their derivatives, which are broadly used as physiologically active compounds, chiral auxiliaries or synthons and as intermediates in pharmaceutical, food and agrochemistry 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis pAcy1 catalyzes the highly stereoselective acylation of amino acids, a useful conversion for the preparation of optically pure N-acyl-L-amino acids, the enzyme acts as chiral catalyst 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis the enzyme can be used for production of optically pure L-Phe and D-Phe by usage of N-acetylated DL-Phe, which is hydrolyzed stereospecifically to L-Phe, the remaining N-acetyl-D-Phe can be chemically hydrolyzed to D-Phe. The method can also be applied to racemic Trp and Tyr 3.5.1.14 N-acyl-aliphatic-L-amino acid amidohydrolase synthesis because of its thermostability, the enzyme is expected to be useful for the production of L-amino acid derivatives from racemates at temperatures over 90°C 3.5.1.30 5-aminopentanamidase synthesis the enzyme is useful to produce the nylon-5 monomer 5-aminovalerate from L-lysine in large scale 3.5.1.30 5-aminopentanamidase synthesis 2-monooxygenase (DavB) and delta-aminovaleramidase (DavA) are coexpressed in Escherichia coli BL21(DE3) to produce nylon-5 monomer 5-aminovalerate from L-lysine. PP2911 (4-aminobutyrate transporter in Pseudomonas putida) and LysP (the lysine specific permease in Escherichia coli) are overexpressed to promote 5-aminovalerate production using whole cells of recombinant Escherichia coli. The constructed Escherichia coli strain overexpressing transport proteins exhibits good 5-aminovalerate production performance and might serve as a promising biocatalyst for 5-aminovalerate production from L-lysine. This strategy not only shows an efficient process for the production of nylon monomers but also might be used in production of other chemicals 3.5.1.41 chitin deacetylase synthesis industrial production of chitosan 3.5.1.41 chitin deacetylase synthesis the enzyme can be used for chitosan production from Aspergillus niger mycelium waste from citric acid production plant in cooperation with lysozyme, snailase, and neutral protease, analysis of chitosan structure 3.5.1.41 chitin deacetylase synthesis the enzyme can be used for production of chitin particles with only 1% deacetylated chitin which is still rigid and insoluble in acidic environment, but shows highly increased ovalbumin binding, chitosan as well is a good material for column chromatography showing no hydrogel formation 3.5.1.41 chitin deacetylase synthesis the enzyme might be useful for large scale production of chitosan from chitin 3.5.1.41 chitin deacetylase synthesis chitosan produced by the enzymatic method offers the possibility of a controlled, non-degradable deacetylation process that results in a more regular pattern of deacetylation 3.5.1.41 chitin deacetylase synthesis recombinant enzyme is used to produce partially acetylated chitosan oligosaccharides from chitin oligomers, whereby the pronounced regioselectivity of the enzyme leads to the production of defined products with novel patterns of acetylation 3.5.1.44 protein-glutamine glutaminase synthesis deamination of food proteins after treatment at 100°C for 15 min followed by proteolysis and alkali solubilization 3.5.1.52 peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase synthesis PNGase F can be utilized for glycosylation of non-glycosylated recombinant proteins produced in prokaryotic cells 3.5.1.57 tryptophanamidase synthesis enzymatic method of L-tryptophan production. The enzyme is useful for the manufacturing process because it is not inhibited by high levels of product 3.5.1.58 N-benzyloxycarbonylglycine hydrolase synthesis can be used in peptide synthesis procedures for removal of the protecting groups Z and Bz from Gly and Ala 3.5.1.69 glycosphingolipid deacylase synthesis a method for generation of novel fluorocarbon derivatives of glycosphingolipids has been developed. SCDase is used to remove the fatty acid from the ceramide moiety, after which a fluorocarbon-rich substituent is incorporated at the free amine of the sphingoid 3.5.1.69 glycosphingolipid deacylase synthesis the enzyme is immobilised on magnetic macroporous cellulose and is used to semisynthesise C17:0 glucosylceramide and C17:0 sulfatide, which are required as internal standards for quantification of the corresponding glycosphingolipids by tandem mass spectrometry. A high rate of conversion is achieved for both lipids, 80% for C17:0 sulfatide and 90% for C17:0 glucosylceramide. In contrast to synthesis with a soluble form of the enzyme, use of immobilised SCDase significantly reduces the contamination of the sphingolipid products with other isoforms, so further purification is not necessary 3.5.1.69 glycosphingolipid deacylase synthesis the enzyme can serve as a biocatalyst in the enzymatic synthesis of glycosphingolipids since it catalyzes the reversible hydrolysis/synthesis of the amide linkage between the fatty acid and the sphingosine base in the ceramide moiety of glycosphingolipids 3.5.1.69 glycosphingolipid deacylase synthesis sphingolipid ceramide N-deacylase is used for lyso-glycosphingolipid production. Application to large-scale preparation of gamglioside lyso-GM1, overview 3.5.1.70 aculeacin-A deacylase synthesis useful in producing peptide nuclei for creating new antifungal agents by introducing different acyl moieties 3.5.1.70 aculeacin-A deacylase synthesis useful in preparing deacylated peptides which are used as starting material for semisynthetic antifungal antibiotics. Enzyme is used industrially on a large scale to produce the peptide nuclei , i.e. deacetylated cyclic hexapeptides 3.5.1.70 aculeacin-A deacylase synthesis AuAAC should be considered an industrial biocatalyst with high potential in the production of semisynthetic penicillins 3.5.1.70 aculeacin-A deacylase synthesis antibiotic acylases are key enzymes for the industrial production of antibiotic drugs 3.5.1.73 carnitinamidase synthesis the enzyme is a valuable tool for L-carnitine production 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis enzyme is used for the industrial production of D-amino acids 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis the enzyme is suitable for use in the production of D-amino acids 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis combination of D-hydantoin hydrolase and N-carbamoyl-D-amino acid amidohydrolase is applicable for the production of various D-amino acids 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis D-amino acid production by Escherichia coli coexpressing three genes encoding hydantoin racemase, D-hydantoinase and N-carbamoyl-D-amino acid amidohydrolase. Efficient conversion of D-Phe, D-Tyr, D-Trp, O-benzyl-D-Ser, D-Val, D-norvaline, D-Leu and D-norleucine from the DL-5-monosubstituted hydantoin compounds 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis important enzyme involved in semi-synthesis of beta-lactam antibiotics in industry 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis D-carbamoylase is used for biocatalytic cascade synthesis of D-tryptophan featuring high enantioselectivity 3.5.1.77 N-carbamoyl-D-amino-acid hydrolase synthesis D-NCAase has wide applications particularly in the pharmaceutical industry, since it catalyzes the production of D-amino acids such as D-p-hydroxyphenylglycine (D-HPG), an intermediate in the synthesis of beta-lactam antibiotics, such as penicillins, cephalosporins and amoxicillin 3.5.1.79 Phthalyl amidase synthesis alternative chemical route in removing the phthalimido protecting group 3.5.1.79 Phthalyl amidase synthesis selective deprotection of phthalyl protected amines 3.5.1.79 Phthalyl amidase synthesis the enzyme catalyzes removal of the phthalyl group from a wide variety of phthalyl-containing compounds with improved yields over processes, exhibits stereochemical selectivity, and eliminates the need for harsh conditions to remove the protecting group 3.5.1.79 Phthalyl amidase synthesis one-step synthesis of the antibiotic loracarbef 3.5.1.79 Phthalyl amidase synthesis synthesis of aspartame 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis the enzyme can be useful in production of D-valine 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis development of a facile biotransformation path to perform Michael additions between 1,3-dicarbonyl compounds to methyl vinyl ketone by utilizing promiscuous D-aminoacylase as biocatalyst 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis the aza-Markovnikov addition reactions of 4-nitroimidazole to vinyl acetate catalyzed by D-aminoacylase is up to 1260fold faster than the respective non-enzymatic reaction. New strategy to perform the Markovnikov addition 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis the F191W mutant is considered to be useful for the enzymatic production of D-Trp which is an important building block of some therapeutic drugs 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis N-acyl-D-amino acid amidohydrolases are often used as tools for the optical resolution of D-amino acids, which are important products with applications in industries related to medicine and cosmetics 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis the enzyme can be useful for industrial production of D-amino acids 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis D-aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6 is used for the biotechnological production of D-amino acid from the racemic mixture of N-acyl-DL-amino acids 3.5.1.81 N-Acyl-D-amino-acid deacylase synthesis the D-stereospecific amidohydrolase hydrolyzes the N-acyl-D-amino acid to produce its corresponding D-amino acid, and N-acyl-D-amino acid is left to be used again for chemical racemization. D-amino acids produced by this method include D-Ala, D-Arg, D-Asp, D-Glu, and D-Leu. Commercial process, overview 3.5.1.82 N-acyl-D-glutamate deacylase synthesis the enzyme is used for production of D-glutamate, Co2+ might be a useful additive for stabilization of the enzyme 3.5.1.85 (S)-N-acetyl-1-phenylethylamine hydrolase synthesis economic synthesis of enantiomerically pure (S)-1-phenylethylamine. Enantiomerically pure chiral amines, (S)-enantiomer of 1-phenylethylamine is an example of such compound of high interest, a field of high interest for pharmaceutical industry 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis production of a cell biocatalyst for the production of L-homophenylalanine from D,L-homophenylalanylhydantoin by coexpression of the pydB gene and a thermostable L-N-carbamoylase gene from Bacillus kaustophilus CCRC11223 in Escherichia coli JM109. The expression levels of dihydropyrimidinase and l-N-carbamoylase in the recombinant Escherichia coli cells are estimated to be about 20% of the respective total soluble proteins. When 1% (w/v) isopropyl-beta-D-thiogalactopyranoside-induced cells are used as biocatalysts, a conversion yield of 49% for D,L-homophenylalanylhydantoin with more than 99% enantiomeric excess can be reached in 16 h at pH 7.0 from 10 mM D,L-homophenylalanylhydantoin. The cells can be reused for at least eight cycles at a conversion yield of more than 43%. Coexpression of pydB and lnc in Escherichia coli might be a potential biocatalyst for production of L-homophenylalanylhydantoin 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis to develop a recombinant Escherichia coli whole cell system for the conversion of racemic N-carbamoyl-L-homophenylalanine to L-homophenylalanine, naaar gene from Deinococcus radiodurans and L-N-carbamoylase gene from Bacillus kaustophilus BCRC11223 are cloned and coexpressed in Escherichia coli cells. Recombinant cells treated with 0.5% toluene at 30°C for 30 min exhibit enhanced N-acylamino acid racemase and L-N-carbamoylase activities, which are about 20fold and 60fold, respectively, higher than those of untreated cells. Using toluene-permeabilized recombinant Escherichia coli cells, a maximal productivity of 7.5 mmol L-homophenylalanine/l h with more than 99% yield could be obtained from 150 mmol racemic N-carbamoyl-D-homophenylalanine. Permeabilized cells show considerable stability in the bioconversion process using 10 mmol racemic N-carbamoyl-D-homophenylalanine as substrate, no significantly decrease in conversion yield for L-homophenylalanine is found in the eight cycles 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis a bi-enzyme process for the synthesis of L-homophenylalanine from N-carbamoyl-D-homophenylalanine with immobilized N-acylamino acid racemase and immobilized L-N-carbamoylase. In batch operation, quantitative conversion is achieved. It is a promising alternative for the synthesis of L-homophenylalanine from racemate of N-carbamoyl-DL-homophenylalanine 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis development of a bienzymatic biocatalyst system comprising an N-succinylamino acid racemase from Geobacillus kaustophilus CECT4264 and the enantiospecific L-N-carbamoylase from Geobacillus stearothermophilus CECT43. The biocatalyst system is able to produce optically pure natural and non-natural L-amino acids starting from racemic mixtures of N-acetyl-, N-formyl- and N-carbamoyl-amino acids by dynamic kinetic resolution. The fastest conversion rate is found with N-formyl-aminoacids, followed by N-carbamoyl- and N-acetyl-amino acids, and the an N-succinylamino acid racemase proves to be the limiting step of the system due to its lower specific activity, overview 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis production of different optically pure L-alpha-amino acids starting from different racemic N-formyl- and N-carbamoyl-amino acids using a dynamic kinetic resolution approach with immobilized L-N-carbamoylase and N-succinyl-amino acid racemase as biocatalysts, the system is effective for the biosynthesis of natural and unnatural L-amino acids (enantiomeric excess over 99.5%), overview 3.5.1.87 N-carbamoyl-L-amino-acid hydrolase synthesis the enzyme shows promise as a potential biocatalyst for L-alpha-amino acid production 3.5.1.88 peptide deformylase synthesis the enzyme is an attractive candidate for the biocatalytic deprotection of formylated peptides that are used in chemoenzymatic peptide synthesis 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis enzymatic production of 7-aminocephalosporanate. Construction of site-directed mutants with enhanced activity and stability 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis immobilized mutant enzyme M269Y/C305S may be a promising enzyme in one-step enzymatic production of 7-aminocephalosporanate from cephalosporin C 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis mutant enzyme M269Y may be a promising enzyme in the one-step enzymic production of 7-amino-cephalosporanic acid from cephalosporin C 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis bioconversion of cephalosporin C into 7-aminocephalosporanate 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis D-amino acid oxidase in the permeabilized Pichia pastoris cells and immobilized glutaryl-7-aminocephalosporanic acid acid acylase on support, are employed to convert cephalosporin to 7-aminocephalosporanic acid in a single reactor 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis production of 7-aminocephalosporanate on a large scale at 25°C using enzyme immobilized by epoxide silanization 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis immobilzation on LX-1000EP, an epoxy support, with a recovery of 33.8% and an activity of 81 U/g wet resin, suggesting a potential in industrial 7-aminocephalosporanic acid production. Under optimal conditions, a very high 7-aminocephalosporanic acid yield of 96.6% is obtained within 60 min 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis the enzyme mutant may be suitable for industrial application of the mono-step process for cephalosporin C conversion to (7R)-7-aminocephalosporanate, which is the precursor for production of semisynthetic cephalosporins 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis antibiotic acylases are key enzymes for the industrial production of antibiotic drugs 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis biocatalytic synthesis by the cephalosporin acylase from Pseudomonas sp. strain N176 is a promising alternative to chemical semisynthesis of cephalosporins antibiotics 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis cephalosporin C acylase (CCA), a proton-forming enzyme, is an important industrial enzyme that can directly catalyze the substrate cephalosporin C (CPC) to 7-aminocephalosporanic acid (7-ACA), which is an intermediate in many types of synthetic cephalosporins. Immobilized CCA on porous carriers is applied in industry for the production of 7-ACA because of its high efficiency and environmentally friendly nature compared with the chemical process and two-step enzymatic process involving two enzymes 3.5.1.93 glutaryl-7-aminocephalosporanic-acid acylase synthesis the enzyme is used for production of the important cephalosporin antibiotic 7-aminocephalosporanic acid (7-ACA). 7-ACA is an important cephalosporin nucleus for the synthesis of many widely used beta-lactam antibiotics 3.5.1.100 (R)-amidase synthesis R-amidase is the first enzyme useful for the enzymatic optical resolution of racemic piperazine-2-tert-butylcarboxamide carried out under mild conditions. Enantiomerically pure piperazine-2-carboxylic acid and its tert-butylcarboxamide derivative are important chiral building blocks for some pharmacologically active compounds such as N-methyl-D-aspartate antagonist for glutamate receptor, cardioprotective nucleoside transport blocker, and HIV protease inhibitor 3.5.1.100 (R)-amidase synthesis expression of amidase in Escherichia coli using a T7 promoter. Amidase activity of the engineered Escherichia coli strain reaches 2963 U/l in a 5-l bioreactor, and can be further increased to 5255 U/l in a 100-l bioreactor. Using cell-free extract prepared from 1 kg (wet cell weight) of recombinant cells as catalyst, 60 kg of R,S-DMCPCA is resolved into S-DMCPCA (28.6 kg) and R-2,2-dimethylcyclopropanecarboxylic acid (31.7 kg) in 18 h, and the enantiomeric excess (ee) value of S-DMCPCA reaches 99.32%. 20.5 kg of pure S-DMCPCA is obtained after concentration and crystallization, corresponding to a total yield of 34.2% from R,S-DMCPCA 3.5.1.101 L-proline amide hydrolase synthesis Escherichia coli cells overexpressing the laaA gene have been demonstrated to be applicable to the S-stereoselective hydrolysis of (R,S)-piperazine-2-tert-butylcarboxamide to produce (S)-piperazine-2-carboxylic acid with high optical purity. Enantiomerically pure piperazine-2-carboxylic acid and its tert-butylcarboxamide derivative are important chiral building blocks for some pharmacologically active compounds such as N-methyl-D-aspartate antagonist for glutamate receptor, cardioprotective nucleoside transport blocker and HIV protease inhibitor 3.5.1.105 chitin disaccharide deacetylase synthesis biocatalytic production of glucosamine from N-acetylglucosamine by diacetylchitobiose deacetylase. Although both strains are biocatalytically active, the performance of Bacillus subtilis is 2fold more effective. Establishment of an efficient biotransformation process for the biotechnological production of GlcN in Bacillus subtilis that is more environmentally friendly than the traditional multistep chemical synthesis approach. The overall optimal conditions are 18.6 g/l cells at 40°C, pH 7.5 for 3 h in 3-l bioreactor 3.5.1.105 chitin disaccharide deacetylase synthesis biotransformation of chitin into chitosan through enzymatic deacetylation can be achieved with chitin deacetylases (EC 3.5.1.41, ChDa). Other enzymes involved in chitin and chitosan conversion are chitinases (EC 3.2.1.14) and chitosanases (EC 3.2.1.132). Both of them catalyze the hydrolysis of glycosidic bonds but differ in substrate specificity, hydrolysing bonds of chitin and chitosan, respectively. Obtained chitooligosaccharides can be further enzymatically modified by chitooligosaccharides deacetylases (EC 3.5.1.105, CODa) to obtain products with desired chain arrangement 3.5.1.105 chitin disaccharide deacetylase synthesis Dac is a key enzyme used in the biodegradation of chitin and chitosan to produce chitosan oligosaccharides and monosaccharides 3.5.1.105 chitin disaccharide deacetylase synthesis enzymatic production of defined chitosan oligomers with a specific pattern of acetylation using a combination of chitin oligosaccharide deacetylases. Production of chitosan oligomers by partial chemical or physical depolymerisation of the respective polymers has severe disadvantages. Not only does the production of the oligomers typically involve harsh thermo-chemical treatments or strong physical forces, which may be environmentally unfriendly and/or partially destructive to the oligosaccharides produced, but also the production is highly difficult to control leading to broad heterogeneous mixtures, and the outcome is strongly dependent on the chemical and physical characteristics of the starting material. Partial enzymatic hydrolysis of chitosan polymers using chitosan hydrolysing enzymes such as chitinases or chitosanases with well-defined cleavage specificities has been proposed as an alternative to chemical or physical depolymerisation. But, this attempt is also strongly dependent on the starting material and it, too, leads to the production of heterogeneous mixtures of chitosan oligomers. Still, due to the cleavage specificities of the enzymes, the resulting mixture will be better defined than the chitosan oligomer mixtures obtained by chemical or physical depolymerisation 3.5.1.108 UDP-3-O-acyl-N-acetylglucosamine deacetylase synthesis construction of immobilized metal affinity membrane via coupling of epichlorohydrin, iminodiacetic acid, and nickel ion on a regenerated cellulose membrane. The D-hydantoin-hydrolyzing enzyme harboring a poly-His tagged residue is immobilized on the prepared membrane. By employing a membrane with nickel ion of 155.5 micromol/disc immersed in 0.1 M Tris-HCl buffer pH 8, with 0.8 M sodium chloride, an enzyme activity of 4.2 U/disc is obtained. The immobilized DHTase membrane can achieve a larger pH and thermaltolerance range than the free enzyme. 99% of enzyme activity can be retained after 15 repeated uses 3.5.1.117 6-aminohexanoate-oligomer endohydrolase synthesis bio-based conversion from wastes/byproducts of nylon-6 to adipate (nylon-66 monomer) is possible, when the NADPH generated by the NylE reaction is re-oxidized to NADP+ by coupling a suitable oxidoreductase reaction and the enzyme activity is high enough at a practical level 3.5.1.136 N,N'-diacetylchitobiose non-reducing end deacetylase synthesis a whole-cell biocatalytic process for the environment-friendly synthesis of glucosamine is developed. Biotechnological production of GlcN in Bacillus subtilis is more environmentally friendly than the traditional multistep chemical synthesis approach. It has great a potential for large-scale production of GlcN in an environment-friendly manner 3.5.1.136 N,N'-diacetylchitobiose non-reducing end deacetylase synthesis production of chitosan oligosaccharides and monosaccharides 3.5.2.2 dihydropyrimidinase synthesis - 3.5.2.2 dihydropyrimidinase synthesis artificial fusion enzyme useful as potential biocatalyst for production of nonnatural amino acids 3.5.2.2 dihydropyrimidinase synthesis production of enantiomeric pure nonproteinogenic amino acids, currently under investigation in bioprocess scale for industrial application 3.5.2.2 dihydropyrimidinase synthesis used technically in combination with carbamoylases for production of D-amino acids and non-proteinogenic L-amino acids 3.5.2.2 dihydropyrimidinase synthesis commercial production of optically pure amino acids for synthesis of antibiotics, pharmaceuticals, artificial sweeteners and biologically active peptides, food ingredients, pyrethroids, pesticides, antimicrobial and antiviral agents and other agrochemicals 3.5.2.2 dihydropyrimidinase synthesis thermostable enzyme with great biotechnological potential 3.5.2.2 dihydropyrimidinase synthesis industrial enzyme, that is widely used in the production of D-amino acids which are precursors for semisynthesis of antibiotics, peptides, and pesticides 3.5.2.2 dihydropyrimidinase synthesis production of D-N-carbamoyl amino acid 3.5.2.2 dihydropyrimidinase synthesis encapsulation of a crude cell extract from Agrobacterium radiobacter containing D-hydantoinase and D-carbamoylase activities into alginate-chitosan polyelectrolyte complexes with negligible leakage from the formed capsules. The most suitable biocatalysts are prepared using a chitosan with a medium molecular weight of 600 kDa and a degree of deacetylation of 0.9. For all of the preparation conditions under study, an encapsulation yield of around 60% was achieved and the enzymatic activity yields ranged from 30 to 80% for D-hydantoinase activity and from 40 to 128% for D-carbamoylase activity relative to the activities of the soluble extract. All of the biocatalysts are able to hydrolyze LD-hydroxyphenylhydantoin into p-hydroxyphenylglycine with yields ranging from 30 to 80% 3.5.2.2 dihydropyrimidinase synthesis to produce optically pure D-amino acids, microbial D-hydantoinase is used for stereospecific hydrolysis of chemically synthesized cyclic hydantoins 3.5.2.2 dihydropyrimidinase synthesis the D-hydantoinase is an important enzyme capable of a reversible enantioselective ring-opening hydrolysis of hydantoins being applied for the efficient production of D-form amino acids which are key compounds applicable for the production of antibiotics, peptide hormones, pyrethroids, and pesticides in industry 3.5.2.2 dihydropyrimidinase synthesis recombinant D-PfHYD can potentially be applied in the synthesis of D-amino acids 3.5.2.B2 (+)-gamma-lactamase synthesis enantioselective resolution of 100 g/l 2-azabicyclo[2.2.1]hept-5-en-3-one, is achieved with 10 g/l dry cells, resulting in 55.2% conversion and 99% enantiomeric excess of the (-)-gamma-lactam, i.e. (1R,4S)-2-azabicyclo[2.2.1]hept-5-en-3-one 3.5.2.B2 (+)-gamma-lactamase synthesis the enantiomers of substrate 2-azabicyclo[2.2.1]hept-5-en-3-one (gamma-lactam) are key chiral synthons in the synthesis of antiviral drugs such as carbovir and abacavir 3.5.2.B2 (+)-gamma-lactamase synthesis (+)-gamma-lactamase catalyzes the specific hydrolysis of (+)-gamma-lactam out of the racemic gamma-lactam (2-azabicyclo[2.2.1]hept-5-en-3-one) to leave optically pure (-)-gamma-lactam, which is the key building block of antiviral drugs such as carbovir and abacavir 3.5.2.B2 (+)-gamma-lactamase synthesis mutant enzyme Q192S/L223Y can be employed for the preparation of (-)-gamma-lactam and also for that of (2S,3R)-ethyl 3-phenylglycidate 3.5.2.B2 (+)-gamma-lactamase synthesis the enzyme may become a potential tool for the production of (-)-gamma-lactam because of its superior physicochemical properties and high enzyme activity 3.5.2.B2 (+)-gamma-lactamase synthesis the transformation of an active but unstable mesophilic enzyme into a stable catalyst, achieved by immobilizing the enzyme on macroporous resins, provides a feasible approach for the preparation of optically active (-)-Vince lactam (i.e. 2-azabicyclo [2.2.1] hept-5-en-3-one), which is an important chiral synthon used as an intermediate in organic chemistry 3.5.2.3 dihydroorotase synthesis growth conditions which maintain DHOase overproduction require supplementation of Yeast Carbon Base with asparagines 5 g/l, glucose 20 g/l at pH 5.5. In this medium DHOase activity decreases with time, thus in terms of DHOase yield, the best time for harvesting cells is reached after 30 to 38 h of growth 3.5.2.B3 (-)-gamma-lactamase synthesis Mhg can be utilized to prepare chiral gamma-lactam, which is an important chiral intermediate for the synthesis of a series of antiviral drugs, such as abacavir (targeting HIV) and peramivir (targeting hepatitis and pandemic influenza viruses) 3.5.2.B3 (-)-gamma-lactamase synthesis (+)-gamma-lactamase catalyzes the specific hydrolysis of (+)-gamma-lactam out of the racemic gamma-lactam (2-azabicyclo[2.2.1]hept-5-en-3-one) to leave optically pure (-)-gamma-lactam, which is the key building block of antiviral drugs such as carbovir and abacavir 3.5.2.B3 (-)-gamma-lactamase synthesis bioprocess catalysed by the enzyme is a promising route for the green manufacture of chiral (+)-gamma-lactam 3.5.2.6 beta-lactamase synthesis production of (-)2-azabicyclo[2.2.1]hept-5-en-3-one, which can be converted into carbovir, carbovir is a potent and selective inhibitor of HIV-1 3.5.2.6 beta-lactamase synthesis production of 4-amino-cyclopent-2-enecarboxylic acid, which is used as building block in the synthesis of carbocyclic nucleosides of the natural configuration 3.5.2.11 L-lysine-lactamase synthesis production of D-alpha-amino-beta-caprolactam and L-lysine as nutrient and food supplement 3.5.3.1 arginase synthesis the enzyme can be used to produce L-ornithine from L-arginine 3.5.3.6 arginine deiminase synthesis L-citrulline can be enzymatically produced by arginine deiminase 3.5.3.20 diguanidinobutanase synthesis preparation of pure agmatine sulfate 3.5.3.22 proclavaminate amidinohydrolase synthesis clavulanic acid intermediates: deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid are heterologously produced in Streptomyces venezuelae recombinant using four sets of early genes from the clavulanic acid biosynthetic pathway 3.5.3.22 proclavaminate amidinohydrolase synthesis expression of four structural clavulanic acid biosynthesis genes, which encode carboxyethylarginine synthase (Ceas2), beta-lactam synthetase (Bls2), clavaminate synthase (Cas2), and proclavaminate amidinohydrolase (Pah2), in Streptomyces venezuelae enables production of clavulanic acid intermediates deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid 3.5.4.6 AMP deaminase synthesis the recombinant enzyme is rapidly inactivated below pH 6.0 and at temperatures above 35 °C but under optimal conditions catalysed the highly efficient transformation of AMP to IMP with minimal by-products, suggesting that it could be used for IMP production following protein engineering 3.5.4.25 GTP cyclohydrolase II synthesis the enzyme is essential for industrial riboflavin production by Bacillus subtilis overproducing strains, overview 3.5.5.1 nitrilase synthesis nitrilase products are intermediates in the synthesis of nylon 3.5.5.1 nitrilase synthesis preparing carboxylic acids from readily available nitrile analogues 3.5.5.1 nitrilase synthesis enzyme can be used as a biocatalyst for the synthesis of a range of alpha-hydroxy carboxylic acids or amides from aldehydes in presence of cyanide, halting of the reaction at the amide intermediate by use of specific inhibitors is possible 3.5.5.1 nitrilase synthesis biotransformation of benzonitrile to benzoic acid. The effect of whole cell immobilisation on the biotransformation of benzonitrile and the use of direct electric current for enhanced product removal 3.5.5.1 nitrilase synthesis biotransformation of trans-3-[(5S,6R)-5,6-dihydroxycyclohexa-1,3-dienyl]-acrylonitrile to trans-3-[(5S,6R)-5,6-dihydroxycyclohexa-1,3-dienyl]-acrylic acid with isolated enzyme, immobilized enzyme and with recombinant cells containing AtNIT1 3.5.5.1 nitrilase synthesis production of (R)-mandelic acid from racemic mandelonitrile using free and immobilized cells of Pseudomonas putida 3.5.5.1 nitrilase synthesis attractive as green, mild, and selective catalysts for setting stereogenic centers in fine-chemical synthesis and enantiospecific synthesis of a variety of carboxylic acid derivatives 3.5.5.1 nitrilase synthesis synthesis of (R)-mandelic acid from (R,S)-mandelonitrile by use of Escherichia coli overexpressing the enzyme in a bioreactor. After a single batch reaction, (R)-mandelic acid can be obtained with 87% yield and 99.99% enantiomeric excess 3.5.5.1 nitrilase synthesis synthesis of beta-hydroxy carboxylic acids with high yield and optical purity by reudction of aromatic beta-ketonitriles with recombinant carbonyl reductase and nitrilase-catalyzed hydrolysis 3.5.5.1 nitrilase synthesis nitrilases are useful biocatalysts for the hydrolysis of nitriles in a mild and environmentally friendly manner assuring a clean process and specificity with high yield 3.5.5.1 nitrilase synthesis recombinant cells expressing the enzyme are promising catalysts for the synthesis of stable chiral quaternary carbon centers from ketones 3.5.5.1 nitrilase synthesis the enzyme is useful fpr synthesis of iminodiacetic acid, which is widely used as an intermediate in the manufacture of chelating agents, glyphosate herbicides and surfactants 3.5.5.1 nitrilase synthesis a cascade reaction for the synthesis of optically pure (S)-beta-phenylalanine from benzoylacetonitrile was developed by coupling HpN with an omega-transaminase from Polaromonas sp. JS666 in toluene-water biphasic reaction system using beta-alanine as an amino donor. Various (S)-beta-amino acids can be produced from benzoylacetonitrile derivatives with moderate to high conversions (73-99%) and excellent enantioselectivity (above 99% enantiomeric exess). Great potential of this cascade reaction for the practical synthesis of (S)-beta-phenylalanine 3.5.5.1 nitrilase synthesis mutant nitrilase F168V/T201N/S192F/M191T/F192S is promising in applications for the upscale production of iminodiacetic acid 3.5.5.1 nitrilase synthesis Nitrilase-catalyzed regioselective hydrolysis of 1-cyanocyclohexaneacetonitrile is a green and efficient approach for the preparation of 1-cyanocyclohexaneacetic acid, a key precursor for the synthesis of gabapentin. The enzyme ecapsulated in ethyleneamine-mediated biosilica an be used as a biocatalyst for the development of an efficient biotransformation process for the synthesis of 1-cyanocyclohexaneacetic acid 3.5.5.1 nitrilase synthesis Pseudomonas aeruginosa RZ44 has the potential to be applied in the biotransformation of nitrile compounds 3.5.5.1 nitrilase synthesis the broad range of substrate specificity, the high activity towards aliphatic, aromatic, and arylacetonitriles, and the highly stability, in terms of temperature, pH, and metal ions make the enzyme a promising biocatalyst for mild nitrile hydrolysis 3.5.5.1 nitrilase synthesis the high-level production of Arthrobacter aurescens CYC705 nitrilase will meet the need of industrial biosynthesis of iminodiacetic acid 3.5.5.1 nitrilase synthesis the purified enzyme reveales its selectivity towards dinitriles, which suggests a possible industrial application in the synthesis of cyanocarboxylic acids 3.5.5.1 nitrilase synthesis the strain mut-D3, with higher toxic substrate tolerance and enhanced robustness to extraneous factors is obtained via several rounds of screening. The free or immobilized catalysts of mut-D3 could serve as a good choice for nicotinic acid production from 3-cyanopyridine 3.5.5.1 nitrilase synthesis the use of mutant enzyme P194A/I201A/F202V is feasible for application in the production of (S)-3-(4-chlorophenyl)-4-cyanobutanoic acid en route to (R)-baclofen 3.5.5.5 Arylacetonitrilase synthesis nitrilases are important industrial enzymes that convert nitriles directly into the corresponding carboxylic acids 3.5.5.5 Arylacetonitrilase synthesis recombinant cells expressing the enzyme are promising catalysts for the synthesis of stable chiral quaternary carbon centers from ketones 3.5.5.5 Arylacetonitrilase synthesis the enzyme is promising for the enantioretentive transformation of (S)-mandelonitrile 3.5.5.5 Arylacetonitrilase synthesis the whole cell arylacetonitrilase of Alcaligenes faecalis MTCC 12629 is employed to develop a bioprocess for the synthesis of 4-hydroxyphenylacetic acid from 4-hydroxyphenylacetonitrile. A fed batch process designed at 500 ml with five feedings of substrate results in 150 mM product formation using 18 U/ml whole cells nitrilase (16 mg/ml dcw) at 45°C and a volumetric and catalytic productivity of 23 g/l and 4.13 g/g dcw/h 3.5.5.7 Aliphatic nitrilase synthesis Candida guilliermondii UFMG-Y65 and the isolated enzyme, respectively, might be useful for the bioproduction of amides and acids 3.5.5.7 Aliphatic nitrilase synthesis an efficient, scaleable synthesis of ethyl (R)-4-cyano-3-hydroxybutyrate, a potential intermediate in the synthesis of Atorvastatin (Lipitor) involves nitrilase-catalyzed desymmetrization of 3-hydroxyglutaronitrile 3.5.5.7 Aliphatic nitrilase synthesis biotransformation of acrylonitrile by thermophilic nitrilase from Streptomyces sp. MTCC 7546. Enzyme converts nitriles to acids without the formation of amides. Immobilization of whole cells in agar-agar or beads allows for 25 cycles at 50°C with conversion of 71% of acrylonitrile to acid at 6 h and 100% conversion at 3 h by free cells 3.5.5.7 Aliphatic nitrilase synthesis the enzyme catalyzes the hydrolysis of 3-aminopropionitrile at high substrate concentration up to 3 M, and optimal pH 7.3 and temperature 30°C. However, with the increase of substrate concentration, 3-aminopropanamide is formed, reaching 33% at the substrate concentration of 3 M. A tandem reaction strategy is developed by introducing the aspartate ammonia-lyase-catalyzed amination of fumarate, which utilizes the by-product ammonia as the amino donor in the formation of aspartic acid. Formation of 3-aminopropanamide is significantly inhibited with its amount being reduced from 33% to 3%. The tandem reaction strategy of removing the byproduct ammonia might offer a possibility of producing x02-alanine and L-aspartic acid in one process if the problem in the separation of these two products was solved 3.5.5.7 Aliphatic nitrilase synthesis the enzyme is a potential candidate for industrial applications for biosynthesis of carboxylic acid 3.5.5.7 Aliphatic nitrilase synthesis under optimized conditions, using the fed-batch reaction mode, total of 1050 mM 3-cyanopyridine is hydrolyzed completely in 20.8 h with eight substrate feedings, yielding 129.2 g/l production of nicotinic acid and thus showing a potential for industrial application 3.6.1.1 inorganic diphosphatase synthesis expression in Escherichia coli. Co-expression of tRNA-ARG, cognate for the rare codon AGA in Escherichia coli, improves yield 3.6.1.1 inorganic diphosphatase synthesis an N-acetylhexosamine 1-kinase from Bifidobacterium infantis (NahK_15697), a guanosine 5'-diphosphate (GDP)-mannose pyrophosphorylase from Pyrococcus furiosus (PFManC), and an Escherichia coli inorganic pyrophosphatase (EcPpA) are used efficiently for a one-pot three enzyme synthesis of GDP-mannose, GDP-glucose, their derivatives, and GDP-talose from simple monosaccharides and derivatives in preparative scale 3.6.1.1 inorganic diphosphatase synthesis HvPPA is useful for hydrolyzing diphosphate under conditions of reduced water activity that are a hurdle to current PPA-based technologies 3.6.1.22 NAD+ diphosphatase synthesis preparation of NMN 3.6.1.23 dUTP diphosphatase synthesis can reduce uracil incorporation into DNA 3.6.4.B7 RadA recombinase synthesis efficient generation of hydrazides in proteins by RadA split intein 3.6.4.B10 chaperonin ATPase (protein-folding, protecting from aggregation, protein stabilizing) synthesis ApCpnB has both foldase and holdase activities and can be used as a powerful molecular machinery for the production of recombinant proteins as soluble and active forms in Escherichia coli 3.7.1.3 kynureninase synthesis of inhibitors of kynureninase could prove to be useful in the development of the successful treatment regimen for neurological disorders such as septicemia, AIDS related dementia, Lyme disease, Huntington's and Alzheimer's disease 3.7.1.11 cyclohexane-1,2-dione hydrolase synthesis the enzyme can be used to obtain highly enantioenriched products 3.7.1.12 cobalt-precorrin 5A hydrolase synthesis co-expression of the cobA gene from Propionibacterium freudenreichii and the cbiA, -C, -D, -E, -T, -F, -G, -H, -J, -K, -L, and -P genes from Salmonella enterica serovar typhimurium in Escherichia coli result in the production of cobyrinic acid a,c-diamide 3.7.1.12 cobalt-precorrin 5A hydrolase synthesis synthesis of cobalt corrinoid intermediates cobalt-precorrin 5A and cobalt-precorrin 5B, with the aid of overexpressed enzymes of the vitamin B12 pathway of Salmonella enterica serovar typhimurium 3.8.1.2 (S)-2-haloacid dehalogenase synthesis production of D-lactate in industry 3.8.1.2 (S)-2-haloacid dehalogenase synthesis production of optically active 2-hydroxyalkanoic acids and 2-haloalkanoic acids for chiral synthesis in industry 3.8.1.2 (S)-2-haloacid dehalogenase synthesis His-tagged L-2-haloacid dehalogenase subtype immobilized for production of D-lactate and D-chloropropionic acid 3.8.1.2 (S)-2-haloacid dehalogenase synthesis L-2-haloacid dehalogenase subtype produces D-2-hydroxyalkanoic acids for chiral synthesis in industry in water and organic solvents 3.8.1.2 (S)-2-haloacid dehalogenase synthesis production of (S)-2-chloropropionic acid, which is used in the synthesis of optically active phenoxypropionic acid herbicides 3.8.1.5 haloalkane dehalogenase synthesis production of (R)-3-chloropropane-1,2-diol + 2-oxo-propionaldehyde, which are chiral synthons for various chiral pharmaceuticals, agrochemicals and ferro-electroliquid crystals 3.8.1.5 haloalkane dehalogenase synthesis DhaA produces reaction products (R)- and (S)-2,3-dichloropropan-1-ol, which can be converted to (S)- and (R)-epihydrins, valuable fine chemicals that find application in synthetic routes to several pharmaceutical and healthcare products 3.8.1.5 haloalkane dehalogenase synthesis haloalkane dehalogenases can be used for the production of highly enantioenriched haloalcohols 3.8.1.5 haloalkane dehalogenase synthesis unique catalytic activity and structural stability of DbjA in a broad pH range, combined with high enantioselectivity with particular substrates, make this enzyme a very versatile biocatalyst 3.8.1.5 haloalkane dehalogenase synthesis enzyme DadB and its host, Alcanivorax dieselolei strain B-5, are of potential use for biocatalysis and bioremediation applications 3.8.1.5 haloalkane dehalogenase synthesis enzyme DspA is very enantioselective towards 2-bromobutane and may be attractive for biocatalysis 3.8.1.5 haloalkane dehalogenase synthesis the enzyme might be useful for biocatalytic syntheses in industrial processes 3.8.1.5 haloalkane dehalogenase synthesis the enzymes are of interest for biocatalysis, due to their ability to create enantiomerically pure alcohols 3.8.1.10 2-haloacid dehalogenase (configuration-inverting) synthesis the enzyme is of interest for their potential use in bioremediation and in the synthesis of industrial chemicals 4.1.1.1 pyruvate decarboxylase synthesis synthesis of (R)-phenylacetylcarbinol from cheap substrates in an aqueous reaction system by W392M mutant PDC, alternative strategy to the current fermentative process free of any unwanted by-product 4.1.1.1 pyruvate decarboxylase synthesis Candida utilis PDC is a stable and high productivity enzyme for the production (R)-phenylacetylcarbinol, a pharmaceutical precursor 4.1.1.1 pyruvate decarboxylase synthesis engineered enzyme mutants are useful for synthesis of both enantiomers of alpha-ketols and acetolactates with good enantiomeric excess, overview 4.1.1.1 pyruvate decarboxylase synthesis PDC is useful for the production (R)-phenylacetylcarbinol, a pharmaceutical precursor 4.1.1.1 pyruvate decarboxylase synthesis the enzyme is useful in ethanol production in bacterial coupled systems, overview 4.1.1.1 pyruvate decarboxylase synthesis alpha-ketoisovalerate decarboxylase Kivd from Lactococcus lactis combined with alcohol dehydrogenase Adh3 from Zymomonas mobilis are the optimum candidates for 3-methyl-1-butanol production in Corynebacterium glutamicum. The recombinant strain produces 0.182 g/l of 3-methyl-1-butanol and 0.144 g/l of isobutanol after 12 h of incubation. Further inactivation of the E1 subunit of pyruvate dehydrogenase complex gene (aceE) and lactic dehydrogenase gene (ldh) improves the 3-methyl-1-butanol titer to 0.497 g/l after 12 h of incubation 4.1.1.1 pyruvate decarboxylase synthesis comparison of relevant properties for isobutanol production of Saccharomyces cerevisiae Aro10 and Lactococcus lactis KivD and KdcA genes. Activity in cell extracts reveals a superior Vmax/Km ratio of KdcA for alpha-ketoisovalerate and a wide range of linear and branched-chain 2-oxo acids. KdcA also shows the highest activity with pyruvate which, in engineered strains, can contribute to formation of ethanol as a by-product. During oxygen-limited incubation in the presence of glucose, strains expressing kdcA or kivD show a ca. twofold higher in vivo rate of conversion of alpha-ketoisovalerate into isobutanol than an Aro10-expressing strain. Cell extracts from cultures grown on different nitrogen sources reveal increased activity of constitutively expressed KdcA after growth on both valine and phenylalanine, while KivD and Aro10 activity is only increased after growth on phenylalanine 4.1.1.1 pyruvate decarboxylase synthesis construction of a bypassed pyruvate decarboxylation pathway, through which pyruvate can be converted to acetyl-CoA, by using a coupled enzyme system consisting of pyruvate decarboxylase from Acetobacter pasteurianus and the CoA-acylating aldehyde dehydrogenase from Thermus thermophilus. A cofactor-balanced and CoA-recycling synthetic pathway for N-acetylglutamate production is designed by coupling the bypassed pathway with the glutamate dehydrogenase from Thermus thermophilus and N-acetylglutamate synthase from Thermotoga maritima. N-Acetylglutamate can be produced from an equimolar mixture of pyruvate and alpha-ketoglutarate with a molar yield of 55% through the synthetic pathway consisting of a mixture of four recombinant Escherichia coli strains having either one of the thermostable enzymes. The overall recycling number of CoA is 27 4.1.1.1 pyruvate decarboxylase synthesis construction of a bypassed pyruvate decarboxylation pathway, through which pyruvate can be converted to acetyl-CoA. The coupled enzyme system consists of pyruvate decarboxylase from Acetobacter pasteurianus and the CoA-acylating aldehyde dehydrogenase from Thermus thermophilus. A cofactor-balanced and CoA-recycling synthetic pathway for N-acetylglutamate production is established by coupling the bypassed pathway with the glutamate dehydrogenase from Thermus thermophilus and N-acetylglutamate synthase from Thermotoga maritima. N-Acetylglutamate can be produced from an equimolar mixture of pyruvate and alpha-ketoglutarate with a molar yield of 55% through the synthetic pathway consisting of a mixture of four recombinant Escherichia coli strains having either one of the thermostable enzymes. The overall recycling number of CoA is 27 4.1.1.1 pyruvate decarboxylase synthesis construction of isobutanol production systems by overexpression of effective 2-oxoacid decarboxylase KivD and combinatorial overexpression of valine biosynthetic enzymes in Saccharomyces cerevisiae D452-2. Isobutanol production by the engineered strain is assessed in micro-aerobic batch fermentations using glucose as a sole carbon source, leading to priduction of 93 mg/l isobutanol, which corresponds to a fourfold improvement as compared with the control strain. Isobutanol production is further enhanced to 151 mg/l by additional overexpression of acetolactate synthase Ilv2p, acetohydroxyacid reductoisomerase Ilv5p, and dihydroxyacid dehydratase Ilv3p in the cytosol 4.1.1.1 pyruvate decarboxylase synthesis engineering of Klebsiella pneumoniae to produce 2-butanol from crude glycerol as a sole carbon source by expressing acetolactate synthase (IlvIH), keto-acid reducto-isomerase (IlvC) and dihydroxyacid dehydratase (IlvD) from Klebsiella pneumoniae, and alpha-oxoisovalerate decarboxylase (Kivd) and alcohol dehydrogenase (AdhA) from Lactococcus lactis. The engineered strain produce 2-butanol (160 mg/l) from crude glycerol. Elimination of the 2,3-butanediol pathway by inactivating alpha-acetolactate decarboxylase (Adc) further improves the yield of 2-butanol from 160 to 320 mg/l 4.1.1.1 pyruvate decarboxylase synthesis enhancement of ethanol production capacity of Clostridium thermocellum by transferring pyruvate decarboxylase and alcohol dehydrogenase genes of the homoethanol pathway from Zymomonas mobilis. Both transferring pyruvate decarboxylase and alcohol dehydrogenase are functional in Clostridium thermocellum, but the presence of and alcohol dehydrogenase severely limits the growth of the recombinant strains, irrespective of the presence or absence of the pyruvate decarboxylase gene. The recombinant strain shows two-fold increase in pyruvate carboxylase activity and ethanol production when compared with the wild type strain 4.1.1.1 pyruvate decarboxylase synthesis expression of branched-chain alpha-oxo acid decarboxylase from Lactococcus lactis subsp. lactis KivD and alcohol dehydrogenase from Zymomonas mobilis AdhB in Escherichia coli for higher alcohol production. In LB medium, the resulting strain produces much more 3-methyl-1-butanol (104 mg/l) than isobutanol (24 mg/l). In 5 g/l glucose-containing medium, the production of two alcohols is similar, 156 and 161 mg/l for C4 (isobutanol) and C5 (3-methyl-1-butanol) alcohol, respectively. The increase of glucose content and the adding of alpha-keto acids facilitate the production of C4 and C5 alcohols. The enzyme activities of pure Kivd on alpha-ketoisovalerate and alpha-ketoisocaproate are 26.77 and 21.24 micromol/min and mg, respectively 4.1.1.1 pyruvate decarboxylase synthesis in order to increase production of isobutanol, 2-oxoacid decarboxylase (KDC) and alcohol dehydrogenase (ADH) are expressed in Saccharomyces cerevisiae to enhance the endogenous activity of the Ehrlich pathway. Overexpression Ilv2, which catalyzes the first step in the valine synthetic pathway, and deletion of the PDC1 gene encoding a major pyruvate decarboxylase alters the abundant ethanol flux via pyruvate. Along with modification of culture conditions, the isobutanol titer is elevated 13fold, from 11 mg/l to 143 mg/l, and the yield is 6.6 mg/g glucose 4.1.1.1 pyruvate decarboxylase synthesis modification of enzyme with the N-succinimide ester of an amylose glycylglycine adduct. Upon conjugation, the optimum temperature shifts from 35°C to 40°C, the conjugate shows higher resistance to heat treatment than the native enzyme 4.1.1.1 pyruvate decarboxylase synthesis Ralstonia eutropha H16 produces polyhydroxybutanoate as an intracellular carbon storage material. The excess carbon can be redirected in engineered strains from polyhydroxybutanoate storage to the production of isobutanol and 3-methyl-1-butanol (branched-chain higher alcohols). Strains of Ralstonia eutropha with isobutyraldehyde dehydrogenase activity, in combination with the overexpression of plasmid-borne, native branched-chain amino acid biosynthesis pathway genes and the overexpression of heterologous ketoisovalerate decarboxylase gene, are employed for the biosynthesis of isobutanol and 3-methyl-1-butanol. One mutant strain produces over 180 mg/l branched-chain alcohols in flask culture, and is significantly more tolerant of isobutanol toxicity than wild-type. After the elimination of genes ilvE, bkdAB, and aceE, the production titer improves to 270 mg/l isobutanol and 40 mg/l 3-methyl-1-butanol. Under semicontinuous flask cultivation, the strain grows and produces more than 14 g/l branched-chain alcohols over the duration of 50 days 4.1.1.2 oxalate decarboxylase synthesis biomimetic dyes may prove to be useful technological tools, suitable for the purification 4.1.1.4 acetoacetate decarboxylase synthesis acetoacetate decarboxylase from Clostridium acetobutylicum can act as a biocatalyst for decarboxylation of levulinic acid in an enzymatic system for synthesis of 2-butanone from levulinic acid 4.1.1.5 acetolactate decarboxylase synthesis cloning and expression of the acetolactate decarboxylase gene of Enterobacter aerogenes in Saccharomaces uvarum is carried out to repress the diacetyl formation 4.1.1.5 acetolactate decarboxylase synthesis production of acetoin, which is used in beer fermentation procedures, where diacetyl is produced as by-product and has to be eliminated 4.1.1.7 benzoylformate decarboxylase synthesis enzyme-catalyzed synthesis of enantiomerically pure (R)-benzoin derivatives in good yield 4.1.1.7 benzoylformate decarboxylase synthesis synthesis of enantiopure unsymmetrical and symmetrical 2-hydroxy ketones 4.1.1.7 benzoylformate decarboxylase synthesis purified recombinant His-tagged enzyme immobilized on magnetic nanoparticles, microspheres, nanospheres and ferrofluids, is used for enantioselective synthesis of (S)-2-hydroxyketones. Its stereoselectivity is highly dependent on the structure of the substrate aldehydes. The heterogeneous biocatalyst is offering the ease of immobilization along with the separation steps of metal affinity ligand (Cu2+-iminodiacetic acid) coated magnetic nanoparticles. Reusability of immobilized enzyme for carboligation reactions 4.1.1.7 benzoylformate decarboxylase synthesis the enzyme recombinantly expressed in Escherichia coli is useful for production of ethyl vanillin, i.e. 3-ethoxy-4-hydroxybenzaldehyde, which is a good substitute for vanillin in foods and perfumes since it is 3.5times stronger in flavor and keeps better condition in storage and transport 4.1.1.11 aspartate 1-decarboxylase synthesis the enzyme is used for industrial production of beta-alanine from L-aspartate 4.1.1.12 aspartate 4-decarboxylase synthesis L-Ala is produced much more efficiently at a reduced production cost by the recycling biotransformation reaction system using immobilized Pseudomonas dacunhae cells as compared to the conventional batch reaction system 4.1.1.12 aspartate 4-decarboxylase synthesis production of L-alanine, which is used in infusion solutions and as food additives 4.1.1.12 aspartate 4-decarboxylase synthesis Asd is a major enzyme used in the industrial production of L-alanine 4.1.1.B12 lysine/ornithine carboxy-lyase synthesis the enzyme can potentially be used for synthesis and production of cadaverine. Cadaverine is a promising chemical platform that has a variety of applications, including the production of polyamides, polyurethanes, chelating agents, and additives. In particular, it is relevant for the production of bio-based nylon, which can be used to replace conventional polyamides from petrochemical routes. Whereas Nylon 66 is a polymer composed of hexamethylenediamine and adipic acid, Nylon 56 is a polymer that is produced by the co-polymerization of cadaverine and adipic acid 4.1.1.15 glutamate decarboxylase synthesis expression of enzyme in Bifidobacterium longum in medium containing monosodium glutamate, significant increase in production of gamma-aminobutyric acid 4.1.1.15 glutamate decarboxylase synthesis glutamate decarboxylase from Lactobacillus brevis is a very promising candidate for biosynthesis of gamma-aminobutanoate and various other bulk chemicals that can be derived from it. The enzyme mutant with deletion the C-terminal residues can be be useful in a bioreactor for continuous production of gamma-aminobutanoate 4.1.1.15 glutamate decarboxylase synthesis the enzyme is a key component of 4-aminobutanoate production through an enzymatic process 4.1.1.15 glutamate decarboxylase synthesis the organism is used for production of 4-aminobutanoate, the enzyme is heterologously expressed in Escherichia coli, method optimization, overview 4.1.1.15 glutamate decarboxylase synthesis can be used as an industrially effective tool for the synthesis of gamma-aminobutyric acid 4.1.1.15 glutamate decarboxylase synthesis the unusual high activity of BmGAD at pH 6 makes it an attractive GABA-producing candidate in industrial application 4.1.1.18 lysine decarboxylase synthesis CadA is the conventional cadaverine producer 4.1.1.18 lysine decarboxylase synthesis enzyme AsLdc has a high potential for the industrial production of cadaverine 4.1.1.18 lysine decarboxylase synthesis enzyme LDC plays a crucial role in the synthesis of cadaverine, an important industrial platform chemical. Cadaverine is utilized with a variety of applications such as the production of polyamides, polyurethanes, chelating agents, and additives 4.1.1.18 lysine decarboxylase synthesis the enzyme can be used for industrial production of cadaverine, especially mutant T88S is a promising biocatalyst 4.1.1.18 lysine decarboxylase synthesis the enzyme can be used for synthesis of cadaverine, a biogenic amine, that has the potential to become an important platform chemical for the production of industrial polymers, such as polyamides and polyurethanes 4.1.1.18 lysine decarboxylase synthesis the enzyme can be used for synthesis of cadaverine, a biogenic amine, that has the potential to become an important platform chemical for the production of industrial polymers, such as polyamides, polyurethanes, and nylon. Nylon is one of the most commonly used polymers, and cadaverine has the potential to substitute hexamethylenediamine to produce nylon-5,4, nylon-5,6, nylon-5,10, or nylon-5,12 4.1.1.18 lysine decarboxylase synthesis the enzyme can potentially be used for synthesis and production of cadaverine. Cadaverine is a promising chemical platform that has a variety of applications, including the production of polyamides, polyurethanes, chelating agents, and additives. In particular, it is relevant for the production of bio-based nylon, which can be used to replace conventional polyamides from petrochemical routes. Whereas Nylon 66 is a polymer composed of hexamethylenediamine and adipic acid, Nylon 56 is a polymer that is produced by the co-polymerization of cadaverine and adipic acid 4.1.1.18 lysine decarboxylase synthesis the enzyme EcLdcC can be used for whole-cell biotransformation (a whole-cell biocatalyst) using a constitutive lysine decarboxylase from Escherichia coli for the high-level production of cadaverine from industrial grade L-lysine. It is more effective in comparison to EcCadA. Cadaverine is used for synthesis of bio-polyamides 4.1.1.18 lysine decarboxylase synthesis the immobilized recombinant enzyme CadACLEA can be used as a potential catalyst for efficient production of cadaverine 4.1.1.25 tyrosine decarboxylase synthesis engineering of a salidroside biosynthetic pathway in Rhodiola crenulata hairy roots via metabolic engineering strategy of overexpression. All the transgenic lines show much higher expression levels of tyrosine decaboxylase than non-transgenic one. The transgenic lines produce tyramine, tyrosol and salidroside at higher levels, which are respectively 3.21–6.84, 1.50–2.19 and 1.27–3.47 folds compared with the corresponding compound in non-transgenic lines 4.1.1.28 aromatic-L-amino-acid decarboxylase synthesis preparation of valuable biochemical and diagnostic tools 4.1.1.28 aromatic-L-amino-acid decarboxylase synthesis artemisinic acid can be used as a promising elicitor, especially for the production of therapeutically important indole alkaloids 4.1.1.31 phosphoenolpyruvate carboxylase synthesis multienzyme system composed of recombinant phosphoenolpyruvate, in combination with mesophilic/thermophilic bacterial carbonic anhydrases, for converting CO2 into oxaloacetate. The catalytic procedure is in two steps: the conversion of CO2 into bicarbonate by carbonic anhydrase, followed by the carboxylation of phosphoenolpyruvate with bicarbonate, catalyzed by PEPC, with formation of oxaloacetate. When coupled with the extremely thermostable carbonic anhydrase from Sulphurhydrogenibium azorense, the production of oxaloacetate is achieved even at temperatures up to 60°C 4.1.1.32 phosphoenolpyruvate carboxykinase (GTP) synthesis heterlogous expression in the Escherichia coli dcuD mutant improves hydrogen and ethanol synthesis when grown in a glycerol-based medium. Hydrogen and ethanol specific productions and glycerol consumption increase by 2.46, 1.73 and 1.95 times, respectively, upon the expression 4.1.1.33 diphosphomevalonate decarboxylase synthesis Agrobacterium rhizogenes-mediated transformation and overexpression in Panax ginseng hairy roots leads to 4.4fold higher stigmasterol content and the accumulation of phytosterols and ginsenosides 4.1.1.43 phenylpyruvate decarboxylase synthesis overexpression of phenylpyruvate decarboxylase ARO10 and alcoholdehydrogenase ADH2 genes of Saccharomyces cerevisiae in Kluyveromyces marxianus results in synthesis of 1 g/l 2-phenylethanol 4.1.1.46 o-pyrocatechuate decarboxylase synthesis construction of an artificial pathway for muconic acid production in Escherichia coli by connecting 2,3-dihydroxybenzoic acid biosynthesis with its degradation pathway. Overexpression of entCBA and the key enzymes in the shikimate pathway leads to the production of 900 mg/l of 2,3-dihydroxybenzoic acid. Expression of 2,3-dihydroxybenzoic acid decarboxylase coupled with catechol 1,2-dioxygenase achieves the conversion of 2,3-dihydroxybenzoic acid into muconic acid. Assembly of the total pathway results in the de novo production of muconic acid up to 480 mg/l 4.1.1.46 o-pyrocatechuate decarboxylase synthesis development of an artificial pathway in Escherichia coli for the biosynthesis of muconic acid. Expression of 2,3-dihydroxybenzoate decarboxylase and catechol 1,2-dioxygenase and metabolic optimization by increasing the flux from chorismate through the enterobactin biosynthesis pathway and by regulating the shikimate pathway leads to synthesis of 605.18 mg/l of muconic acid from glucose in a shaking flask culture 4.1.1.50 adenosylmethionine decarboxylase synthesis coexpression of S-adenosylmethionine decarboxylase with chimeric protein KPf, which is made up of the ATPase domain of Escherichia coli DnaK and the substrate binding domain of Plasmodium falciparum Hsp70, and DnaK in Escherichia coli cells. The recombinant protein exhibits improved activity compared to protein coexpressed with overexpressed DnaK 4.1.1.52 6-methylsalicylate decarboxylase synthesis biotechnological de novo production of m-cresol from sugar in complex yeast extract-peptone medium with the yeast Saccharomyces cerevisiae. A heterologous pathway based on the decarboxylation of the polyketide 6-methylsalicylic acid is introduced into a CEN.PK yeast strain. Overexpression of codon-optimized 6-methylsalicylic acid synthase from Penicillium patulum together with activating phosphopantetheinyl transferase npgA from Aspergillus nidulans results in up to 367 mg/l 6-methylsalicylic acid production. Additional genomic integration of the genes have a strongly promoting effect and 6-methylsalicylic acid titers reach more than 2 g/l. Simultaneous expression of 6-methylsalicylic acid decarboxylase patG from Aspergillus clavatus leads to the complete conversion of 6-methylsalicylic acid and production of up to 589 mg/L m-cresol 4.1.1.59 gallate decarboxylase synthesis synthesis of pyrogallol from readily available tannic acid by using the combined action of gallate decarboxylase and tannase in a two-enzyme resting cell bioconversion 4.1.1.61 4-hydroxybenzoate decarboxylase synthesis construction of a phenol synthetic pathway in Escherichia coli via overexpression of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase and chorismate pyruvate lyase. Phenol titer increases 147-fold after modulating the 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase, chorismate pyruvate lyase, and 4-hydroxybenzoate decarboxylase genes in the chromosome. Tributyrin and dibutyl phthalate are the best two solvents for improving phenol production 4.1.1.61 4-hydroxybenzoate decarboxylase synthesis increase in protocatechuate decarboxylase activity in Escherichia coli through enhanced expression of both protocatechuate decarboxylase AroYand kpdB encoding the B-subunit of 4-hydroxybenzoate decarboxylase. This permits expression of protocatechuate decarboxylase activity at a level approximately 14fold greater than the strain with aroY only. The expression level of AroY increases, apparently as a function of the coexpression of AroY and KpdB 4.1.1.63 protocatechuate decarboxylase synthesis engineering of Pseudomonas putida KT2440 to produce muconate from either aromatic molecules or sugars. Coexpression of protocatechuate decarboxylase and associated proteins reduces protocatechuate accumulation and enhances muconate production relative to strains expressing the protocatechuate decarboxylase alone. In bioreactor experiments, coexpression increases the specific productivity of muconate from the lignin monomer p-coumarate by 50% and results in a titer of >15 g/l. In strains engineered to produce muconate from glucose, coexpression more than triples the titer, yield, productivity, and specific productivity, with the best strain producing 4.92±0.48 g/l muconate 4.1.1.63 protocatechuate decarboxylase synthesis increase in protocatechuate decarboxylase activity in Escherichia coli through enhanced expression of both protocatechuate decarboxylase AroYand kpdB encoding the B-subunit of 4-hydroxybenzoate decarboxylase. This permits expression of protocatechuate decarboxylase activity at a level approximately 14fold greater than the strain with aroY only. The expression level of AroY increases, apparently as a function of the coexpression of AroY and KpdB 4.1.1.63 protocatechuate decarboxylase synthesis production of the 4-hydroxybenzoate, protocatechuate, and catechol in Escherichia coli. To enhance endogenous biosynthesis of 4-hydroxybenzoate, native chorismate pyruvate lyase ubiC is overexpressed. 4-Hydroxybenzoate is converted to protocatechuate by hydroxylase pobA from Pseudomonas aeruginosa. Catechol is produced by the additional coexpression of protocatechuate decarboxylase from Enterobacter cloacae. Systematic expression of appropriate pathway elements in phenylalanine overproducing Escherichia coli enables initial titers of 32, 110, and 81 mg/l for 4-hydroxybenzoate, protocatechuate, and catechol, respectively. Disruption of chorismate mutase/prephenate dehydratase (pheA) to preserve endogenous chorismate then allows maximum titers of 277, 454, and 451 mg/l, respectively, at glucose yields of 5.8, 9.7, and 14.3% of their respective theoretical maxima 4.1.1.72 branched-chain-2-oxoacid decarboxylase synthesis the enzyme is able to catalyze carboligation reactions with an exceptionally broad substrate range, a feature that makes KdcA a potentially valuable biocatalyst for C-C bond formation, in particular for the enzymatic synthesis of diversely substituted 2-hydroxyketones with high enantioselectivity 4.1.1.72 branched-chain-2-oxoacid decarboxylase synthesis comparison of relevant properties for isobutanol production of Saccharomyces cerevisiae Aro10 and Lactococcus lactis KivD and KdcA genes. Activity in cell extracts reveals a superior Vmax/Km ratio of KdcA for alpha-ketoisovalerate and a wide range of linear and branched-chain 2-oxo acids. KdcA also shows the highest activity with pyruvate which, in engineered strains, can contribute to formation of ethanol as a by-product. During oxygen-limited incubation in the presence of glucose, strains expressing kdcA or kivD show a ca. twofold higher in vivo rate of conversion of alpha-ketoisovalerate into isobutanol than an Aro10-expressing strain. Cell extracts from cultures grown on different nitrogen sources reveal increased activity of constitutively expressed KdcA after growth on both valine and phenylalanine, while KivD and Aro10 activity is only increased after growth on phenylalanine 4.1.1.72 branched-chain-2-oxoacid decarboxylase synthesis expression of branched-chain 2-oxoacid decarboxylase gene from Lactococcus lactis subsp. lactis CICC 6246 and alcohol dehydrogenase gene from Zymomonas mobilis CICC 41465 in Escherichia coli. Upon incubation in LB medium, much more 3-methyl-1-butanol (104 mg/l) than isobutanol (24 mg/l) is produced. In 5 g/l glucose-containing medium, 156 and 161 mg/l isobutanol and 3-methyl-1-butanol, respectively, is produced 4.1.1.72 branched-chain-2-oxoacid decarboxylase synthesis use of the enzyme for catalytic asymmetric synthesis of (R)-phenylacetylcarbinol derivatives 4.1.1.73 tartrate decarboxylase synthesis production of D-glycerate, which is a useful chiral synthon in synthetic organic chemistry 4.1.1.74 indolepyruvate decarboxylase synthesis engineering of a strain of Corynebacterium glutamicum, based on inactivation of the pyruvate dehydrogenase complex, pyruvate:quinone oxidoreductase, transaminase B, and additional overexpression of the ilvBNCD genes, encoding acetohydroxyacid synthase, acetohydroxyacid isomeroreductase, and dihydroxyacid dehydratase, for the production of isobutanol from glucose under oxygen deprivation conditions by inactivation of L-lactate and malate dehydrogenases, implementation of ketoacid decarboxylase from Lactococcus lactis, alcohol dehydrogenase 2 (ADH2) from Saccharomyces cerevisiae, and expression of the pntAB transhydrogenase genes from Escherichia coli. The resulting strain produces isobutanol with a substrate-specific yield (YP/S) of 0.60 mol per mol of glucose. Chromosomally encoded alcohol dehydrogenase AdhA rather than the plasmid-encoded ADH2 from S. cerevisiae is involved in isobutanol formation, and overexpression of the corresponding AdhA gene increases the YP/S to 0.77 mol of isobutanol per mol of glucose. Inactivation of the malic enzyme significantly reduces the YP/S, indicating that the metabolic cycle consisting of pyruvate and/or phosphoenolpyruvate carboxylase, malate dehydrogenase, and malic enzyme is responsible for the conversion of NADH + H+ to NADPH + H+. In fed-batch fermentations with an aerobic growth phase and an oxygen-depleted production phase, the most promising strain produces about 175 mM isobutanol, with a volumetric productivity of 4.4 mM per h, and shows an overall YP/S of about 0.48 mol per mol of glucose in the production phase 4.1.1.74 indolepyruvate decarboxylase synthesis engineering of Clostridium thermocellum to produce isobutanol. Both the native 2-oxoisovalerate-oxidoreductase KOR, EC 1.2.7.7, and the heterologous Lactococcus lactis 2-oxoisovalerate decarboxylase KIVD, EC 4.1.1.74, expressed are responsible for isobutanol production.The plasmid is integrated into the chromosome by single crossover. The resulting strain is stable without antibiotic selection pressure and produces 5.4g/l of isobutanol from cellulose in minimal medium at 50°C within 75 h, corresponding to 41% of theoretical yield 4.1.1.76 arylmalonate decarboxylase synthesis asymmetric synthesis of both enantiomers of [alpha-2H]phenylacetic acid 4.1.1.76 arylmalonate decarboxylase synthesis synthesis of enantiomeric pure (R)-alpha-fluorophenylacetic acid 4.1.1.76 arylmalonate decarboxylase synthesis synthesis of the anti-cancer drug (R)-flurbiprofen 4.1.1.76 arylmalonate decarboxylase synthesis by immobilization on an amino C2 acrylate carrier the operational stability of the (S)-selective AMDase variant G74C/M159L/C188G/V43I/A125P/V156L is 158fold increased to a half-life of about 8.6 days. Further optimization is achieved by simple immobilization of the cell lysate 4.1.1.77 2-oxo-3-hexenedioate decarboxylase synthesis expression and purification of a monomeric, stable and active enzyme in the absence of vinylpyruvate hydratase 4.1.1.90 peptidyl-glutamate 4-carboxylase synthesis development of expression vectors which enable expressing mammalian gamma-glutamyl carboxylase, vitamin K epoxide reductase complex, subunit 1, and/or PDIA2 along with human coagulation factor VII in Drosophila Schnieder S2 cells. Mammalian GGCX i indispensable to synthesize active factor VII while mammalian vitamin K epoxide reductase complex, subunit 1, and PDIA2 are not critical but supportive factors for S2 cells 4.1.1.91 salicylate decarboxylase synthesis selective and ecological production by carboxylation of phenol to form salicylate, the enzymatic Kolbe–Schmitt reaction 4.1.1.99 phosphomevalonate decarboxylase synthesis isopentenol production in Escherichia coli by utilizing phosphomevalonate decarboxylase and Escherchia coli-endogenous phosphatase AphA. The enzymes bypass the isopentenyl diphosphate mevalonate pathways, have reduced energetic requirements, are further decoupled from intrinsic regulation, and are free from isopentenyl diphosphate-related toxicity. Reduced aeration rate has less impact on the bypass pathway than the original mevalonate pathway. The performance of the bypass pathway is primarily determined by the activity of phosphomevalonate decarboxylase 4.1.1.102 phenacrylate decarboxylase synthesis synthesis of vanillin by use of ferulic acid decarboxylase Fdc from Bacillus pumilus and 4-vinylguaiacol oxygenase Cso2 from Caulobacter segnis. In the first stage, Escherichia coli cells expressing Fdc rapidly decarboxylate ferulic acid and completely convert 75 mM of this substrate to 4-vinylguaiacol within 2 h at pH 9.0. In the second stage, Escherichia coli cells expressing Cso2 efficiently oxidize 4-vinylguaiacol to vanillin. The concentration of vanillin reaches 52 mM (7.8 g/l) in 24 h 4.1.1.102 phenacrylate decarboxylase synthesis by expressing Rhodobacter sphaeroides tyrosine ammonia lyase, in Streptomyces mobaraense, which permits the synthesis of p-coumaric acid from glucose, a strain is obtained that produces high amounts of 4-vinylphenol 4.1.1.102 phenacrylate decarboxylase synthesis for 4-vinylphenol production directly from cellulose, L-tyrosine ammonia lyase derived from Rhodobacter sphaeroides and phenolic acid decarboxylase from Streptomyces sviceus are introduced into endoglucanase-secreting Streptomyces lividans, and the 4-vinylphenol biosynthetic pathway is constructed therein. The created transformants successfully produce 4-vinylphenol directly from cellulose 4.1.1.102 phenacrylate decarboxylase synthesis styrene production from biomass-derived carbon sources, by culture of Streptomyces lividans expressing FDC1 together with Streptomyces lividans/p-encP, which produces trans-cinnamic acid. The coculture system combined with the recovery of styrene using polystyrene resin beads XAD-4 allows the production of styrene from glucose, cellobiose, or xylooligosaccharides, respectively 4.1.1.102 phenacrylate decarboxylase synthesis 4-hydroxystilbene is synthesized from p-coumaric acid in four parallel continuous flow reactors, using a 3D printing process with agarose bioinks, and a subsequent palladium(II) acetate-catalysed Heck reaction, with a total yield of 14.7% on a milligram scale. The enzyme shows 38% residual activity after the printing process 4.1.1.103 gamma-resorcylate decarboxylase synthesis enzyme catalyzes the regioselective carboxylation of phenol, 1,2-dihydroxybenzene and 1,3-dihydroxybenzen to 4-hydroxybenzoate, 2,3-dihydroxybenzoate, and 2,6-dihydroxybenzoate, respectively. When the efficient production of 2,6-dihydroxybenzoate is optimized using whole cells of Pandoraea sp. 12B-2, the productivity of 2,6-dihydroxybenzoate tops out at 1.43 M. No formation of any other products is observed after the carboxylation reaction 4.1.1.112 oxaloacetate decarboxylase synthesis optimization of oxaloacetate may increase lysine productivity of commercially used strains 4.1.1.117 2-[(L-alanin-3-ylcarbamoyl)methyl]-2-hydroxybutanedioate decarboxylase synthesis the SbnCEF synthetases and decarboxylase SbnH are necessary and sufficient to produce staphyloferrin B in reactions containing component substrates L-2,3-diaminopropionic acid, citric acid and 2-oxoglutaric acid. 1,2-diaminoethane is not required, this component arises from the SbnH-dependent decarboxylation of a 2,3-diaminoproprionic acid-containing intermediate 4.1.2.4 deoxyribose-phosphate aldolase synthesis stereocontrolled aldol condensation of 3-hydroxy aldehydes and ketones 4.1.2.4 deoxyribose-phosphate aldolase synthesis synthesis of a variety of sugar analogs, including 2-deoxy-L-fucose and analogs, thiosugars and glycolipid precursors. The sequential aldol reaction is utilized in the synthesis of a variety of 2,4-dideoxyhexoses and 2,4,6-trideoxyhexoses 4.1.2.4 deoxyribose-phosphate aldolase synthesis mutant D-2-deoxyribose-5-phosphate aldolase Ser238Asp is used to prepare beta-hydroxy-delta lactol synthons and tert-butyl [(4R,6R)-6-aminoethyl-2,2-dimethyl-1,3-dioxn-4-yl]acetate, a key intermediate for atorvastatin synthesis 4.1.2.4 deoxyribose-phosphate aldolase synthesis production of enzyme using a continuous lactose induction strategy. The lactose concentration in the feed medium affects directly the expression of the target protein. The use of 50 g/L in the feed medium results in an expression level of above 30%, and the maximum final enzyme concentration of 16200 U/l. The acetate concentration remains at a low level in the fed-batch phase, less than 0.5 g/l 4.1.2.4 deoxyribose-phosphate aldolase synthesis biosynthesis of 2-deoxysugars using whole-cell catalyst expressing 2-deoxy-D-ribose 5-phosphate aldolase. A whole-cell transformation strategy using resting cells of the BL21(pKDERA12) strain, containing the expressed plasmid pKDERA12 (S238D/F200I/DELTAY259), results in increase in 2-deoxy-D-ribose yield from 0.41 mol/mol D-glyceraldehyde to 0.81 mol/mol D-glyceraldehyde and higher substrate tolerance from 0.5 to 3 M compared to in vitro assays. With further optimization of the transformation process, the BL21(pKDERA12) strain produces 2.14 M (287.06 g/l) 2-deoxy-D-ribose, with a yield of 0.71 mol/mol D-glyceraldehyde and average productivity of 0.13 mol/l*h (17.94 g/l*h). The results demonstrate the potential for large-scale production of 2-deoxy-D-ribose using the BL21(pKDERA12) strain. Furthermore, the BL21(pKDERA12) strain also exhibits the ability to efficiently produce 2-deoxy-D-altrose from D-erythrose, as well as 2-deoxy-L-xylose and 2-deoxy-L-ribose from L-glyceraldehyde 4.1.2.4 deoxyribose-phosphate aldolase synthesis industrial application of the enzyme for the synthesis of a key building block for the pharmaceutical blockbuster atorvastatin. The main drawback of the enzyme being used in such a process is its sensitivity to industrially relevant concentrations of the substrate and the occurrence of product inhibition. Product inhibition can be avoided by coupling the catalytic transformation with transport processes, which, in turn, would require an immobilization of the enzyme within a thin film that can be deposited on a membrane support. A fabrication process for such films is developed that is based on the formation of DERA-poly(N-isopropylacrylamide) conjugates that are subsequently allowed to self-assemble at an air-water interface to yield the respective film 4.1.2.4 deoxyribose-phosphate aldolase synthesis synthesis of statin intermediates. The recombinant enzyme can be potentially applied in the production of (3R,5S)-6-chloro-2,4,6-trideoxy-erythro-hexose. The bioconversion process for production of (3R,5S)-6-chloro-2,4,6-trideoxy-erythro-hexose from chloroacetaldehyde and acetaldehyde using the recombinant enzyme is studied and this process took 3 h for maximal conversion 4.1.2.4 deoxyribose-phosphate aldolase synthesis the enzyme is an interesting candidate for bio-catalysis of carbo-ligation reactions, which are central to synthetic chemistry 4.1.2.5 L-threonine aldolase synthesis threonine aldolase is a very promising enzyme that can be used to prepare biologically active compounds or building blocks for pharmaceutical industry. Rational design is applied to thermophilic threonine aldolase from Thermotoga maritima to improve thermal stability by the incorporation of salt and disulfide bridges between subunits in the functional tetramer 4.1.2.9 phosphoketolase synthesis a phosphoketolase disruption mutant harboring the pXylRAB gene for catabolism of xylose lacks the phosphoketolasepathwas pathway and produces predominantly lactic acid from xylose via the pentose phosphate pathway, although its fermentation rate slightly decreases. Further introduction of the transketolase gene to disrupted phosphoketolase locus leads to restoration of the fermentation rate. As a result, the strain produces 50.1 g/l of L-lactic acid from xylose with a optical purity of 99.6% and a yield of 1.58 mol per mole xylose consumed 4.1.2.9 phosphoketolase synthesis metabolic engineering strategy to express the fungal genes of the phosphoketolase pathway in Saccharomyces cerevisiae. The utilization of the phosphoketolase pathway does not interfere with glucose assimilation through the Embden-Meyerhof-Parnas pathway and the expression of this route can contribute to increase the acetyl CoA supply 4.1.2.9 phosphoketolase synthesis given that acetyl-CoA is a key intermediate in several biosynthetic pathways, phosphoketolase overexpression offers a viable strategy to enhance the economics of an array of biological methane conversion processes 4.1.2.9 phosphoketolase synthesis expression of mutant T2A/I6T/H260 in Corynebacterium glutamicum Z188 results in 16.67% and 18.19% improvement in L-glutamate titer and yield, respectively, compared with the wildtype 4.1.2.10 (R)-mandelonitrile lyase synthesis used as a catalyst in the preparation of optically active cyanohydrins 4.1.2.10 (R)-mandelonitrile lyase synthesis (R)-oxynitrilase immobilized as a cross-linked enzyme aggregate via precipitation with 1,2-dimethoxyethane and subsequent cross-linking using glutaraldehyde is stable and recyclable. Application in microaqueous medium, superior biocatalyst for the enantioselective hydrocyanation of slow-reacting aldehydes 4.1.2.10 (R)-mandelonitrile lyase synthesis development and validation of a process model for production of (R)-cyanohydrins in an aqueous-organic biphasic-stirred tank reactor with an unknown interfacial area operated in batch mode. The formation of (R)-mandelonitrile from benzaldehyde and cyanide catalyzed by Prunus amygdalus hydroxynitrile lyase is chosen as a model reaction. Methyl tert-butyl ether is selected as the organic solvent and the reaction conditions are 5°C and pH 5.5 at which the nonenzymatic reaction towards rac-mandelonitrile is largely suppressed 4.1.2.10 (R)-mandelonitrile lyase synthesis hydroxynitrile lyases are proficient biocatalysts for the stereospecific synthesis of cyanohydrins 4.1.2.10 (R)-mandelonitrile lyase synthesis optically active cyanohydrins are expedient starting materials for preparation of alpha-hydroxyketones, alpha-hydroxy acids, beta-aminoalcohols, aminonitriles and aziridines 4.1.2.10 (R)-mandelonitrile lyase synthesis optically active cyanoydrins are expedient starting materials for preparation of alpha-hydroxyketones, alpha-hydroxy acids, beta-aminoalcohols, aminonitriles and aziridines 4.1.2.10 (R)-mandelonitrile lyase synthesis production of (R)-hydroxypivalaldehyde cyanohydrin, the chiral key precursor in hydroxynitrile lyase based (R)-pantlactone synthesis 4.1.2.10 (R)-mandelonitrile lyase synthesis the enantiospecific formation of alpha-hydroxynitriles 4.1.2.10 (R)-mandelonitrile lyase synthesis fusion of enzyme to different fluorescent reporter proteins. Flavin-based fluorescent reporter fusions convert 95% benzaldehyde to (R)-mandelonitrile within 60 min at pH 4.75, with 96% enantiomeric excess 4.1.2.10 (R)-mandelonitrile lyase synthesis purification of enzyme as crosslinked enzyme aggregates and application for synthesis of enantiopure cyanohydrins 4.1.2.10 (R)-mandelonitrile lyase synthesis synthesis of (R)-beta-nitro alcohols using nitromethane as donor. Highest enantioselectivity is obtained with n-butyl acetate as solvent with an optimum aqueous phase content of 50% (v/v) 4.1.2.10 (R)-mandelonitrile lyase synthesis Amygdalus pedunculata hydroxynitrile lyase is an excellent biocatalyst and has very high potential for synthesis of enantiopure cyanohydrins 4.1.2.10 (R)-mandelonitrile lyase synthesis an easy one-step immobilization of a complex Arabidopsis thaliana hydroxynitrile lyase fusion protein is possible starting from an Escherichia coli crude cell extract (producing a carbohydrate-binding module-containing fusion protein) and cost efficient cellulosic carrier materials. Highly-specific site-selective immobilization of the target protein is achieved by fusing the family 2 carbohydrate-binding module of the exoglucanase/xylanase Cex from Cellulomonas fimi to the target enzyme. This yields a cheap, active, stable and recyclable immobilizate, which can be employed in micro-aqueous reaction systems to enable enantiopure cyanohydrin synthesis 4.1.2.10 (R)-mandelonitrile lyase synthesis hydroxynitrile lyases are involved in the synthesis of enantiomerically pure cyanohydrins which are important intermediates in the production of pharmaceuticals and agrochemicals. The enzyme synthesizes (R)-mandelonitrile in both, batch reaction and fed-batch reaction and can be effectively used in the synthesis of (R)-mandelonitrile 4.1.2.10 (R)-mandelonitrile lyase synthesis the enzyme has very high potential for synthesis of cyanohydrins and can be used for the production of enantiopure cyanohydrins. Cyanohydrins are important intermediates in the production of pharmaceuticals and agrochemicals 4.1.2.10 (R)-mandelonitrile lyase synthesis the enzyme is a powerful and cheap biocatalyst in the synthesis of (R)-mandelonitrile and may be used in combination with nitrilases to produce enantiopure mandelic acids 4.1.2.10 (R)-mandelonitrile lyase synthesis the enzyme may find application as a biocatalyst for the synthesis of enantiomerically pure cyanohydrins. DtHNL1-CLEA preparation results in a more robust biocatalyst under acidic conditions, which is an interesting feature for the efficient production of enantiomerically pure cyanohydrins 4.1.2.10 (R)-mandelonitrile lyase synthesis the high specific activity toward benzaldehyde along with wide temperature, pH stabilities and high enantioselectivity in the synthesis of various cyanohydrins make this enzyme suitable as an industrial biocatalyst 4.1.2.10 (R)-mandelonitrile lyase synthesis the immobilization of partially purified hydroxynitrile lyase from Prunus dulcis as crosslinked enzyme aggregate is a very promising tool owing to its ease of preparation, cheapness, and efficiency in the preparation of enantiopure cyanohydrins. Response surface methodology provides an effective way for the optimization of the preparation of PdHNL-crosslinked enzyme aggregate. The synthesis of (R)-mandelonitrile, (R)-2-chloromandelonitrile, (R)-3,4-dihydroxymandelonitrile, (R)-2-hydroxy-4-phenyl butyronitrile, (R)-4-bromomandelonitrile, (R)-4-fluoromandelonitrile, and (R)-4-nitromandelonitrile is achieved with high yield and enantiomeric excess 4.1.2.11 hydroxymandelonitrile lyase synthesis biocatalyst for the enantiospecific addition of hydrogen cyanide to aldehydes in organic solvents 4.1.2.11 hydroxymandelonitrile lyase synthesis first (S)-HNL used in organic solvents for the preparation of (S)-cyanohydrins 4.1.2.13 fructose-bisphosphate aldolase synthesis enzyme FruA has a flexible substrate specificity and accepts various aldehydes. It is applcable for synthesis of a number of ketoses using simple aliphatic aldehydes as acceptors in a one-pot reaction system. The yields are quite similar for reactions of which acetaldehyde, propionaldehyde, and butyraldehyde are acceptors 4.1.2.13 fructose-bisphosphate aldolase synthesis tea oil yield can be improved by enhanced expression of fructose-1,6-bisphosphate aldolase and stearoyl-ACP desaturase genes in transgenic plants 4.1.2.14 2-dehydro-3-deoxy-phosphogluconate aldolase synthesis application of a KDPG-aldolase gene-dependent addiction system for enhanced production of cyanophycin in Ralstonia eutropha strain H16 4.1.2.14 2-dehydro-3-deoxy-phosphogluconate aldolase synthesis Methylophilus methylotrophus AS1 is choosen as a host strain to produce L-lysine from methanol 4.1.2.14 2-dehydro-3-deoxy-phosphogluconate aldolase synthesis stereospecific formation of carbon-carbon bonds is one of the major interests in organic synthetic chemistry, aldolases are part of the most important group of asymmetric C-C-bonding enzymes 4.1.2.14 2-dehydro-3-deoxy-phosphogluconate aldolase synthesis use of a recombinant strain of Ralstonia eutropha H16 presenting a 2-keto-3-desoxy-phosphogluconate aldolase gene-dependent catabolic addiction system for plasmid maintenance when using gluconate or fructose as sole carbon source for synthesis of cyanophycin granule polypeptides. Maximum contents of water-insoluble cyanophycin granule polypeptide and water-soluble cyanophycin granule polypeptide, contributing to 47.5% and 5.8% (w/w) of cell dry matter, respectively, are obtained at the 30-L scale cultivation under optimum conditions 4.1.2.17 L-fuculose-phosphate aldolase synthesis a multienzyme system composed by recombinant dihydroxyacetone kinase from Citrobacter freundii, fuculose-1-phosphate aldolase from Escherichia coli and acetate kinase, allows a practical one-pot C-C bond formation catalyzed by dihydroxyacetone phosphate-dependent aldolases from dihydroxyacetone and an aldehyde 4.1.2.17 L-fuculose-phosphate aldolase synthesis expression of fuculose-1-phosphate aldolase FucA in Escherichia coli as active inclusion bodies in batch cultures and modulation of the activity of insoluble FucA by a proper selection of producing strain, culture media, and process conditions. In an optimized defined medium, FucA inclusion bodies are more active (in terms of specific activity) than the soluble protein version obtained in the same process with a conventional defined medium. Either directly or immobilized into LentikatVR beads, FucA gives product yields ranging from 65 to 76% in an aldolic reaction between DHAP and (S)-Cbz-alaninal 4.1.2.19 rhamnulose-1-phosphate aldolase synthesis highly efficient enzymatic synthesis of L-fructose based on rhamnulose-1-phosphate aldolase and acid phosphatase using racemic glyceraldehyde and dihydroxyacetone phosphate as substrates. Enantiomerically pure L-fructose is obtained 4.1.2.19 rhamnulose-1-phosphate aldolase synthesis use as model enzyme for the immobilization onto gold nanoparticles and application as biocatalyst of the aldol addition reaction between dihydroxyacetone phosphate and (S)-Cbz-alaninal during two reaction cycles. In these conditions, an improved reaction yield and selectivity, together with a fourfold activity enhancement are observed 4.1.2.21 2-dehydro-3-deoxy-6-phosphogalactonate aldolase synthesis the enzyme can stereospecifically catalyze aldol addition with unnatural electrophilic substrates at rates practical for preparative scale organic synthesis 4.1.2.28 2-dehydro-3-deoxy-D-pentonate aldolase synthesis biosynthesis route for ethylene glycol production from D-xylose, consisting of conversion steps D-xylose to D-xylonate to 2-dehydro-3-deoxy-D-pentonate to glycoaldehyde to ethylene glycol. Implementation of the respective enzymes, D-xylose dehydrogenase, D-xylonate dehydratase, 2-dehydro-3-deoxy-D-pentonate aldolase, and glycoaldehyde reductase, in a metabolically engineered Escherichia coli, in which the D-xylose to D-xylulose reaction is prevented by disrupting the D-xylose isomerase gene, leads to a construct producing 11.7 g/l of ethylene glycol from 40.0 g/l of D-xylose 4.1.2.38 benzoin aldolase synthesis application of benzaldehyde lyase as a heterogeneous catalyst in the continuous synthesis (in a packed bed in a continously operated plug flow reactor for over 140 h) of the chiral 2-hydroxy ketone (R)-2-hydroxy-1-phenyl-propanone 4.1.2.38 benzoin aldolase synthesis stereoselective synthesis of novel benzoins catalysed by benzaldehyde lyase in a gel-stabilised two-phase system. (R)-1,2-di(3-furanyl)-2-hydroxyethanone, (R)-2-hydroxy-1,2-di(3-thienyl) ethanone, (R)-1,2-di(4-ethoxyphenyl)-2-hydroxyethanone, (R)-1,2-di(3-ethoxyphenyl)-2-hydroxyethanone, (R)-2-hydroxy-1,2-di(3-tolyl)ethanone, and (R)-1,2-di(benzofuran-2-yl)-2-hydroxyethanone are prepared with yields up to 31.8% and enantiomeric excess of more than 99% 4.1.2.38 benzoin aldolase synthesis the enzyme is a tool in chemoenzymatic synthesis of chiral alpha-hydroxyketones 4.1.2.38 benzoin aldolase synthesis (R)-benzoin is synthesized from benzaldehyde using the most stable variant immobilized in packed-bed reactors via the SpyCatcher/SpyTag system. Over a period of seven days, (R)-benzoin is produced with a stable spacetime-yield of 9.3 mmol/l and day 4.1.2.42 D-threonine aldolase synthesis alanine racemase with engineered function of D-threonine aldolase, capable of synthesizing beta-hydroxy-alpha-amino acids, stereoselectivity is comparable to that of D-threonine aldolase 4.1.2.42 D-threonine aldolase synthesis efficient, environmentally friendly process for the production of (2R,3S)-2-amino-3-hydroxy-3-(pyridin-4-yl)-propanoic acid by a recombinant D-threonine aldolase catalyzed aldol addition of glycine and pyridine 4-carboxaldehyde. (2R,3S)-2-amino-3-hydroxy-3-(pyridin-4-yl)-propanoic acid, is a key intermediate in the synthesis of the (2R,3S)-2-amino-3-hydroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one, a developmental drug candidate. The aldol addition product directly crystallizes out from the reaction mixture in high purity and high diastereo- and enantioselectivity, contributing to high yield and allowing easy isolation, processing, and downstream utilization 4.1.2.42 D-threonine aldolase synthesis the enzyme can be used for the asymmetric synthesis alpha-quaternary alpha-amino acids 4.1.2.42 D-threonine aldolase synthesis the enzyme has a considerable potential in biocatalysis for the stereospecific synthesis of various beta-hydroxy amino acids, which are valuable building blocks for the production of pharmaceuticals 4.1.2.42 D-threonine aldolase synthesis the enzyme is a powerful tool for the stereospecific synthesis of various beta-hydroxy amino acids in synthetic organic chemistry 4.1.2.42 D-threonine aldolase synthesis the enzyme might be a promising biocatalyst for producing chiral aromatic beta-hydroxy-alpha-amino acids 4.1.2.43 3-hexulose-6-phosphate synthase synthesis enzymatic preparation is suitable for the synthesis of sugars labeled with 13C at specific positions, enzymatic preparation of [1-13C]D-fructose-6-phosphate by using D-ribose-5 phosphate and [1-13C]-formaldehyde as substrates 4.1.2.46 aliphatic (R)-hydroxynitrile lyase synthesis optically active aliphatic omega-hydroxycyanohydrins are valued materials in organic synthesis 4.1.2.46 aliphatic (R)-hydroxynitrile lyase synthesis production of organosilicon compounds by enantioselective transcyanation. Under optimum conditions, both acetyltrimethylsilane conversion to (R)-2-trimethylsilyl-2-hydroxy-ethylcyanide and enantiomeric excess of the product are above 99%. the silicon atom in acetyltrimethylsilane has a great effect on the eaction and both the substrate conversion and the product enantiomeric excess are much higher than those in its carbon counterpart 3,3-dimethyl-2-butanone 4.1.2.47 (S)-hydroxynitrile lyase synthesis of aliphatic and aromatic cyanohydrins 4.1.2.47 (S)-hydroxynitrile lyase synthesis biocatalyst for the enantiospecific addition of hydrogen cyanide to aldehydes in organic solvents 4.1.2.47 (S)-hydroxynitrile lyase synthesis catalyzes the industrially interesting formation of (S)-cyanohydrins from aldehydes or ketones and HCN 4.1.2.47 (S)-hydroxynitrile lyase synthesis enzyme-catalyzed preparation of optically active cyanohydrins 4.1.2.47 (S)-hydroxynitrile lyase synthesis industrial important biocatalyst used in enantiospecific syntheses of alpha-hydroxynitriles from aldehydes and methyl-ketones 4.1.2.47 (S)-hydroxynitrile lyase synthesis production of chiral ferrocene derivatives as ligands in asymmetric catalysis, to bioelectrochemistry and development of new pharmaceuticals against malaria 4.1.2.47 (S)-hydroxynitrile lyase synthesis (S)-mandelonitrile production on a commercial scale 4.1.2.47 (S)-hydroxynitrile lyase synthesis crude cell lysate containing a hydroxynitrile lyase can be used for the enantioselective synthesis of several cyanohydrins in a microchannel. These enzymatic reactions show a high initial reaction rate and enantioselectivity, which in a batchwise process can only be achieved by vigorous stirring 4.1.2.47 (S)-hydroxynitrile lyase synthesis industrially relevant C-C-coupling reaction 4.1.2.47 (S)-hydroxynitrile lyase synthesis recombinant Pichia pastoris strains are constructed which simultaneously express the (S)-oxynitrilase of Manihot esculenta and the arylacetonitrilase of Pseudomonas fluorescens EBC191 each under the control of individual AOX1 promoters in order to obtain a whole cell catalyst for the synthesis of (S)-mandelic acid from benzaldehyde and cyanideproduction of optically active cyanohydrin compounds 4.1.2.47 (S)-hydroxynitrile lyase synthesis synthesis of enantiopure (S)-3-phenoxybenzaldehyde cyanohydrin (a useful intermediate in the pyrethroid synthesis) by applying a high pH two-phase system to reduce nonenzymatic reaction 4.1.2.47 (S)-hydroxynitrile lyase synthesis synthesis of optically active cyanohydrins that are interesting intermediates for the synthesis of alpha-hydroxy acids, alpha-hydroxy ketones, or beta-ethanolamines, all of which are important building blocks in organic synthesis 4.1.2.47 (S)-hydroxynitrile lyase synthesis the cyanohydrin reaction of formylferrocene catalysed by the hydroxynitrile lyase from Hevea brasiliensis provides an access to chiral ferrocene derivatives in high enantiomeric excess. Since cyanohydrins are versatile synthetic intermediates, the possibility for many preparative transformations is opened. This synthetic potential is enlarged even further with the transformation of 1,1'-diformylferrocene leading to biscyanohydrins 4.1.2.47 (S)-hydroxynitrile lyase synthesis the enzyme is a potent biocatalyst for the industrial production of chemicals 4.1.2.47 (S)-hydroxynitrile lyase synthesis the enzyme is a valuable catalysts for the synthesis of cyanohydrins, which are versatile chiral building blocks in the pharmaceutical and agrochemical industries 4.1.2.47 (S)-hydroxynitrile lyase synthesis hydroxynitrile lyase is a useful enzyme for production of optically active cyanohydrin compounds 4.1.2.47 (S)-hydroxynitrile lyase synthesis synthesis of pure 2-hydroxycarboxylic acids as valuable synthetic building blocks. Development of bienzymatic cascades to convert aldehydes, via hydrocyanation and subsequent hydration or hydrolysis, into the corresponding (S)-2-hydroxycarboxylic amides and acids. The biocatalysts comprise an (S)-specific hydroxynitrile lyase combined with a nonselective nitrile hydratase or nitrilase. The key to success is preventing racemisation of the intermediate (S)-2-hydroxynitrile while adequately protecting the nitrile-converting enzyme, either in a cross-linked enzyme aggregate (CLEA) or in resting cells.Two biocatalyst systems for the synthesis of (S)-mandelic acid are developed: a combined cross-linked enzyme aggregate (combi-CLEA) of an (S)-hydroxynitrile lyase and a nitrilase, as well as a whole-cell Escherichia coli biocatalyst expressing both enzymes. The nitrilase formed large amounts of mandelicamide, which was remedied by including an amidase in the combi-CLEA as well as by using nitrilasevariants obtained by directed mutagenesis in the whole-cell biocatalyst. Excellent results with more than 95% conversion of benzaldehyde into (S)-mandelic acid with near-quantitative enantiomeric purity are obtained with both biocatalyst systems. Directed mutagenesis of the nitrilase provides an amide-selective whole-cell biocatalyst, which produces (S)-mandelic amide in near-stoichiometric yields. (S)-2-hydroxylalkanoic carboxamides are synthesised in the presence of CLEAs of hydroxynitrile lyase and nitrile hydratase 4.1.2.47 (S)-hydroxynitrile lyase synthesis use of biocatalytic active static emulsions (BASE) for the hydroxynitrile lyase-catalyzed synthesis of enantiopure cyanohydrins. With this technique a full suppression of the undesired racemic non-enzymatic side reaction is facilitated, even at unusually high pH within the aqueous phase. BASE is an inclusion immobilization. It consists of a hydrophobic matrix, typically polydimethylsiloxane (PDMS), with dispersed domains of an aqueous phase 4.1.2.48 low-specificity L-threonine aldolase synthesis production of L-threo-3,4-dihydroxyphenylserine. At the optimized conditions, a mixture of L-threo-3,4-dihydroxyphenylserine and L-erythro-3,4-dihydroxyphenylserine is synthesized by diastereoselectivity-enhanced L-threonine aldolase expressed in Escherichia coli in a continuous process for 100 h, yielding an average of 4.0 mg/ml of L-threo-3,4-dihydroxyphenylserine and 60% diastereoselectivity 4.1.2.48 low-specificity L-threonine aldolase synthesis synthesis of optically active beta-hydroxy-alpha-amino acids by immobilized Escherichia coli cells expressing the enzyme. The immobilized cells can be continuously used 10 times, yielding an average conversion rate of 60.4% 4.1.2.48 low-specificity L-threonine aldolase synthesis the enzyme may be exploited for bioorganic synthesis of L-3-hydroxyamino acids that are biologically active or constitute building blocks for pharmaceutical molecules 4.1.2.55 2-dehydro-3-deoxy-phosphogluconate/2-dehydro-3-deoxy-6-phosphogalactonate aldolase synthesis stereoselectivity is induced into aldol reactions of this naturally promiscuous enzyme by employing D-glyceraldehyde acetonide and L-glyceraldehyde acetonide as substrates 4.1.2.61 feruloyl-CoA hydratase/lyase synthesis biotechnological production of vanillin 4.1.2.61 feruloyl-CoA hydratase/lyase synthesis useful for commercial microbial batch fermentation of vanillin, economical production of vanillic acid instead of chemical oxidation of vanillin 4.1.3.3 N-acetylneuraminate lyase synthesis enzymatic synthesis of N-acetylneuraminic acid with immobilized enzyme 4.1.3.3 N-acetylneuraminate lyase synthesis enzymatic synthesis of N-acetylneuraminic acid, large scale production using N-acetylneuraminate lyase and N-acetyl-D-glucosamine-2-epimerase 4.1.3.3 N-acetylneuraminate lyase synthesis production of N-acetyl-D-neuraminic acid, which is the major representative of amino sugars. The synthesis of N-acetyl-D-neuraminic acid is of interest in studies towards inhibitors of neuraminidase, hemagglutinin and selectin-mediated leucocyte adhesion 4.1.3.3 N-acetylneuraminate lyase synthesis method to produce N-acetylneuraminic acid efficiently. Using a recombinant human renin binding protein (rhRnBp) showing GlcNAc-2-epimerase activity and Escherichia coli sialic acid aldolase, about 80% conversion yield of Neu5Ac is obtained in the coupling reaction under 10fold excess of pyruvate to GlcNAc based on the initial concentration of GlcNAc. The equilibrium of GlcNAc-2-epimerase reaction is not affected by temperature, whereas that of sialic acid aldolase reaction is shifted toward Neu5Ac by lowering the reaction temperature. Low temperatures improve the conversion yield of Neu5Ac, but decrease the reaction rate in the coupling reaction. A high reaction rate as well as a high conversion yield can be achieved by shifting the temperature of the coupling reaction during the reaction 4.1.3.3 N-acetylneuraminate lyase synthesis the enzyme can be a useful and cheap biocatalyst to produce N-acetylneuraminic acid (Neu5Ac) 4.1.3.16 4-Hydroxy-2-oxoglutarate aldolase synthesis use as synthetic tool for C-C bond formation considered 4.1.3.27 anthranilate synthase synthesis Oryza sativacalli overexpressing OASA1D:OASA1D is a system for the production of significant amounts of pharmacologically useful indole alkaloids in rice 4.1.3.42 (4S)-4-hydroxy-2-oxoglutarate aldolase synthesis preparation of (4S)-4-hydroxy-2-oxoglutarate at more than 95% enantiomeric excess by stereospecific synthesis from glyoxylate and pyruvate. Preparation of (4R)-4-hydroxy-2-oxoglutarate at 60% enantiomeric excess by selective cleavage of the (4S)-isomer of racemic 4-hydroxy-2-oxoglutarate leaving the (4R)-isomer in solution 4.1.99.1 tryptophanase synthesis production of L-tryptophan and related amino acids 4.1.99.2 tyrosine phenol-lyase synthesis development of a multienzyme reactor with ec 4.1.99.2 and ec 4.1.1.25 for dopamine synthesis 4.1.99.2 tyrosine phenol-lyase synthesis production of L-dopa, which is applied for the treatment of Parkinsonism 4.1.99.2 tyrosine phenol-lyase synthesis major applications of tyrosine phenol lyase in the production of L-tyrosine and L-DOPA. Tyrosine phenol lyase also has the ability to modify and synthesize natural and non-natural amino acids 4.1.99.2 tyrosine phenol-lyase synthesis tyrosine phenol lyase is an important enzyme for the synthesis of pharmaceutical aromatic amino acids, viz. L-tyrosine and L-DOPA 4.1.99.2 tyrosine phenol-lyase synthesis high activity and high level expression of Fn-TPL in Escherichia coli provides a cheap and robust biocatalyst for large-scale production of L-DOPA 4.1.99.2 tyrosine phenol-lyase synthesis the tyrosine phenol lyase (TPL) catalyzed synthesis of L-DOPA is regarded as one of the most economic routes for L-DOPA synthesis 4.1.99.2 tyrosine phenol-lyase synthesis the tyrosine phenol lyase (TPL) catalyzes synthesis of L-DOPA and can be used for large scale L-DOPA synthesis with direct incorporation of L-DOPA into proteins 4.1.99.12 3,4-dihydroxy-2-butanone-4-phosphate synthase synthesis the enzyme is essential for industrial riboflavin production by Bacillus subtilis overproducing strains, overview 4.1.99.20 3-amino-4-hydroxybenzoate synthase synthesis sweet sorghum juice is a potentially suitable feedstock for 3,4-AHBA production by recombinant Corynebacterium glutamicum expressing enzyme GriH from Streptomyces griseus. 3-Amino-4-hydroxybenzoic acid (3,4-AHBA) is a precursor for production of the bioplastic 4.2.1.2 fumarate hydratase synthesis highly stable enzyme would be ideal for use in various industrial processes, especially since the specific activity is also very high 4.2.1.2 fumarate hydratase synthesis economic production of L-malate in an enzyme membrane reactor 4.2.1.2 fumarate hydratase synthesis production of (S)-malic acid, which is used as an acidurant in fruit and vegetable juices, carbonated soft drinks, jams and candies, in amino acid infusions and for the treatment of hepatic malfunctioning 4.2.1.2 fumarate hydratase synthesis a pfl ldhA double mutant Escherichia coli strain NZN11 is used to produce succinic acid by overexpressing the Escherichia coli malic enzyme gene sfcA. This strain, however, produces a large amount of malic acid as well as succinic acid. The fumB gene encoding the anaerobic fumarase of Escherichia coli is co-amplified to solve the problem of malic acid accumulation, and subsequently improve the succinic acid production 4.2.1.2 fumarate hydratase synthesis stFUMC is a highly efficient, thermostable fumarase C with industrial potential 4.2.1.2 fumarate hydratase synthesis fumarase is used for the industrial production of L-malate from the substrate fumarate 4.2.1.2 fumarate hydratase synthesis overexpression of mutant P160A in Torulopsis glabrata leads to production 5.2 g/l fumarate. Additional deletion of adenylosuccinate synthase, a component of the purine nucleotide cycle, results in production of 9.2 g/l fumarate 4.2.1.2 fumarate hydratase synthesis synthesis of fumarate in 50% ethylene glycol to shift the reaction equilibrium to fumaric acid. 54.7% conversion is observed using fumarase for transforming 1 mmol L-malic acid. 27% total yield is obtained with 99% purity 4.2.1.2 fumarate hydratase synthesis use of tryptophan synthase in the generation of L-dihalotryptophans and L-alkynyltryptophans 4.2.1.3 aconitate hydratase synthesis overexpression of enzyme in Escherichia coli. Presence of co-expressed GroEL reduces the aconitase over-expression drastically, however, exogenous GroEL and GroES together compensate this reduction. For over-expressing cells, growth-rate decreases by 30% at 25°C, however, in presence of co-expressed GroEL and GroES the growth rate of aconitase producing cells is enhanced by 30% at 37°C 4.2.1.9 dihydroxy-acid dehydratase synthesis the enzyme can be used for a convenient one-step synthesis route from D-gluconate to 2-dehydro-3-deoxy-D-gluconate 4.2.1.12 phosphogluconate dehydratase synthesis synthesis of 6-phospho-2-dehydro-3-deoxy-D-gluconate 4.2.1.20 tryptophan synthase synthesis Recycling of hair by acid hydrolysis has enormous economic importance. Enzymatic synthesis of L-tryptophan from hair acid hydrolysis industries wastewater with tryptophan synthase. The L-serine conversion rate reaches 95.1% with a final L-tryptophan concentration of 33.2 g/l 4.2.1.20 tryptophan synthase synthesis synthesis of L-2-methyltryptophan L-serine and 2-methylindole by recombinant Escherichia coli with tryptophan synthase activity. Under the optimal conditions including 100 mmol/l L-serine, 100 mmol/l 2-methylindole, 1 g tryptophan synthase cells, 0.1mmol/l Ca2+, 50 ml reaction volume, 37ºC, pH 8.0, the bioconversion rate of L-2-methyltryptophan reaches 93% after 6 h 4.2.1.20 tryptophan synthase synthesis the reaction of mutant L166V with L-amino acid oxidase, halogenase enzymes and palladium chemocatalysts provides access to further D-configured and regioselectively halogenated or arylated beta-methyltryptophan derivatives 4.2.1.20 tryptophan synthase synthesis use of enzyme in production of psilocybin formation from 4-hydroxyindole and L-serine, and similarly of 7-phosphoryloxytryptamine (isonorbaeocystin), and of serotonin, together with Psilocybe cubensis enzymes PsiD, PsiK, and PsiM, which provide decarboxylase, kinase, and methyltransferase activity, respectively 4.2.1.B20 onoceroid synthase synthesis the onoceroid synthase function of BmeTC enables the synthesis of (+)-ambrein, a major constituent of ambergris that is difficult to obtain naturally, via a mutated squalene-hopene cyclase-catalyzed reaction from easily available squalene 4.2.1.24 porphobilinogen synthase synthesis in addition to hemA and hemL, hemB, hemD, hemF, hemG and hemH are also major regulatory targets of the heme biosynthesis pathway. Up-regulation of hemD and hemF benefits ALA accumulation whereas overexpression of hemB, hemG and hemH diminishes ALA accumulation. By combinatorial overexpression of hemA, hemL,hemD and hemF with different copy-number plasmids, the titer of ALA can be improved to 3.25 g/l 4.2.1.28 propanediol dehydratase synthesis production of 2-butanol, using a TEV-protease based expression system to achieve equimolar expression of the B12-dependent dehydratase subunit and use of NADH-dependent secondary alcohol dehydrogenase Sadh from Gordonia sp. able to catalyze the subsequent conversion of butanone to 2-butanol. A final concentration of 460.2 mg/l 2-butanol and 260.1 mg/l of butanone is found. Availability of NADH is achieved by growing cells lacking the GPD1 and GPD2 isogenes under anaerobic conditions 4.2.1.30 glycerol dehydratase synthesis 1,3-propanediol, starting material for producing plastics, from glycerol relies on the activity of glycerol or diol dehydratases 4.2.1.30 glycerol dehydratase synthesis 1,3-Propanediol is a valuable chemical intermediate,which is particularly suitable as a monomer for polycondensations to produce polyesters, polyethers and polyurethanes 4.2.1.30 glycerol dehydratase synthesis synthesis of 1,3-propanediol 4.2.1.30 glycerol dehydratase synthesis synthesis of 1,3-propanediol, which is a suitable monomer for polycondensations to produce polyesters, polyethers, and polyurethanes 4.2.1.30 glycerol dehydratase synthesis 7.4fold increased production of 3-hydroxypropionic acid from glycerol in the DELTAtpiA DELTAzwf DELTAyqhD Escherichia coli strain by the expression of glycerol dehydratase from Klebsiella pneumoniae and aldehyde dehydrogenase 4.2.1.30 glycerol dehydratase synthesis enzymatic production of 1,3-propanediol, which is a compound of great potential application in many synthesis reactions, in particular as a monomer for polycondensations to produce polyethers, polyurethanes, and polyesters. An optimal PCR system is developed for effective and reproducible PCR amplification of unknown long gene segments from the genome. Using this method, a new glycerol dehydratase is obtained, which eliminates the need for coenzyme B12. This reduces the cost of 1,3-propanediol production from renewable resources 4.2.1.30 glycerol dehydratase synthesis Escherichia coli is engineered to produce 1,3-propanediol from glycerol, an inexpensive carbon source. This is done by introducing a synthetic 1,3-propanediol production pathway in recombinant Escherichia coli consisting of glycerol dehydratase complex (dhaB123) and glycerol dehydratase reactivation factors (gdrAB) from Klebsiella pneumoniae and 1,3-propanediol oxidoreductase isoenzyme (yqhD) from Escherichia coli. When 10 mM succinate is added to the medium, the titer of 1,3-propanediol and the glycerol consumption increase to 9.9 and 23.84 g/l, respectively. In addition, the ratio of NADH to NAD+ increases by 43%. Succinate addition is a promising route for the biotechnological production of NADH-dependent microbial metabolites 4.2.1.30 glycerol dehydratase synthesis glycerol dehydratase is a key and rate-limiting enzyme in the bioconversion process of glycerol to 3-hydroxypropionaldehyde, which is further reduced to 1,3-propanediol by 1,3-propanediol dehydrogenase. 3-Hydroxypropionaldehyde is an important chemical industry material, being a potent bacterial inhibitor and precursor in the production of numerous industrial chemicals, such as 1,3-propanediol 4.2.1.30 glycerol dehydratase synthesis glycerol dehydratase is a key enzyme for the production of 1,3-propanediol in soluble cell extract. Klebsiella pneumoniae J2B shows a high potential for the production of 1,3-propanediol from glycerol. Optimization of the culture conditions and the elimination of lactate synthesis improves 1,3-propanediol production significantly 4.2.1.30 glycerol dehydratase synthesis industrial production of 3-hydroxypropionic acid, 3-hydroxypropionaldehyde and 1,3-propanediol. Lactobacillus reuteri DSM 20016 is amenable to metabolic engineering and a wide variety of methods for its genetic manipulation are available. Engineering of the pdu operon to increase the glycerol-utilization rate is a good strategy to increase specific production rates, and further manipulation could render a robust strain for industrial applications 4.2.1.30 glycerol dehydratase synthesis synthesis of 3-hydroxypropionic acid. UTR engineering is used to maximally increase the activities of glycerol dehydratase and aldehyde dehydrogenase for the high conversion of glycerol to 3-hydroxypropionic acid. Thereafter, the activity of glycerol dehydratase is precisely controlled to avoid the accumulation of 3-hydroxypropionaldehyde by varying expression of dhaB1, a gene encoding a main subunit of glycerol dehydratase. The optimally balanced Escherichia coli HGL_DBK4 shows a substantially enhanced 3-hydroxypropionic acid titer and productivity compared with the parental strain. The yield on glycerol is 0.97 g 3-hydroxypropionic acid/g glycerol, in a fed-batch experiment 4.2.1.30 glycerol dehydratase synthesis synthesis of butanone 4.2.1.31 maleate hydratase synthesis biosynthesis of synthons for fine chemicals industry, description of a crystal-liquid two-phase system and soluble metal-organic acid complex system for fermentation which minimizes substrate inhibition and thereby increases efficiency 4.2.1.31 maleate hydratase synthesis use for synthesis of chiral synthons considered 4.2.1.31 maleate hydratase synthesis production of (R)-malic acid, which can be used as chiral synthon in organic chemistry or as resolving agent in resolution of racemic compounds 4.2.1.36 homoaconitate hydratase synthesis the enzymes' stereospecific hydrolyase activity make it an attractive catalyst to produce diastereomers from unsaturated precursors, analysis of the structural basis for engineering of new stereospecific hydro-lyase enzymes for chemoenzymatic syntheses, overview 4.2.1.36 homoaconitate hydratase synthesis glutarate biosynthetic pathway by incorporation of a +1 carbon chain extension pathway from 2-oxoglutarate in combination with 2-oxo acid decarboxylation pathway in Escherichia coli. Introduction of homocitrate synthase, homoaconitase and homoisocitrate dehydrogenase from Saccharomyces cerevisiae into Escherichia coli enables +1 carbon extension from 2-oxoglutarate to 2-oxoadipate, which is subsequently converted into glutarate by a promiscuous 2-oxo acid decarboxylase (KivD) and a succinate semialdehyde dehydrogenase (GabD). The recombinant Escherichia coli coexpressing all five genes produces 0.3 g/l glutarate from glucose. To further improve the titers, 2-oxoglutarate is rechanneled into carbon chain extension pathway via the clustered regularly interspersed palindromic repeats system mediated interference (CRISPRi) of essential genes sucA and sucB in tricarboxylic acid cycle. The final strain can produce 0.42 g/l glutarate, which is increased by 40% compared with the parental strain. Glutarate is one of the most potential building blocks for bioplastics 4.2.1.39 gluconate dehydratase synthesis utilization for fermentation 2-keto-3-deoxy-D-gluconate proposed, conditions for fermentation 4.2.1.39 gluconate dehydratase synthesis highly economic one-step biocatalytic synthesis procedure of 2-keto-3-deoxy-D-gluconate. D-Gluconate is completely converted tostereochemically pure 2-dehydro-3-deoxy-D-gluconate without side-product formation. The final 2-dehydro-3-deoxy-D-gluconate yield is approximately 90% 4.2.1.39 gluconate dehydratase synthesis highly economic one-step biocatalytic synthesis procedure of 2-oxo-3-deoxy-D-gluconate from D-gluconate using recombinant gluconate dehydratase (GAD) from the hyperthermophilic crenarchaeon Thermoproteus tenax 4.2.1.39 gluconate dehydratase synthesis one-step biocatalytic synthesis procedure for 2-keto-3-deoxy-D-gluconate from D-gluconate using recombinant gluconate dehydratase. The D-gluconate is completely converted to stereochemically pure D-2-keto-3-deoxy-D-gluconate without side-product formation, the final 2-keto-3-deoxy-D-gluconate yield is approximately 90%, the quantitative and qualitative LC-MS analysis method enables the simultaneous detection of D-gluconate and 2-keto-3-deoxy-D-gluconate and the enzyme can be provided by a simple and rapid procedure involving only two precipitation steps 4.2.1.46 dTDP-glucose 4,6-dehydratase synthesis hemo-enzymatic syntheses of artificial and naturally occuring deoxysugars 4.2.1.46 dTDP-glucose 4,6-dehydratase synthesis Synthesis of dTDP-6-deoxy-4-keto-D-glucose (intermediate for synthesis of activated deoxysugars) 4.2.1.47 GDP-mannose 4,6-dehydratase synthesis GMD knockout CHO cells are ideal host cells for the manufacture of completely non-fucosylated therapeutic antibodies and completely O-fucose-negative therapeutics as well 4.2.1.47 GDP-mannose 4,6-dehydratase synthesis repression of the GMD gene is thus very useful for deleting immunogenic total fucose residues and facilitating the production of pharmaceutical glycoproteins in plants 4.2.1.51 prephenate dehydratase synthesis synthesis of L-Phe 4.2.1.51 prephenate dehydratase synthesis synthesis of L-Phe, pheA gene expressed in Corynebacterium glutamicum 4.2.1.51 prephenate dehydratase synthesis synthesis of L-phenylalanine (an important amino acid that is widely used in the production of food flavors and pharmaceuticals) by engineered Escherichia coli. Coexpression of Vitreoscilla hemoglobin gene, driven by a tac promoter, with the genes encoding 3-deoxy-D-arabinoheptulosonate-7-phosphate synthetase (aroF) and feedback-resistant chorismate mutase/prephenate dehydratase (pheAfbr), leads to increased productivity of L-phenylalanine and decreased demand for aeration by Escherichia coli CICC10245 4.2.1.53 oleate hydratase synthesis (R)-10-hydroxyoctadecanoic acid, produced by the oleate hydratase is a precursor for gamma-dodecalactone which is an important fragrance and flavour compound 4.2.1.53 oleate hydratase synthesis one of the largest bottlenecks in producing lactones is the hydroxylation of the fatty acids. The exploitation of oleate hydratase to hydrate oleic acid, which can easily be turned into gamma-dodecalactone after beta-elimination, is therefore the perfect solution to produce this valuable compound from cheap renewable materials (e.g. vegetable oil). The specificity and enantioselectivity of the enzyme will provide the correct lactone 4.2.1.78 (S)-norcoclaurine synthase synthesis development of an efficient, stereoselective, green synthesis of (S)-norcoclaurine, i.e. higenamine, using the recombinant (S)-norcoclaurine synthase enzyme, starting from the cheap tyrosine and dopamine substrates in a one-pot, two step process, overview. The optimized process affords enantiomerically pure (S)-norcoclaurine (93%) in a yield higher than 80% and allows good recovery of the enzyme for recycling, by a green Pictet-Spengler synthesis 4.2.1.78 (S)-norcoclaurine synthase synthesis the enzyme can catalyse the Pictet-Spengler reaction between dopamine and unactivated ketones, thus facilitating the facile biocatalytic generation of 1,1'-disubstituted tetrahydroisoquinolines. Variants of the enzyme showing improved conversions are identified and used to synthesize novel chiral 1,10-disubstituted and spiro-tetrahydroisoquinolines 4.2.1.78 (S)-norcoclaurine synthase synthesis the enzyme can serve as a tool to synthesize unnatural, optically active tetrahydroisoquinolines. The enzyme is a promising catalyst that functions to stereoselectively produce various 1-substituted-1,2,3,4-tetrahydroisoquinolines 4.2.1.78 (S)-norcoclaurine synthase synthesis the enzyme has also shown great potential as a biocatalyst for the formation of chiral isoquinolines 4.2.1.78 (S)-norcoclaurine synthase synthesis the enzyme is a highly suitable catalyst for the Pictet-Spengler reaction 4.2.1.78 (S)-norcoclaurine synthase synthesis the enzyme is a highly suitable catalyst for the Pictet-Spengler reaction. The enzyme is successfully immobilized on various carriers whereby EziG3 proves to be the best suited for biotransformations 4.2.1.84 nitrile hydratase synthesis - 4.2.1.84 nitrile hydratase synthesis useful for acrylamide production 4.2.1.84 nitrile hydratase synthesis industrial production of nicotinamide 4.2.1.84 nitrile hydratase synthesis useful for modification of polyacrylonitrile fibers and granulates 4.2.1.84 nitrile hydratase synthesis H-NHase is used in the industrial production of acrylamide and nicotinamide 4.2.1.84 nitrile hydratase synthesis production of nicotinamide (=vitamin B3) for vitamin supplement for food and animal feed 4.2.1.84 nitrile hydratase synthesis production of 2-naphthylacetamide by one-pot chemo-enzymatic conversion 4.2.1.84 nitrile hydratase synthesis production of alpha-hydroxy nitriles by use of enzyme 4.2.1.84 nitrile hydratase synthesis production of D-tert-leucine-nitrile from racemic tert-leucine-nitrile by use of enzyme plus D-selective amidase from Variovorax paradoxus 4.2.1.84 nitrile hydratase synthesis production of propionamide by use of enzyme in ultrafiltration-membrane reactor with maximum volumetric production of 0.5 g propionamide per litre and h 4.2.1.84 nitrile hydratase synthesis industrial production of (S)-2,2-dimethylcyclopropanecarboxylic acid 4.2.1.84 nitrile hydratase synthesis enzyme can be used in conjunction with a stereoselective amidase to synthesize ethyl (S)-4-chloro-3-hydroxybutyrate, an intermediate for a hypercholesterolemia drug, Atorvastatin 4.2.1.84 nitrile hydratase synthesis nitrile hydratase is an enzyme used in the industrial biotechnological production of acrylamide 4.2.1.84 nitrile hydratase synthesis nitrile hydratase is used for large scale industrial production of important commodities such as acrylamide and nicotinamide 4.2.1.84 nitrile hydratase synthesis the enzyme is useful in synthesis of compounds by hydrating biotransformations, optimization of strain cultivation and enzyme production and activity, overview 4.2.1.84 nitrile hydratase synthesis bioconversion of 3-cyanopyridine using the in situ nitrile hydratase-amidase cascade system of resting Microbacterium imperiale CBS 498-74 cells in an ultrafiltration-membrane reactor, carried out in continuously stirred tank UF-membrane bioreactors arranged in series, the reactor configuration enables both enzymes, involved in the cascade reaction, to work with optimized kinetics, without any purification, exploiting their differing temperature dependences, method optimization, overview 4.2.1.84 nitrile hydratase synthesis bioconversion of 3-cyanopyridine using the in situ nitrile hydratase-amidase cascade system of resting Microbacterium imperiale CBS 498-74 cells in an ultrafiltration-membrane reactor, operated in either batch or continuous mode, method optimization, overview 4.2.1.84 nitrile hydratase synthesis nitrile hydratase-catalyzed preparation of 2-amino-2,3-dimethylbutyramide, ADBA, a key intermediate for imidazolinone herbicides, method development and optimization, evaluation of the appropriate organism, overview 4.2.1.84 nitrile hydratase synthesis transformation of benzonitrile into benzohydroxamic acid performed by a cascade bienzymatic reaction involving nitrile hydration and acyl transfer of the intermediate benzamide onto hydroxylamine. The first step is catalyzed by a cell-free extract from recombinant Escherichia coli strain expressing nitrile hydratase from Klebsiella oxytoca strain 38.1.2, the second step is a cellfree extract from Rhodococcus erythropolis A4 amidase, EC 3.5.1.4 4.2.1.84 nitrile hydratase synthesis transformation of benzonitrile into benzohydroxamic acid performed by a cascade bienzymatic reaction involving nitrile hydration and acyl transfer of the intermediate benzamide onto hydroxylamine. The first step is catalyzed by a cell-free extract from recombinant Escherichia coli strain expressing nitrile hydratase from Raoultella terrigena srain 77.1, the second step is a cell-free extract from Rhodococcus erythropolis A4 amidase, EC 3.5.1.4 4.2.1.84 nitrile hydratase synthesis transformation of benzonitrile into benzohydroxamic acid performed by a cascade bienzymatic reaction involving nitrile hydration and acyl transfer of the intermediate benzamide onto hydroxylamine. The first step is catalyzed by a cell-free extract from Rhodococcus erythropolis A4 containing nitrile hydratase, the second step is a cell-free extract from Rhodococcus erythropolis A4 amidase, EC 3.5.1.4 4.2.1.84 nitrile hydratase synthesis NHases are important for large scale production of acrylamide and nicotinamide 4.2.1.84 nitrile hydratase synthesis industrial production of highly purified acrylamide and nicotinamide. The thermostability and catalytic efficiency of the subunit-fused nitrile hydratase is improved by semi-rational engineering 4.2.1.84 nitrile hydratase synthesis the enzyme from Rhodococcus aetherivorans JB1208 shows high regioselectivity and strong substrate tolerance for alicyclic dinitrile and affords a potentially industrial route to gabapentin (a precursor of gamma-aminobutyric acid, which has been approved for treatment of a variety of central nervous system disorders, partial seizures, and restless legs syndrome) 4.2.1.84 nitrile hydratase synthesis use of recombinant Corynebacterium cells for the production of acrylamide from acrylonitrile results in a conversion yield of 93% and a final acrylamide concentration of 42.5% within 6 h when the total amount of fed acrylonitrile is 456 g 4.2.1.92 hydroperoxide dehydratase synthesis enzymatic synthesis of volatile C6 aldehydes (hexanal, 2(E)-hexenal and 3(Z)-hexenal) using lipoxygenase (LOX) and hydroperoxide lyase (HPL) as biocatalysts. C6 aldehydes are widely used in perfume and food industry, because they are important contributors to the distinctive scent of fresh fruits and vegetables 4.2.1.92 hydroperoxide dehydratase synthesis the recombinant 13-HPL enzyme is useful biocatalyst to produce C6-aldehydes 4.2.1.92 hydroperoxide dehydratase synthesis the recombinant 13-HPL enzyme is valuated as biocatalyst to produce C6-aldehydes 4.2.1.95 kievitone hydratase synthesis due to its catalytic properties and apparent substrate promiscuity, enzyme NhKHS is a promising enzyme for the biocatalytic production of tertiary alcohols. The production of enantiopure tertiary alcohols is a major challenge of organic synthesis as these functional groups are widely applicable for the generation of pharmaceuticals or other bioactive compounds 4.2.1.95 kievitone hydratase synthesis due to its catalytic properties and apparent substrate promiscuity, enzyme NhKHS is a promising enzyme for the biocatalytic production of tertiary alcohols. The production of enantiopure tertiary alcohols is a major challenge of organic synthesis as these functional groups are widely applicable for the generation of pharmaceuticals or other bioactive compounds. The compatibility of enzyme NhKHS with nonpolar organic solvents is beneficial for a number of biocatalytic applications, e.g. in general for conversion of hydrophobic substrates 4.2.1.118 3-dehydroshikimate dehydratase synthesis introduction of both 3-dehydroshikimate dehydratase and protocatechuic acid decarboxylase into an Escherichia coli construct synthesizing elevated levles of 3-dehydroshikimic acid leads to production of up to 18.5 mM catechol from 56 mM D-glucose on 1 l-scale 4.2.1.118 3-dehydroshikimate dehydratase synthesis production of vanillin by engineered pathway in Schizosaccharomyces pombe or Saccharomyces cerevisiae. Pathway involves incorporation of 3-dehydroshikimate dehydratase, an aromatic carboxylic acid reductase from a bacterium of the Nocardia genus, and an O-ethyltransferase from Homo sapiens. In Saccharomyces cerevisiae, the aromatic carboxylic acid reductase enzyme requires activation by phosphopantetheinylation, achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol is achieved by knockout of the host alcohol dehydrogenase ADH6. In Schizosaccharomyces pombe, the biosynthesis is further improved by introduction of an Arabidopsis thaliana family 1 UDPglycosyltransferase, converting vanillin into vanillin beta-D-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts 4.2.1.122 tryptophan synthase (indole-salvaging) synthesis the enzyme participates in psilocybin formation from 4-hydroxyindole and L-serine, which are less cost-intensive substrates, compared to the previous method. The pharmaceutical interest in this psychotropic natural product as a future medication to treat depression and anxiety is strongly reemerging. Enzymatic production of 7-phosphoryloxytryptamine (isonorbaeocystin), a non-natural congener of the Psilocybe alkaloid norbaeocystin (4-phosphoryloxytryptamine), and of serotonin (5-hydroxytryptamine) by means of the same in vitro approach 4.2.1.125 dammarenediol II synthase synthesis the production of dammarenediol-II in a cell suspension culture of transgenic tobacco can be applied to the large-scale production of this compound and utilized as a source of pharmacologically active medicinal materials 4.2.1.127 linalool dehydratase synthesis the dehydration reaction of Ldi or genetically engineered variants thereof can be used for the transformation of smaller substrate molecules into butadiene and isoprene 4.2.1.135 UDP-N-acetylglucosamine 4,6-dehydratase (configuration-retaining) synthesis sugar capsule capsular polysaccharide A (CPSA), which coats the surface of the mammalian symbiont Bacteroides fragilis, is a key mediator of mammalian immune system development, the enzyme, coupled to a Bacteroides fragilis-encoded aminotransferase (WcfR), is used in sythetic construction system for CPSA for synthesis of the rare stereoconfiguration sugar acetamido-4-amino-6-deoxygalactopyranose in a two-step coupled reaction with WcfR, overview 4.2.1.146 L-galactonate dehydratase synthesis extracellular production of L-galactonate in gram quantities from D-galacturonic and polygalacturonic acids. DELTAgaaB strains produce L-galactonate at yields of 0.6 to 0.9 g per g of substrate consumed. Intracellular accumulation of L-galactonate suggests that export may be limiting. Deletion of the L-galactonate dehydratase from Aspergillus niger delays induction of D-galacturonate reductase and overexpression of the reductase improves initial production rates. Deletion of the L-galactonate dehydratase from Aspergillus niger also delays or prevents induction of the putative D-galacturonate transporter An14g04280. Aspergillus nigergaaB produces L-galactonate from polygalacturonate as efficiently as from the monomer 4.2.1.159 dTDP-4-dehydro-6-deoxy-alpha-D-glucopyranose 2,3-dehydratase synthesis construction of a Streptomyces albus strain harboring the oleG2 glycosyltransferase gene and transformation with plasmid constructs containing a set of genes proposed to be required for the biosynthesis of dTDP-L-olivose and dTDP-L-oleandrose, respectively. Incubation of these strains with the erythromycin aglycon (erythronolide B) gives rise to two new glycosylated compounds, identified as L-3-O-olivosyl- and L-3-O-oleandrosyl-erythronolide B. Plasmid constructs pOLV and pOLE containing the oleW, oleV, oleL, oleS, oleE, and oleU genes and the 5' end of oleNI encode all enzyme activities required for the biosynthesis of these two 2,6-dideoxysugars 4.2.1.173 ent-8alpha-hydroxylabd-13-en-15-yl diphosphate synthase synthesis CPS4 and kaurene synthase-like2 act together to produce ent-13-epi-manoyl oxide 4.2.2.1 hyaluronate lyase synthesis enzymatic production of fully O-sulfated oligosaccharides by large scale depolymerization of hyaluronan polymer 4.2.2.3 mannuronate-specific alginate lyase synthesis alginate production is improved by use of the constructed mutant strain SML2, due to enzyme-deficiency the alginate production reaches a higher level and the alginate polymers have a higher molecular weight 4.2.2.3 mannuronate-specific alginate lyase synthesis enzyme-catalyzed gel-sol transition of calcium-alginate obtained by internal gelling strategy with the help of an entrapped alginate lyase. Alginate molecules and enzyme-produced oligoalginates shorten the gel time of physical gelatin gels (5% and 1.5%), probably due to local protein concentration increase. Interpenetrated networks composed of calcium-alginate and of gelatin are obtained only if elongation of gelatin helices inside a pre-existing calcium-alginate network could occur and only for low gelatin concentration (1.5%). The physical gelatin network is almost reversible inside the alginate one. Both networks can be obtained in the presence of alginate lyase, but gel-sol transition of calcium-alginate cannot be obtained in the presence of gelatin 4.2.2.3 mannuronate-specific alginate lyase synthesis aly-SJ02 may be a good tool to produce dimers and trimers from alginate 4.2.2.3 mannuronate-specific alginate lyase synthesis the recombinant MJ-3 alginate lyase can be used as a biocatalyst for saccharification of alginate since it can efficiently degrade poly-M block, poly-G block, poly-MG block, alginate oligosaccharides, and alginate into alginate monosaccharides 4.2.2.3 mannuronate-specific alginate lyase synthesis Alg17C can be used as the key enzyme to produce alginate monomers in the process of utilizing alginate for biofuels and chemicals production 4.2.2.5 chondroitin AC lyase synthesis the enzyme can serve as a suitable replacement for the original enzyme, from Arthrobacter aurescens IAM 110 65, UniProt ID P84141, which is no longer commercially available 4.2.2.7 heparin lyase synthesis expression as fusion protein, fused to the C-terminus of soluble partners translation initiation factor 2 domain I, glutathione S-transferase, maltose-binding protein, small ubiquitin modifying protein and N-utilization substance A, and purification of hybrid proteins. Except for NusA, the fusion partners dramatically improve the soluble expression of recombinant HepA, with translation initiation factor 2 domain I-HepA and small ubiquitin modifying protein-HepA creating almost completely soluble HepA where 98% and 94% of expressed HepA fusions are soluble, respectively 4.2.2.7 heparin lyase synthesis expression in Escherichia coli using a hexahistidine-tagged small ubiquitin-like modifier. The fusion protein exhibits high enzyme activity without requirements of in vitro refolding and SUMO-tag releasing process, and the optimum enzyme activity is obtained at 30°C, pH 7.0 and 10 mmol/l Ca2+ in the reaction buffer 4.2.2.7 heparin lyase synthesis expression of HepI with the chitin binding domain of Bacillus circulans chitinase A1 as a chitin-affinity tag, and the small ubiquitin-like modifier (SUMO) linker as a solvation enhancer in different fusion sequence. The constructs chitin binding domain-HepI, chitin binding domain-SUMO-HepI, SUMO-chitin binding domain-HepI and chitin binding domain-HepI-SUMO show specific enzyme activities of 1.88, 3.69, 3.44, and 2.73 IU/mg total proteins, respectively, with unfractionated heparin as substrate. Chitin binding domain-SUMO-HepI exhibits the maximum half-life (48 min) at 30°C and best thermostability under 15-50°C. All the fusion enzymes show broad pH-stability in the range of 5.4-9.0 4.2.2.B7 mannuronate-specific alginate exolyase synthesis production of highly pure 4-deoxy-L-erythro-5-hexoseulose uronic acid using 0.78 mg of TcAlg1 per ml substrate solution (2% w/v of sodium alginate) during 12 h, overall yield is 28% 4.2.2.8 heparin-sulfate lyase synthesis construcution of a photoswitchable biocatalyst consisating of a heparinase III mutant K130C-N,N-dimethylacrylamide-co-4-phenyl azophenyl acrylate copolymer. Upon photoswitch, the enzymatic degradation of heparin can be artificially controlled to produce low molecular weight heparin with more uniform molecular weight and an increase in anticoagulant activity 4.2.2.10 pectin lyase synthesis cheap and simple method of immobilization is attractive for the preparation of tailored systems for industrial application as well as for studies on substrate-enzyme-support interactions 4.2.2.10 pectin lyase synthesis production of pectin lyase by Aspergillus giganteus. The highest activity, using citrus pectin as carbon source, is obtained in 11-day-old standing cultures, but the highest specific activity is obtained in 6.5-day-old shaken cultures, at pH 6.5 and 35°C. Using orange waste as carbon source, the highest activity is observed in 8-day-old standing cultures, at pH 7.0 and 30°C 4.2.2.10 pectin lyase synthesis expression in a Kluyveromyces lactis ku80- strain defective in the non-homologous end-joining pathway increases the efficiency of homologous recombination with pKLAC1 vectors and the production and secretion of recombinant enzyme 4.2.2.10 pectin lyase synthesis maximal production of functional Pnl1 is obtained after induction by 0.4 mM isopropyl beta-D-thiogalactoside at 30°C and 150 rpm for 16 h. The use of Origami host strain significantly improves the yield of the soluble active form of the enzyme 4.2.2.10 pectin lyase synthesis maximum pectin lyase activity in a synthetic medium (42 g/l pectin, 40 g/l yeast extract, and 0.02 g/l iron sulfate) is 31 U/ml, and 46 U/ml in the agro-industrial medium (160 g/l orange peel, 150 g/l corn steep liquor, and 300 g/l parboiled rice water), obtained over 60 and 124 h of bioproduction, 180 r/min, 30 x02C, pH initial 5.5, and 5 million spores/ml, respectively 4.2.2.10 pectin lyase synthesis maximum production of pectinase (3315 U/gds) and pectin lyase (10.5 U/gds) by Bacillus subtilis using solid state fermentation in the presence of a combination of orange peel and coconut fiber (4:1), with a moisture content of 60% at 35 °C and pH 4.0 after 4 days and 8 days of incubation, respectively. Pectin lyase production is unaffected by adding carbon and nitrogen source to the basal medium 4.2.2.10 pectin lyase synthesis production of Aspergillus niger on wheat bran and citrus pectin in submerged culture 4.2.2.10 pectin lyase synthesis purification and surface immobilization using various concentrations of chitosan and glutaraldehyde as cross-linking agent. The immobilized fractions are highly stable over a pH and temperature range from 4-9 and 45-85°C, respectively. Chitosan-immobilized enzyme fractions retain more than 75% of their operational activities even after seven consecutive cycles 4.2.2.11 guluronate-specific alginate lyase synthesis Alg17C can be used as the key enzyme to produce alginate monomers in the process of utilizing alginate for biofuels and chemicals production 4.2.2.11 guluronate-specific alginate lyase synthesis alginate lyase is a promising biocatalyst because of its application in saccharification of alginate for the production of biochemicals. Alg2A can be a good tool for the large-scale preparation of alginate oligosaccharides with high degree of polymerization 4.2.2.14 glucuronan lyase synthesis production of glucuronan oligosaccharides 4.2.2.14 glucuronan lyase synthesis production of oligoglucuronans by enzymatic depolymerization of nascent glucuronan 4.2.2.14 glucuronan lyase synthesis large-scale production of oligocellouronic acids 4.2.2.14 glucuronan lyase synthesis production of highly O-acetylated oligoglucuronan in order to envisage new therapeutic applications fields such as the realization of biological tests among animals and vegetals 4.2.2.15 anhydrosialidase synthesis synthesis of sialyl oligosaccharides using enzyme, since it is difficult to obtain them from natural sources 4.2.2.16 levan fructotransferase (DFA-IV-forming) synthesis synthesis of di-D-fructose-2,6':2',6 dianhydride 4.2.2.16 levan fructotransferase (DFA-IV-forming) synthesis development of a di-D-fructofranosyl-2,6':2',6-anhydride production system with single culture of Bacillus subtilis directly from sucrose. This system can avoid the purification procedure of levan in which organic solvent is used for precipitation. The levan fructotransferase gene is cloned from Arthrobacter nicotinovorans GS-9 and expressed in levan producing Bacillus subtilis 168. LFTase activity is detected in the culture supernatant of the transformant with maximal activity of 0.062 U/ml after 15.5 h post induction. Then sucrose is added as substrate and incubated. About 78 h after addition of sucrose, 20.5 g/l of di-D-fructofranosyl-2,6':2',6-anhydride is produced from 139.3 g/l of sucrose consumed. The yield of di-D-fructofranosyl-2,6':2',6-anhydride from sucrose is 14.7 wt.% 4.2.2.16 levan fructotransferase (DFA-IV-forming) synthesis enzymatic production of ascorbic acid 2-fructoside, that can be applied to cosmetics, food products, and pharmaceuticals 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis enzyme immobilized on Chitopearl BCW 3510 can be used 10 times without a significant loss in activity 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis production of dry powder of inulin fructotransferase from fermented liquor. Lyophilized enzyme shows good storage stability 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis use of enzyme in an ultrafiltration membrane bioreactor for the production of difructose anhydrides III from enzymatic conversion of inulin. The enzyme can be continuously used for six runs in the ultrafiltration membrane bioreactor. When the substrate concentration is 100 g/l, the concentration of difructose anhydrides III is about 78.4 g/l, while the productivity and purity of difructose anhydrides III can attain about 2385 and 92%, respectively 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis conversion ratio of inulin to DFA III reaches 81% when 100 g/l inulin is catalyzed by 80 nM recombinant enzyme for 20 min at pH 6.5 and 55°C 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis use of high hydrostatic pressure to improve the catalytic efficiency. The maximum activity of IFTase is obtained under 200 MPa at 60°C. High hydrostatic pressure lowers the energy barrier of the enzymatic reaction and decreases the volume between the reactants and the transition state. Under this condition, the optimal pH for the enzymatic reaction shifts from 5.5 to 6.0. The activity is further enhanced by 65.2% in the presence of 1.5 M NaCl 4.2.2.18 inulin fructotransferase (DFA-III-forming) synthesis using expression of IFTase in Pichia pastoris under the control of the formaldehyde dehydrogenase 1 promoter, efficient secretion with four substrate fed-batch strategies in a 3-L fermenter can be achieved. The cofeeding induction strategy with methylamine hydrochloride and methanol achieves the maximum extracellular IFTase activity of 62.72 U/ml. Carbon sources such as glucose and glycerol can also be utilised using methylamine hydrochloride induction but should be strictly controlled at low concentrations of 0.5-1.5 % (v/v) and 1-1.5 % (w/v), respectively 4.2.2.21 chondroitin-sulfate-ABC exolyase synthesis Production of enzyme for industrial use. Induction of enzyme by addition of chondroitin sulfate C to growth medium. Optimal conditions are pH 8.0, 25°C, plus the addition of yeast extract, peptone and casamino acid. Induction is inhibited by chloramphenicol and actinomycin D 4.2.2.22 pectate trisaccharide-lyase synthesis the highly thermostable enzyme constitutes a useful catalyst for a simplified synthesis of (4-deoxy-alpha-L-threo-hex-4-enopyranosyluronic acid)-(1->4)-(alpha-D-galactopyranosyluronic acid)-(1->4)-alpha-D-galactopyranuronic acid which is extremely difficult to obtain via chemical synthesis 4.2.2.26 oligo-alginate lyase synthesis saccharification of alginate into alginate monosaccharides. Unsaturated monosaccharides up to 3.3 mg/ml are successfully prepared from 1% (w/v) alginate by using the recombinant oligoalginate lyase of Sphingomonas sp. MJ-3 4.2.3.3 methylglyoxal synthase synthesis strains in which the genes for glycerol dehydrogenase, methylglyoxal synthase or both are overexpressed produce 1,2-propanediol as a fermentation product of glucose 4.2.3.3 methylglyoxal synthase synthesis engineering of a functional 1,2-propanediol pathway through a combination of overexpression of genes involved in its synthesis from the key intermediate dihydroxyacetone phosphate and the manipulation of the fermentative glycerol utilization pathway, including the overexpression of methylglyoxal synthase, glycerol dehydrogenase, and aldehyde oxidoreductase. Rreplacement of the native Escherichia coli phosphoenolpyruvate-dependent dihydroxyacetone kinase with an ATP-dependent ihydroxyacetone kinase from Citrobacter freundii allows for higher 1,2-propanediol production. Ethanol is required as co-product, and inreases in 1,2-PDO titer and yield are achieved through the disruption of the pathways for acetate and lactate production. Manipulations result in an engineered Escherichia coli strain capable of producing 5.6 g/l 1,2-propanediol, at a yield of 21.3% w/w. This strain also performs well when crude glycerol is used as the substrate 4.2.3.3 methylglyoxal synthase synthesis expressing Escherichia coli mgs gene in Corynebacterium glutamicum increases 1,2-propanediol yield 100fold. Simultaneous overexpression of mgs and cgR_2242, annotated as encoding putative aldo-keto reductase, enhances 1,2-propanediol production to 24 mM 4.2.3.7 pentalenene synthase synthesis production of pentalenene by expression of pentalenene synthase gene in Xanthophyllomyces dendrorhous with concurrent knock out of the native crtE or crtYB genes. Culturing the yeast in presence of dodecane traps the volatile compounds produced, and no other terpenoids are found as by products 4.2.3.8 casbene synthase synthesis overexpression of truncated protein in Saccharomyces cerevisiae demonstrates casbene production but also high levels of nerolidol and farnesol and mevalonate 4.2.3.8 casbene synthase synthesis overexpression of truncated protein in Saccharomyces cerevisiae demonstrates casbene production up to 26 mg/l 4.2.3.8 casbene synthase synthesis overexpression of truncated protein in Saccharomyces cerevisiae demonstrates casbene production up to 30 mg/l 4.2.3.8 casbene synthase synthesis optimized Agrobacterium-mediated transient expression assay in Nicotiana benthamiana for plant diterpene synthase expression and product analysis. Expression of casbene synthases leads to the accumulation of diterpenes within 3 days of infiltration and with a maximum at 5 days. Over 50% of the products aere exported onto the leaf surface for structure elucidation of unknown diterpenes 4.2.3.13 (+)-delta-cadinene synthase synthesis expression in Saccharomyces cerevisiae leads to a titer of (-)-alpha-copaene of 9 mg/l at 48 h and around 7 mg/l in Escherichia coli, and the titer of (+)-delta-cadinene is 6 mg/l in Saccharomyces cerevisiae and 3.5 mg/l in Escherichia coli 4.2.3.16 (4S)-limonene synthase synthesis engineering of Echerichia coli with a heterologous mevalonate pathway and limonene synthase for production of limonene followed by coupling with a cytochrome P450,which specifically hydroxylates limonene to produce perillyl alcohol. A strain containing all mevalonate pathway genes in a single plasmid produces limonene at titers over 400 mg/l from glucose. Incorporation of a cytochrome P450 to hydroxylate limonene yields approximately 100mg/l of perilyl alcohol 4.2.3.17 taxadiene synthase synthesis constitutive expression in Nicotiana benthamiana leads to de novo production of taxadiene. Transformed homozygous lines produce 11-27 microg taxadiene/g of dry weight. Treatment with an elicitor, methyl jasmonate, and metabolic pathway shunting by suppression of the phytoene synthase gene expression results in increased taxadiene accumulation by 1.4- or 1.9fold, respectively 4.2.3.17 taxadiene synthase synthesis transformation of the roots of cultured Panax ginseng C.A. Meyer to produce taxadiene. Without any change in phenotypes or growth difference, a taxadiene synthase-transgenic ginseng line accumulates 9.1 microg taxadiene per gram of dry weight. In response to the treatment of methyl jasmonate for 3 or 6 days, the accumulation is 14.6 and 15.9 microg per g of dry weight, respectively 4.2.3.17 taxadiene synthase synthesis taxadiene synthase ist he rate-limiting enzyme in the biosynthesis of the anticancer compound paclitaxel 4.2.3.18 abieta-7,13-diene synthase synthesis increase of levopimaradiene synthesis in Escherichia coli by amplification of the flux toward isopentenyl diphosphate and dimethylallyl diphosphate precursors and reprogramming the rate-limiting downstream pathway by generating combinatorial mutations in geranylgeranyl diphosphate synthase and levopimaradiene synthase. The most productive pathway, combining precursor flux amplification and mutant synthases, confers approximately 2600fold increase in levopimaradiene levels. A maximum titer of approximately 700 mg/l is obtained by cultivation in a benchscale bioreactor 4.2.3.B23 neocembrene synthase synthesis overexpression of truncated protein in Saccharomyces cerevisiae demonstrates casbene production up to 26 mg/l 4.2.3.24 amorpha-4,11-diene synthase synthesis expression of enzyme plus mevalonate isoprenoid pathway genes from Saccharomyces cerevisiae in Escherichia coli results in synthesis of amorpha-4,11-diene up to 0.024 mg caryophyllene per ml 4.2.3.24 amorpha-4,11-diene synthase synthesis production of amorpha-4,11-diene, expression in Nicotiana tabacum gives 0.2 to 1.7 ng per g fresh weight 4.2.3.27 isoprene synthase synthesis construction of a synthetic pathway of isoprene in Escherichia coli by introducing an isoprene synthase gene from Populus nigra and overexpression of the native or Bacillus subtilis 1-deoxy-D-xylulose-5-phosphate synthase gene and 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene. Transformants expressing ispS alone accumulate around 94 mg isoprene per liter of broth.Transformants expressing ispS plus the native 1-deoxy-D-xylulose-5-phosphate synthase gene and 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene produce 160 mg isoprene per liter. Transformants expressing ispS plus Bacillus subtilis 1-deoxy-D-xylulose-5-phosphate synthase gene and 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene produce 314 mg isoprene per liter 4.2.3.27 isoprene synthase synthesis heterologous expression of the isoprene synthase gene in the cyanobacterium Synechocystis PCC 6803 in combination with mevalonate pathway enzymes results in photosynthetic isoprene yield improvement by approximately 2.5fold, compared with that measured in cyanobacteria transformed with the isoprene synthase gene only. The mevalonate pathway introduces a bypass in the flux of endogenous cellular substrate in Synechocystis to isopentenyl diphosphate and dimethylallyl diphosphate, overcoming flux limitations of the native methylerythritol-phosphate pathway 4.2.3.27 isoprene synthase synthesis production of isoprene in Escherichia coli by improvement of the efficiency of the mevalonate pathway and expression of isoprene synthase. The final genetic strain containing the optimized mevalonate pathway and isoprene synthase from Populus alba can accumulate isoprene up to 6.3 g/l after 40 h of fed-batch cultivation 4.2.3.B31 manool synthase synthesis synthesis of sclareol by co-expression of producing labda-13-en-8-ol diphosphate as major product and manool synthase in Saccharomyces cerevisiae 4.2.3.32 levopimaradiene synthase synthesis increase of levopimaradiene synthesis in Escherichia coli by amplification of the flux toward isopentenyl diphosphate and dimethylallyl diphosphate precursors and reprogramming the rate-limiting downstream pathway by generating combinatorial mutations in geranylgeranyl diphosphate synthase and levopimaradiene synthase. The most productive pathway, combining precursor flux amplification and mutant synthases, confers approximately 2600fold increase in levopimaradiene levels. A maximum titer of approximately 700 mg/l is obtained by cultivation in a benchscale bioreactor 4.2.3.32 levopimaradiene synthase synthesis the development of yeast strains carrying the engineered Erg20p, which support efficient isoprenoid production, e.g. by abietadiene synthase, and can be used as a dedicated chassis for diterpene production or biosynthetic pathway elucidation. The design developed can be applied to the production of any GGPP-derived isoprenoid and is compatible with other yeast terpene production platforms, method overview 4.2.3.36 terpentetriene synthase synthesis CYC2 is a promising tool for introducing a buta-1,3-diene moiety into molecules by the enzymatic reactions 4.2.3.42 aphidicolan-16beta-ol synthase synthesis synthesis of aphidicolin, inhibitor of DNA polymerase alpha, by heterologous expression of the genes encoding geranlygeranyl diphosphate synthase, aphidicolan-16beta-ol synthase, and monooxygenases P450-1 and P450-1 from Phoma betae in Aspergillus oryzae. Host Saccharomyces cerevisiae, carrying the genes encoding geranlygeranyl diphosphate synthase and aphidicolan-16beta-ol, produces aphidicolan-16beta-ol 4.2.3.46 alpha-farnesene synthase synthesis expression in Sacchaomyces cerevisiae leads to a titer of (-)-alpha-copaene of 9 mg/l at 48 h and around 7 mg/l in Eschrichia coli, and the titer of (+)-delta-cadinene is 6 mg/l in Saccharomyces cerevisiae and 3.5 mg/l in Escherichia coli 4.2.3.61 5-epiaristolochene synthase synthesis construction of fusion proteins with farnesyl diphosphate synthase FPPS from Artemisia annua. The fusion enzymes produce epi-aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, are essentially the same for the single and fused enzymes. The bifunctional enzymes show a more efficient conversion of isopentenyl diphosphate to epi-aristolochene than the corresponding amount of single enzymes 4.2.3.61 5-epiaristolochene synthase synthesis expression system in Escherichia coli, highest expression level after 3 h induction with low concentration of IPTG and incubation at 27°C. Epiaristolochene synthase protein constitutes up to 35% of total Escherichia coli proteins 4.2.3.68 beta-eudesmol synthase synthesis introduction of a gene cluster encoding six enzymes of the mevalonate pathway into Escherichia coli and coexpression with ZSS2. When supplemented with mevalonate, the engineered Escherichia coli produces a similar sesquiterpene profile to that produced in the in vitro enzyme assay, and the yield of beta-eudesmol reaches 100 mg/l 4.2.3.70 patchoulol synthase synthesis construction of several fusion protein variants in which farnesyl diphosphate synthase of yeast has been coupled to patchoulol synthase of plant origin. Expression of the fusion proteins in Saccharomyces cerevisiae increases the production of patchoulol up to 2-fold. The fusion strategy can be used in combination with traditional metabolic engineering to further increase the production of patchoulol 4.2.3.70 patchoulol synthase synthesis fusion to thioredoxin and careful codon optimization of the eukaryotic sequence improves solubl expression on average by 42% in comparison to an unoptimized, His-tagged construct 4.2.3.70 patchoulol synthase synthesis use of a bifunctional enzymatic chimera composed as a fusion between farnesyl diphosphate synthase from yeast and patchoulol synthase from patchouli improves patchoulol production compared to that with the use of free enzymes up to 2fold. The effect is additive to improvements obtained by traditional metabolic engineering 4.2.3.73 valencene synthase synthesis 8fold improvement in the production of valencene, in yeast co-engineered with a truncated and deregulated HMG1, mitochondrion-targeted heterologous farnesyl diphosphate synthase and a mitochondrion-targeted sesquiterpene synthase,i.e.valencene synthase. Production of the Citrus sesquiterpene valencene in yeast is affected by deletion of geranylgeranyl diphosphate synthase BTS1 but not of phosphatases DPP1 or LPP1 4.2.3.73 valencene synthase synthesis expression of valencene synthase in Saccharomyces cerevisiae indicates potential for higher yields. In an optimized Rhodobacter sphaeroides strain, expression of valencene synthase increases valencene yields 14fold to 352 mg/l 4.2.3.73 valencene synthase synthesis heterologous expression of the (+)-valencene synthase gene in Corynebacterium glutamicum is not sufficient to enable (+)-valencene production, likely because provision of farnesyl diphosphate by endogenous prenyltransferases is too low. Upon deletion of two endogenous prenyltransferase genes and heterologous expression of either farnesyl diphosphate synthase gene ispA from Escherichia coli or ERG20 from Saccharomyces cerevisiae (+)-valencene production is observed. n-Dodecane is suitable for extraction of (+)-valencene from cultures and compatible with growth of Corynabacterium glutamicum. Production based on (+)-valencene synthase from Nootka cypress is superior to production by the enzyme from Citrus sinensis 4.2.3.73 valencene synthase synthesis Introduction of a valencene synthase gene into mushroom-forming fungus Schizophyllum commune results in production of the sesquiterpene (+)-valencene, both in mycelium and in fruiting bodies. Levels of (+)-valencene in culture media of strains containing a mutated RGS regulatory protein gene are increased fourfold compared to those in wild-type transformants. Up to 16 mg l/1 (+)-valencene can be produced. The amount of (+)-valencene containing n-dodecane recovered from the culture medium increases six- to sevenfold in the mutant strains due to the absence of schizophyllan 4.2.3.77 (+)-germacrene D synthase synthesis improved conditions for expression of Zingiber officinale (+)-germacrene synthase in Escherichia coli. Comparison of bacterial strains; BL21 (DE3), BL21 (DE3) Tuner BL21(DE3) pLysS and BL21 (DE3) pLysS Tuner using different inducing agents. The change between BL21 (DE3) cells and BL21 (DE3) Tuner, induced with IPTG, leads to a twofold increase in enzyme activity in the soluble fraction while a reduction in activity is observed when using the pLysS strains. The same doubling of activity is observed for germacrene synthase when the commonly used BL.21 (DE3) is induced with The Inducer. Addition of 2.5 mM glycine betaine and 660 mM sorbitol to the bacterial growth media results in reduction of growth rate and biomass yield but under these conditions the best overall protein production is obtained 4.2.3.82 alpha-santalene synthase synthesis construction of a Saccharomyces cerevisiae strain capable of producing high levels of alpha-santalene through a rationally designed metabolic engineering approach.Optimal sesquiterpene production is obtained by modulating the expression of one of the keymetabolic steps of the mevalonate pathway, squalene synthase Erg9. Further optimization yields a final titer of 92 mg/l of alpha-santalene 4.2.3.84 10-epi-gamma-eudesmol synthase synthesis introduction of a gene cluster encoding six enzymes of the mevalonate pathway into Escherichia coli and coexpression with ZSS2. When supplemented with mevalonate, the engineered Escherichia coli produces a similar sesquiterpene profile to that produced in the in vitro enzyme assay, and the yield of beta-eudesmol reaches 100 mg/l 4.2.3.85 alpha-eudesmol synthase synthesis introduction of a gene cluster encoding six enzymes of the mevalonate pathway into Escherichia coli and coexpression with ZSS2. When supplemented with mevalonate, the engineered Escherichia coli produces a similar sesquiterpene profile to that produced in the in vitro enzyme assay, and the yield of beta-eudesmol reaches 100 mg/l 4.2.3.95 (-)-alpha-cuprenene synthase synthesis expression in Xanthophyllomyces dendrorhous leads to production of alpha-cuprenene up to 80 mg/l. At this expression levels the pool of terpene precursors is not a limiting factor since the expression of the Cop6 gene in the genomic rDNA of the yeast allows production of both alpha-cuprenene and astaxanthin without affecting the growth or the accumulation levels of both compounds 4.2.3.104 alpha-humulene synthase synthesis chemolithoautotrophic production of a terpene from carbon dioxide, hydrogen, and oxygen, involving the engineered Cupriavidus necator cells and enzyme alpha-humulene synthase, is a promising starting point for the production of different high-value terpene compounds from abundant and simple raw materials. The production system is used to produce 17 mg alpha-humulene per gram cell dry mass (CDW) from CO2 and electrical energy in microbial electrosynthesis (MES) mode 4.2.3.114 gamma-terpinene synthase synthesis recombinant expression in Aspergillus nidulans resulting in production of gamma-terpinene. Aspergillus nidulans is capable of heterologous terpene production and thus has potential as a production host for industrially relevant compounds, e.g. gamma-terpinene 4.2.3.119 (-)-alpha-pinene synthase synthesis co-expression with over-expressed native 1-deoxy-D-xylulose-5-phosphate synthase and isopentenyl diphosphate isomerase from Corynebacterium glutamicum plus geranyl diphosphate synthase from Abies grandis leads to synthesis of up to 27 microg alpha-pinene per g cell weight 4.2.3.131 miltiradiene synthase synthesis expression of copalyl diphosphate synthase and miltiradiene synthase in Saccharomyces cerevisiae, results in production of 4.2 mg/l miltiradiene. Miltiradiene production increases to 8.8 mg/l by improving supply of geranylgeranyl diphosphate by over-expression of a fusion gene of farnesyl diphosphate synthase ERG20 and endogenous geranylgeranyl diphosphate synthase BTS1 together with a heterologous geranylgeranyl diphosphate synthase from Sulfolobus acidocaldarius. Combinatorial overexpression of truncated 3-hydroxyl-3-methylglutaryl-CoA reductase with a mutated global regulatory factor upc2.1 and ERG20-BTS1-geranylgeranyl diphosphate synthase genes has synergetic effects on miltiradiene production, increasing titer to 61.8 mg/l. In fed-batch fermentation 488 mg/l miltiradiene can be produced 4.2.3.133 alpha-copaene synthase synthesis expression in Saccharomyces cerevisiae leads to a titer of (-)-alpha-copaene of 9 mg/l at 48 h and around 7 mg/l in Escherichia coli, and the titer of (+)-delta-cadinene is 6 mg/l in Saccharomyces cerevisiae and 3.5 mg/l in Escherichia coli 4.2.3.135 DELTA6-protoilludene synthase synthesis protoilludene is a valuable sesquiterpene and serves as a precursor for several medicinal compounds and antimicrobial chemicals. It can be synthesized by heterologous overexpression of protoilludene synthase in Escherichia coli with coexpression of mevalonate or methylerythritol-phosphate pathway enzymes, and farnesyl diphosphate synthase as a cell factory for protoilludene production 4.2.3.141 sclareol synthase synthesis synthesis of sclareol by co-expression of producing labda-13-en-8-ol diphosphate as major product and manool synthase in Saccharomyces cerevisiae 4.2.3.141 sclareol synthase synthesis as biotechnological production systems for sesquiterpenoids, a number of diterpenoid engineering platforms are described that use plant diTPSs (and in some cases P450s) to produce, for example, the fragrance precursors sclareol and cis-abienol in the prefume industry. Sclareol from clary sage (Salvia sclarea) is used as a precursor for ambroxide fragrance and fixatives in perfume manufacture 4.2.3.146 cyclooctat-9-en-7-ol synthase synthesis in vivo CotB2 W288G reconstitution in an Escherichia coli based terpene production system allows efficient production of (1R,3E,7E,11S,12S)-3,7,18-dolabellatriene, which displays antibacterial activity against multi-drug resistant Staphylococcus aureus (MRSA) 4.2.3.205 sodorifen synthase synthesis expression of the sodorifen biosynthetic gene clusterSodABCD in Escherichia coli with a very low volatile organic compound background leads to a significant increase in both sodorifen product yield and purity compared to the native producer 4.2.99.21 isochorismate lyase synthesis alternative computational rational approach to improve the secondary catalytic activity of enzymes, taking as a test case the IPL enzyme. The approach is based on the use of molecular dynamic simulations employing hybrid quantum mechanics/molecular mechanics methods that allow describing breaking and forming bonds 4.2.99.22 tuliposide A-converting enzyme synthesis facile method of enzyme-mediated conversion of 6-tuliposide to alpha-methylene-gamma-butyrolactone, i.e.tulipalin by use of a tuliposide-converting enzyme for the conversion of 6-tuliposides. 6-Tuliposides are extracted from tulip tissues into the corresponding tulipalins in high yields within 2 h at pH 7.0. The resulting tulipalins are selectively extracted by using several organic solvents 4.2.99.22 tuliposide A-converting enzyme synthesis PaA and PaB are considered to be useful natural products with potential applications as synthetic intermediates in the production of several bioactive compounds, antimutagenic agents, insect repellents, and as monomers of functional biobased polymers, application of TCE to produce PaB, overview 4.2.99.23 tuliposide B-converting enzyme synthesis PaA and PaB are considered to be useful natural products with potential applications as synthetic intermediates in the production of several bioactive compounds, antimutagenic agents, insect repellents, and as monomers of functional biobased polymers, application of TCE to produce PaB, overview 4.3.1.1 aspartate ammonia-lyase synthesis industrial production of L-aspartic acid using polyurethane-immobilized cells containing aspartase 4.3.1.1 aspartate ammonia-lyase synthesis enzyme or whole cells are used for the industrial production of L-aspartate, which together with L-phenylalanine is the basis of the sweetener aspartame 4.3.1.1 aspartate ammonia-lyase synthesis production of L-aspartic acid, which is used as a precursor for the synthesis of the low calorie synthetic sweetener aspartame 4.3.1.1 aspartate ammonia-lyase synthesis production of L-aspartic acid, which is used as an intermediate for aspartame, an artificial sweetener. It is also used as acidulant in pharmaceuticals and foods 4.3.1.1 aspartate ammonia-lyase synthesis construction of hybrid enzyme from alpha-aspartyl dipeptidase and L-aspartase. The hybrid enzyme can be used in synthesis of the precursor for aspartame. Synthesizing aspartate from fumarate and NH4+, and then taking advantage of the catalytic action of alpha-aspartyl dipeptidase, the precursor of aspartame is produced 4.3.1.1 aspartate ammonia-lyase synthesis bioproduction of L-aspartic acid and cinnamic acid. From an initial concentration of 1000 mM of fumarate and 30 mM of L-phenylalanine, the enzyme converts 0.395 mM L-aspartic acid and 3.47 mM cinnamic acid, respectively 4.3.1.2 methylaspartate ammonia-lyase synthesis enzyme is used for L-aspartic acid production 4.3.1.2 methylaspartate ammonia-lyase synthesis starting from simple non-chiral dicarboxylic acids (either fumaric acid or mesaconic acid), vitamin B5 and both diastereoisomers of alpha-methyl-substituted vitamin B5, which are valuable precursors for promising antimicrobials against Plasmodium falciparum and multidrug-resistant Staphylococcus aureus, can be generated in good yields (up to 70%) and excellent enantiopurity (>99% ee). Access to vitamin B5 ((R)-pantothenic acid) and both diastereoisomers of alpha-methyl-substituted vitamin B5 ((R)- and (S)-3-((R)-2,4-dihydroxy-3,3-dimethylbutanamido)-2-methylpropanoic acid) is achieved using a modular three-step biocatalytic cascade involving 3-methylaspartate ammonia lyase (MAL), aspartate-a-decarboxylase (ADC), beta-methylaspartate-alpha-decarboxylase (CrpG) or glutamate decarboxylase (GAD), and pantothenate synthetase (PS) enzymes 4.3.1.3 histidine ammonia-lyase synthesis enzyme can be successfully microcapsulated within cellulose nitrate artificial cells 4.3.1.16 threo-3-hydroxy-L-aspartate ammonia-lyase synthesis the recombinant enzyme expressed in Escherichia coli is used to produce optically pure L-erythro-3-hydroxyaspartate and D-threo-3-hydroxyaspartate from the corresponding DL-racemic mixtures 4.3.1.19 threonine ammonia-lyase synthesis production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from a single unrelated carbon source via threonine biosynthesis in Escherichia coli, by overexpression of threonine deaminase, which is the key factor for providing propionyl-coenzyme A (propionyl-CoA), from different host bacteria, removal of the feedback inhibition of threonine by mutating and overexpressing the thrABC operon in Escherichia coli, and knock-out of the competitive pathways of catalytic conversion of propionyl-CoA to 3-hydroxyvaleryl-CoA. Construction of a series of strains and mutants leads to production of the poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer with differing monomer compositions in a modified M9 medium supplemented with 20 g/liter xylose. The largest 3-hydroxyvalerate fraction obtained in the copolymer is 17.5 mol% 4.3.1.19 threonine ammonia-lyase synthesis the mutant L-threonine deaminase G323D/F510L/T344A together with thermostable L-leucine dehydrogenase from Bacillus sphaericus DSM730 and formate dehydrogenase from Candida boidinii constitute a one-pot system for biosynthesis of L-2-aminobutyric acid 4.3.1.23 tyrosine ammonia-lyase synthesis biotransformation of tyrosine to p-coumaric acid reaching a maximum 2.4fold increase when 0.5 mM tyrosine is supplemented to the growth medium with Synechocystis PCC 6803 expressing tyrosine ammonia-lyase from Rhodobacter sphaeroides under Ptrc1O promoter 4.3.1.23 tyrosine ammonia-lyase synthesis by expressing Rhodobacter sphaeroides tyrosine ammonia lyase, in Streptomyces mobaraense, which permits the synthesis of p-coumaric acid from glucose, a strain is obtained that produces high amounts of 4-vinylphenol 4.3.1.23 tyrosine ammonia-lyase synthesis for 4-vinylphenol production directly from cellulose, L-tyrosine ammonia lyase derived from Rhodobacter sphaeroides and phenolic acid decarboxylase from Streptomyces sviceus are introduced into endoglucanase-secreting Streptomyces lividans, and the 4-vinylphenol biosynthetic pathway is constructed therein. The created transformants successfully produce 4-vinylphenol directly from cellulose 4.3.1.24 phenylalanine ammonia-lyase synthesis production of L-phenylalanine, which is used in the manufacture of the artificial sweetener aspartame and in parenteral nutrition, it is also used as a building block for the synthesis of the macrolide antibiotic rutamycin B 4.3.1.24 phenylalanine ammonia-lyase synthesis use of enzyme for production of enantiopure D- and L-heteroaryl-2-alanines, i.e. R- and S-2-amino-3-(heteroaryl)propanoic acids 4.3.1.24 phenylalanine ammonia-lyase synthesis phenylalanine ammonia-lyase's reverse reaction is exploited for the commercial production of optically pure L-phenylalanine from trans-cinnamic acid 4.3.1.24 phenylalanine ammonia-lyase synthesis the enzyme is involved in and useful for production salidroside, an effective adaptogenic drug from the medicinal plant Rhodiola sachalinensis 4.3.1.24 phenylalanine ammonia-lyase synthesis the enzyme is useful for an economic way for biosynthesis of 15NL-phenylalanine, yield and purity of 15NL-phenylalanine reach 71% and 99.3%, respectively 4.3.1.24 phenylalanine ammonia-lyase synthesis heterologous expression of enzyme in Streptomyces lividans. After 4 days of cultivation using glucose as carbon source, the maximal level of cinnamic acid reaches 210 mg/l. When glycerol is used as carbon source az 30 g/l, the maximal level of produced cinnamic acid reaches 450 mg/l. Using raw starch, xylose or xylan as carbon source, the maximal level of cinnamic acid reaches 460, 300, and 130 mg/l, respectively 4.3.1.24 phenylalanine ammonia-lyase synthesis improvement of recombinant phenylalanine ammonia-lyase stability in Escherichia coli during the enzymatic methods of L-phenylalanine production. The optimum values for testing variables are 13.04 mM glycerol, 1.87 mM sucrose, 4.09 mM DTT, and 69 mM Mg2+. The maximum phenylalanine ammonia-lyase activity is retained as 67.73 units/g after three successive cycles of bioconversion. In comparison to initial phenylalanine ammonia-lyase activity, the loss of phenylalanine ammonia-lyase activity was only 22%. Phenylalanine ammonia-lyase activity is enhanced about 23% in comparison to the control 4.3.1.24 phenylalanine ammonia-lyase synthesis AtPAL2 is a very good catalyst for the formation of 3-fluoro-L-phenylalanine, 4-fluoro-L-phenylalanine and 2-chloro-L-phenylalanine. Such noncanonical amino acids are valuable building blocks for the formation of various drug molecules 4.3.1.24 phenylalanine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including (2S)-pinocembrin, (2S)-naringenin, p-hydroxystyrene, p-coumarate, trans-cinnamic acid 4.3.1.24 phenylalanine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including dicinnamoylmethane, 6-fluoro-dicinnamoylmethane, 6,6'-difluoro-dicinnamoylmethane or pinosylvin 4.3.1.24 phenylalanine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including trans-cinnamic acid 4.3.1.24 phenylalanine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including trans-cinnamic acid, (2RS)-pinocembrin, styrene, pinosylvin or (2S)-naringenin 4.3.1.24 phenylalanine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including trans-cinnamic acid, p-coumaric acid, (2RS)-pinocembrin, (2RS)-naringenin, trans-resveratrol, pinosylvin, genistein 4.3.1.24 phenylalanine ammonia-lyase synthesis synthesis of analogues of L-phenylalanine, that are incorporated as pharmacophores in several peptidomimetic drug molecules and are therefore of particular interest to the fine chemical industry. Engineering of phenylalanine ammonia lyase from Rhodotorula graminis and identification of variants with very high levels of activity towards a panel of substituted cinnamic acids including; 4-bromo-, 3-bromo-, 4-fluoro-, 3-fluoro- and 3-nitro-cinnamic acid. Optimisation studies for use of one of these variants in the preparative synthesis of related variants of L-phenylalanine. Identification of variants used in a preparative scale biotransformation resulting in a 94% conversion to L-4-Br-phenylalanine (more than 99% enantiomeric excess) 4.3.1.24 phenylalanine ammonia-lyase synthesis synthesis of substituted D-phenylalanines in high yield and excellent optical purity, starting from inexpensive cinnamic acids, is achieved with a one-pot approach by coupling phenylalanine ammonia lyase amination with a chemoenzymatic deracemization (based on stereoselective oxidation and nonselective reduction). A simple high-throughput solid-phase screening method is developed to identify phenylalanine ammonia lyases with higher rates of formation of non-natural D-phenylalanines. The best variants are exploited in the chemoenzymatic cascade, thus increasing the yield and enantiomeric excess value of the D-configured product 4.3.1.24 phenylalanine ammonia-lyase synthesis the enzyme is specifically advantageous for the production of 2-chloro-L-phenylalanine, an important intermediate for the synthesis of angiotensin 1-converting enzyme inhibitors 4.3.1.24 phenylalanine ammonia-lyase synthesis the recombinant ZmPAL2 is a good candidate for the production of trans-cinnamic acid. The recombinant ZmPAL2 can effectively catalyze L-phenylalanine to trans-cinnamic acid, and the trans-cinnamic acid concentration can reach up to 5 g/l 4.3.1.25 phenylalanine/tyrosine ammonia-lyase synthesis immobilization of Escherichia coli cells stably expressing the enzyme on calcium alginate beads for use in batch coversion of L-tyrosine to p-hydroxycinnamic acid. Immobilization and controlling of pH value to 9.8 results in stabilization. In 1 l batch reactions, 50 g/l tyrosine can be converted to 39 g/l p-hydroxycinnamic acid 4.3.1.25 phenylalanine/tyrosine ammonia-lyase synthesis use of enzyme for synthesis of optically pure L-phenylalanine from trans-cinnamate 4.3.1.25 phenylalanine/tyrosine ammonia-lyase synthesis reconstructed phenylpropanoid pathway in engineered Escherichia coli or Saccharomyces cerevisiae leads to the biosynthesis of a wide range of phenylpropanoid-derived compounds, including pinocembrin, naringenin, styrene, 2',4',6'-trihydroxydihydrochalcone, trans-resveratrol, trans-cinnamic acid, p-coumarate, p-hydroxystyrene 4.3.1.25 phenylalanine/tyrosine ammonia-lyase synthesis synthesis of p-hydroxycinnamic acid methyl ester, which shows antibacterial activity 4.3.1.27 threo-3-hydroxy-D-aspartate ammonia-lyase synthesis optically pure synthesis of L-threo-3-hydroxyaspartate and D-threo-3-hydroxyaspartate, promising intermediates for the synthesis of beta-benzyloxyaspartate by using a purified enzyme as a biocatalyst for the resolution of racemic DL-threo-3-hydroxyaspartate and DL-erythro-3-hydroxyaspartate. Considering 50% of the theoretical maximum, efficient yields of L-threo-3-hydroxyaspartate (38.9 %) and D-erythro-3-hydroxyaspartate (48.9 %) as isolated crystals are achieved with more than 99 % enantiomeric excess 4.3.3.2 strictosidine synthase synthesis immobilized enzyme can be used to synthesize large quantities of 3-alpha(S)-strictosidine, that is a common intermediate in the biosynthesis of indole alkaloids 4.3.3.2 strictosidine synthase synthesis involved in the biosynthesis of almost all plant monoterpenoid indole alkaloids 4.3.3.2 strictosidine synthase synthesis decreased production of campothecin on subculturing due to a cross-species biosynthetic pathway is where the endophyte Fusarium solani utilizes indigenous geraniol 10-hydroxylase, secologanin synthase, and tryptophan decarboxylase to biosynthesize precursors. Fusarium solani requires host Campotheca acuminata strictosidine synthase in order to form strictosidine. Biosynthetic genes of campothecin in the seventh subculture generation of the endophyte reveal random and unpredictable nonsynonymous mutations. These random base substitutions lead to dysfunction at the amino acid level. The controls, Top1 gene and rDNA, remained intact over subculturing 4.3.3.2 strictosidine synthase synthesis production of campothecin in hairy root culture of Ophiorrhiza pumila by infection with Agrobacterium tumefaciens. The culture also produces a high level of anthraquinones besides camptothecin. Camptothecin is present not only in hairy root cells, but excreted out to medium at a substantial amount. Camptothecin content in the medium is increased by the presence of a polystyrene resin that absorbs camptothecin 4.3.3.2 strictosidine synthase synthesis overexpression of tryptophan decarboxylase and strictosidine synthase enhances terpenoid indole alkaloid pathway activity and antineoplastic vinblastine biosynthesis in Catharanthus roseus. Vinblastine and vincristine are highly expensive antineoplastic molecules 4.3.3.7 4-hydroxy-tetrahydrodipicolinate synthase synthesis the enzyme can be commercially exploited for high-yield production of (S)-lysine 4.4.1.1 cystathionine gamma-lyase synthesis effective at the end of trans-sulfuration pathway 4.4.1.1 cystathionine gamma-lyase synthesis effective in detoxification and biotransformation of Se 4.4.1.1 cystathionine gamma-lyase synthesis effective in metabolism of Se compounds 4.4.1.4 alliin lyase synthesis immobilized alliinase in reversibly soluble N-succinyl-chitosan is suitable to catalyze the conversion of alliin to allicin, as active ingredient of pharmaceutical compositions and food additive 4.4.1.11 methionine gamma-lyase synthesis presence of a hybrid plasmid in Escherichia coli K12 containing the enzyme gene leads to a decrease in efficiciency of EcoKI restriction 4.4.1.17 Holocytochrome-c synthase synthesis mutants E159A and W118A display enhanced release of cytochrome c from the active site. The mutant allows for synthesis of cytochrome c variants such as cyt c H19M (bis-Met), cyt c M81H (bis-His), cyt c M81A (His/OH), cyt c C15S, which exhibit proper folding 4.4.1.26 olivetolic acid cyclase synthesis synthesis of olivetolic acid from a single carbon source through combining OLA synthase and OLA cyclase expression with the required modules of a beta-oxidation reversal for hexanoyl-CoA generation. The use of additional auxiliary enzymes to increase hexanoyl-CoA and malonyl-CoA, along with evaluation of varying fermentation conditions, enables the synthesis of 80 mg/L olivetolic acid 4.4.1.29 phycobiliprotein cysteine-84 phycobilin lyase synthesis simultaneous expression of the phycocyanobilin-producing genes hox1 and pcyA of Arthrospira platensis with the phycobiliprotein gene cpcB and the lyase gene cpcS/U in Escherichia coli. Phycobiliprotein cpcB is successfully synthesized in Escherichia coli and co-expressed phycocyanobilins are attached though at a relatively low efficiency. The attachment of phycocyanobilins to phycobiliprotein cpcB is carried out mainly by co-expressed lyase cpcS/U but cpcB also shows some autocatalytic activity 4.4.1.32 C-phycocyanin alpha-cysteine-84 phycocyanobilin lyase synthesis production of of a fluorescent holo-C-phycocyanin subunit alpha in Escherichia coli by construction of an expression vector containing five essential genes in charge of biosynthesis of cyanobacterial C-phycocyanin holo-alpha subunit. The vector harbors two cassettes: one cassette carries genes hox1 and pcyA required for conversion of heme to phycocyanobilin, and the other cassette carries cpcA encoding C-phycocyanin subunit alpha along with lyase subunits cpcE and cpcF both of which are necessary and sufficient for the correct addition of phycobiliprotein to CpcA. The maximum peak of absorbance spectrum is at 623 nm, and the maximum peak of fluorescence emission and excitation are at 648 and 633 nm, respectively, which are similar to those of native C-phycocyanin 4.4.1.32 C-phycocyanin alpha-cysteine-84 phycocyanobilin lyase synthesis reconstitution of the entire pathway for the synthesis of a fluorescent holophycobiliprotein subunit from Synechocystis sp. PCC6803 in Escherichia coli. Heme oxygenase 1 and 3Z-phycocyanobilin:ferredoxin oxidoreductase are expressed from one plasmid. Genes for the apoprotein C-phycocyanin a subunit, cpcA and the heterodimeric lyase subunit cpcE and cpcF that catalyze chromophore attachment are expressed from a second plasmid. Upon induction, recombinant Escherichia coli uses the cellular pool of heme to produce holo-CpcA with spectroscopic properties qualitatively and quantitatively similar to those of the same protein produced endogenously in cyanobacteria. About a third of the apo-CpcA is converted to holo-CpcA 4.4.1.36 hercynylcysteine S-oxide lyase synthesis the ergothioneine biosynthetic pathway (EgtA-EgtE catalysis) provides an opportunity for ergothioneine production through metabolic engineering 4.6.1.12 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase synthesis preparation of 4-diphosphocytidyl-2C-methyl-D-erythritol 2-phosphate in multigram quantities 4.6.1.18 pancreatic ribonuclease synthesis expression of enzyme in Escherichia coli and Saccharomyces cerevisiae. Recombinant proteins are indistinguishable from the Sulfolobus solfataricus enzyme on the basis of heat stability, pH optimum and RNA digestion pattern as well as NMR analysis, the only exceptions being that residues K4 and K6 are not methylated in the recombinant enzyme 4.8.1.2 aliphatic aldoxime dehydratase synthesis may be a useful biocatalyst for the production of various nitriles from the corresponding aldoximes 4.8.1.2 aliphatic aldoxime dehydratase synthesis enzyme is able to catalyze the non-natural Kemp elimination reaction 4.8.1.2 aliphatic aldoxime dehydratase synthesis mutant H320D converts 1-methoxynaphthalene in a peroxidase reaction to 1-methoxy-2-naphtalenol, i.e. Russig's blue 4.8.1.4 phenylacetaldoxime dehydratase synthesis high yield synthesis of 3-phenylpropionitrile from unpurified (E/Z)-3-phenylpropionaldoxime, which is spontaneously formed from 3-phenlypropionaldehyde and hydroxylamine in a butyl acetate/water biphasic system, production of further nitriles from their corresponding aldoximes 4.8.1.4 phenylacetaldoxime dehydratase synthesis production of enzyme by expression in Escherichia coli and Bacillus subtilis, at 37°C, yield of mainly inactive inclusion bodies, at 30°C, enzyme is largely soluble and active. Enhancement of production by increasing the volume of culture medium 5.1.1.3 glutamate racemase synthesis production of D-glutamate from L-glutamate with glutamate racemase and L-glutamate oxidase from Streptomyces sp. X119-6. Both enzymes are highly stabilized by immobilization 5.1.1.10 amino-acid racemase synthesis enzymatic production of L-Trp from DL-Ser and indole by a coupled reaction of tryptophan synthase and amino acid racemase 5.1.1.10 amino-acid racemase synthesis the enzyme might be useful for applications in dynamic kinetic resolution for D- or L-amino acid production 5.1.1.10 amino-acid racemase synthesis the high reactivity toward Trp and Tyr, as well as extremely high thermostability, is likely a major advantage of using BAR for biochemical conversion of these aromatic amino acids 5.1.1.11 phenylalanine racemase (ATP-hydrolysing) synthesis production of phenylalanine racemase in continuous culture 5.1.1.15 2-aminohexano-6-lactam racemase synthesis Achromobacter obae which produces alpha-amino-epsilon-caprolactam racemase, is utilized in industry to produce L-Lys from DL-alpha-amino-epsilon-caprolactam with a high yield in the presence of Cryptococcus laurentii, an L-alpha-amino-epsilon caprolactamase producing yeast 5.1.1.15 2-aminohexano-6-lactam racemase synthesis production of DL-alpha-amino-beta-caprolactam as nutrient and food supplement 5.1.1.15 2-aminohexano-6-lactam racemase synthesis the enzyme might be useful for applications in dynamic kinetic resolution for D- or L-amino acid production 5.1.1.17 isopenicillin-N epimerase synthesis epimerization is a rate-limiting step in cephalosporin biosynthesis in Acremonium chrysogenum. When epimerase activity is enhanced, there is a clear increase in cephalosporin production. Thus, this method provides a strategy that could easily be applied to improve industrial cephalosporin producing strains 5.1.2.2 mandelate racemase synthesis DEAE-cellulose-immobilized mandelate racemase can be efficiently used in repeated batch reactions for the racemization of (R)-mandelic acid under mild conditions 5.1.2.2 mandelate racemase synthesis mandelate racemase and mandelate dehydrogenase, coexpressed in Escherichia coli, are useful for the synthesis of benzoylformate by converting racemic mandelate 5.1.3.1 ribulose-phosphate 3-epimerase synthesis enzyme is involved in a cascade of eleven immobilized enzyme reactors in series used for the production of D-ribulose-1,5-bisphosphate from 3-phospho-D-glycerate 5.1.3.1 ribulose-phosphate 3-epimerase synthesis efficient utilization of the available glucose and xylose in the lignocellulosic hydrolysates is the important issue for economic cellulosic ethanol production. Simultaneous utilization of xylose is realized by the coupling of glucose metabolism and xylose utilization through RPE1 deletion in xylose-utilizing Saccharomyces cerevisiae 5.1.3.1 ribulose-phosphate 3-epimerase synthesis the enzyme overexpression is part of the engineering of Synechocystis 6803 for photosynthetic limonene production 5.1.3.2 UDP-glucose 4-epimerase synthesis preparation of a fusion enzyme consisting of UDP-galactose 4-epimerase and galactose-1-phosphate uridylyltransferase with an intervening Ala3 linker, shows kinetic advantages in that the initial velocity to produce glucose 1-phosphate from UDPgalactose an 5.1.3.3 Aldose 1-epimerase synthesis improvement of a bacterial L-arabinose utilization pathway consisting of L-arabinose isomerase from Bacillus licheniformis and L-ribulokinase and L-ribulose-5-phosphate 4-epimerase from Escherichia coli after expression of the corresponding genes in Saccharomyces cerevisiae. After adaptation of codon usage, yeast transformants show strongly improved L-arabinose conversion rates. The ethanol production rate from L-arabinose can be increased more than 2.5fold from 0.014 g ethanol per h and g dry weight to 0.036 g ethanol per h and g dry weight and the ethanol yield can be increased from 0.24 g ethanol per g consumed L-arabinose to 0.39 g ethanol per g consumed L-arabinose 5.1.3.4 L-ribulose-5-phosphate 4-epimerase synthesis improvement of a bacterial L-arabinose utilization pathway consisting of L-arabinose isomerase from Bacillus licheniformis and L-ribulokinase and L-ribulose-5-phosphate 4-epimerase from Escherichia coli after expression of the corresponding genes in Saccharomyces cerevisiae. After adaptation of codon usage, yeast transformants show strongly improved L-arabinose conversion rates. The ethanol production rate from L-arabinose can be increased more than 2.5fold from 0.014 g ethanol per h and g dry weight to 0.036 g ethanol per h and g dry weight and the ethanol yield can be increased from 0.24 g ethanol per g consumed L-arabinose to 0.39 g ethanol per g consumed L-arabinose 5.1.3.7 UDP-N-acetylglucosamine 4-epimerase synthesis method for the enzymatic synthesis of UDP-N-acetylgalactosamine from UDP-N-acetylglucosamine, which is commercially available and inexpensive 5.1.3.7 UDP-N-acetylglucosamine 4-epimerase synthesis large scale preparation of UDP-N-acetylgalactosamine from UDP-N-acetylglucosamine by means of microbial enzymes 5.1.3.8 N-acylglucosamine 2-epimerase synthesis production of N-acetyl-D-neuraminic acid by coupling bacteria expressing N-acetyl-D-glucosamine 2-epimerase and N-acetyl-D-neuraminic acid synthetase. Microbial production of N-acetylneuraminic acid is carried out using Escherichia coli overexpressing N-acylglucosamine 2-epimerase and NeuAc synthetase as enzyme sources. Phosphoenolpyruvate and ATP, required as a substrate or a cofactor of the enzymes, are supplied by the activities of Escherichia coli and Corynebacterium ammoniagenes cells. Starting with 800 mM GlcNAc and 360 mM glucose, NeuAc accumulates at 39.7 mM after 22 h 5.1.3.8 N-acylglucosamine 2-epimerase synthesis production of N-acetylneuraminic acid from N-acetylglucosamine and pyruvate using recombinant human renin binding protein showing GlcNAc-2-epimerase activity and sialic acid aldolase in a coupling reaction 5.1.3.8 N-acylglucosamine 2-epimerase synthesis BoAGE2 a promising biocatalyst for sialic acid production using cheap GlcNAc as starting material 5.1.3.8 N-acylglucosamine 2-epimerase synthesis recombinant Escherichia coli cells synchronously expressing N-acetyl-D-glucosamine-2-epimerase and N-acetyl-D-neuraminic acid aldolase as biocatalysts can potentially be used in the industrial mass production of Neu5Ac 5.1.3.11 cellobiose epimerase synthesis cellobiose 2-epimerase from Caldicellulosiruptor saccharolyticus is used in the presence of borate to increase the production of lactulose from lactose, method optimization, incubation at pH 7.5 and 80°C for 3 h, with a conversion yield of 88% and a productivity of 205 g/l/h, overview 5.1.3.11 cellobiose epimerase synthesis application of the enzyme for production of functional oligosaccharides, overview 5.1.3.11 cellobiose epimerase synthesis industrial application of Caldicellulosiruptor saccharolyticus cellobiose 2-epimerase (CsCE) for lactulose synthesis is limited by low enzyme activity and formation of epilactose as by-product, the mutant G4-C5, i.e. R5M/I52V/A12S/K328I/F231L, shows improved enzyme activity without compromised thermostability. The yield of lactulose catalyzed by mutant G4-C5 increases to approximately 76% with no obvious epilactose detected, indicating that mutant G4-C5 is more suitable for lactulose production than the wild-type enzyme. The mutant does not produc epilactose as side-product from lactose, in contrast to the wild-type enzyme. Although epilactose is also a bifidus factor, formation of epilactose can complicate lactulose purification 5.1.3.11 cellobiose epimerase synthesis industrial application of Caldicellulosiruptor saccharolyticus cellobiose 2-epimerase for lactulose synthesis is limited by low enzyme activity and formation of epilactose as by-product. The yield of lactulose catalyzed by mutant R5M/A12S/I52V/F231L/K328I increases to approximately 76% with no epilactose detected, indicating that the mutant is more suitable for lactulose production than the wild-type enzyme 5.1.3.11 cellobiose epimerase synthesis most efficient enzyme for the enzymatic synthesis of lactulose. Ethanol-permeabilised Escherichia coli cells containing the enzyme are used as biocatalysts for lactulose production. The reaction conditions for maximum lactulose production are optimised to be 600 g/l lactose, pH 7.5, 80°C and 12.5 U/ml of whole-cell biocatalyst. After incubated at Na2HPO4-NaH2PO4 buffer for 2 h, approximately 390.59 g/l lactulose is obtained with a conversion yield of 65.1%. The lowest production and conversion yield of epilactose are also found in Na2HPO4-NaH2PO4 buffer with a final concentration of 11.7 g/l and a conversion yield less than 2%. The results represent a promising technology to attain high production and conversion yield of lactulose with a high purity on industrial scale 5.1.3.11 cellobiose epimerase synthesis synthesis of lactulose, a non-digestible disaccharide widely used in food and pharmaceutical industries. Thermostability enhancement of cellobiose 2-epimerase from Caldicellulosiruptor saccharolyticus by site-directed mutagenesis. Compared to the wild-type enzyme the mutant enzyme E161D/N365P shows approximately 4fold increase in the t1/2 value at 80°C and a 1.3fold increase in catalytic efficiency for lactulose production 5.1.3.11 cellobiose epimerase synthesis the enzyme can be used for production of the probiotic lactulose 5.1.3.11 cellobiose epimerase synthesis the enzyme from Caldicellulosiruptor saccharolyticus is applied for epilactose and lactulose production in milk at lower temperatures 5.1.3.11 cellobiose epimerase synthesis the enzyme mutant Y114E can be used for production of lactulose for the nutritional industry. Using Y114E, isomerization of lactose to lactulose is optimnized, resulting in 86.9 g/l of lactulose and 4.6 g/l of epilactose for 2 h from 200 g/l of lactose 5.1.3.11 cellobiose epimerase synthesis the recombinant enzyme shows both epimerisation and isomerisation activities against lactose, making it an alternative promising biocatalyst candidate for lactulose production 5.1.3.B12 Agrobacterium tumefaciens D-psicose 3-epimerase synthesis the enzyme can be used for synthesis of D-psicose (D-ribo-2-hexulose or D-allulose), a C3 epimer of D-fructose and considered as a rare sugar 5.1.3.B12 Agrobacterium tumefaciens D-psicose 3-epimerase synthesis the enzyme from Agrobacterium tumefaciens can be used for production of D-psicose in a coexpression system with xylose isomerase gene from Escherichia coli. Method optimization and evaluation, overview 5.1.3.13 dTDP-4-dehydrorhamnose 3,5-epimerase synthesis potential application in altering the enzyme behavior for use in synthesis of bioactive carbohydrates 5.1.3.13 dTDP-4-dehydrorhamnose 3,5-epimerase synthesis the enzyme is part of an enzyme module system for the synthesis of dTDP-activated deoxysugars from dTMP and sucrose 5.1.3.14 UDP-N-acetylglucosamine 2-epimerase (non-hydrolysing) synthesis transgenic Arabidopsis thaliana plants expressing three key enzymes of the mammalian Neu5Ac biosynthesis pathway: UDPN-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase, N-acetylneuraminic acid phosphate synthase, and CMP-Nacetylneuraminic acid synthetase. Simultaneous expression of UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase and N-acetylneuraminic acid phosphate synthase results in the generation of significant Neu5Ac amounts of 1275 nmol per g fresh weight in leaves, which can be further converted to cytidine monophospho-N-acetylneuraminic acid by coexpression of CMP-N-acetylneuraminic acid synthetase 5.1.3.17 heparosan-N-sulfate-glucuronate 5-epimerase synthesis the enzyme, coexpressed with 2-O-sulfotransferase in bacteria, can be used for production of bioengineered heparin as a potential substitute for the animal-sourced anticoagulant drug 5.1.3.30 D-psicose 3-epimerase synthesis for the high production of D-psicose from D-fructose, the reaction reaches equilibrium after 160 min with the highest turnover yield (28.8%) observed at 55 °C 5.1.3.30 D-psicose 3-epimerase synthesis suitable for the industrial production of D-psicose from fructose 5.1.3.30 D-psicose 3-epimerase synthesis synthesis of the rare sugar D-psicose, that is an ideal sucrose substitute for food products, due to having 70% of the relative sweetness but 0.3% of the energy of sucrose. It also shows important physiological functions 5.1.3.30 D-psicose 3-epimerase synthesis useful in the bioproduction of D-psicose, a rare hexose sugar, from D-fructose, found plenty in nature 5.1.3.30 D-psicose 3-epimerase synthesis D-psicose 3-epimerase is the key enzyme for producing D-allulose from inexpensive D-fructose sugar 5.1.3.30 D-psicose 3-epimerase synthesis the enzyme is a potential D-psicose producer for industrial production 5.1.3.30 D-psicose 3-epimerase synthesis the recombinant Dorea sp. DPEase displays significantly higher specific activity at acidic pHs and remarkably higher productivity of D-psicose at pH 6.0, indicating that it is appropriate for use as a different source of D-psicose producing enzyme 5.1.3.30 D-psicose 3-epimerase synthesis Trpr-DPEase might be appropriate for the industrial production of D-psicose. Comparison of D-psicose productivity of various ketose 3-epimerases, overview 5.1.3.30 D-psicose 3-epimerase synthesis the recombinant Escherichia coli expressing D-psicose-3-epimerase (DPE), ribitol dehydrogenase (RDH) and formate dehydrogenase (FDH) is constructed and used together with immobilized GI for allitol bioproduction from D-glucose. The conditions of allitol biotransformation, the cell catalytic activity resistance, the cell cultivation medium, and fed-batch culture conditions are optimized 5.1.3.31 D-tagatose 3-epimerase synthesis rare sugar production 5.1.3.31 D-tagatose 3-epimerase synthesis D-tagatose 3-epimerase is a useful enzyme for the production of expensive, rare sugars, e.g. D-sorbose and D-psicose, from inexpensive sugars, e.g. D-tagatose and D-fructose 5.1.3.31 D-tagatose 3-epimerase synthesis enzyme PcDTE is used industrially to produce D-psicose from the more abundant sugar D-fructose 5.1.99.1 methylmalonyl-CoA epimerase synthesis production of polyketides like myxothiazol requiring methylmalonyl-CoA as an extender unit 5.1.99.1 methylmalonyl-CoA epimerase synthesis recombinant methylmalonyl-CoA epimerase enables biocatalytic epimerisation of C-2 methyl- and ethylmalonyl-CoA. When coupled with variant carboxymethylproline synthase CarB and crotonyl-CoA carboxylase reductase, the methylmalonyl-CoA epimerase forms part of a three-enzyme, one-pot procedure for the production of functionalised C-5 carboxyalkylprolines from crotonyl-CoA, carbon dioxide and L-glutamate semialdehyde/5-hydroxyproline/pyrroline-5-carboxylate 5.1.99.5 hydantoin racemase synthesis enzymatic production of D-amino acids via a combination of hydantoin hydrolysis and hydantoin racemization. The D-forms of amino acids D-phenylalanine, D-tyrosine, D-tryptophan, O-benzyl-D-serine, D-valine, D-norvaline, D-leucine and D-norleucine can efficiently be converted from the corresponding DL-5-monosubtituted hydantoin compounds 5.1.99.5 hydantoin racemase synthesis reaction system for the production of D-amino acids from D,L-5-monosubstituted hydantoins based on recombinant Escherichia coli whole cell biocatalysts containing separately expressed D-hydantoinase, D-carbamoylase, and hydantoin recemase. The system shows high substrate specificity and is effective toward both aliphatic and aromatic D,L-5-monosubstituted hydantoins. At pH 8 and 50°C, 100% conversion of D,L-5-(2-methylthioethyl)-hydantoin (15 mM) into D-methionine is obtained in 30 min 5.1.99.5 hydantoin racemase synthesis stereospecific production of L-amino acids from the corresponding DL-5-substituted hydantoins. DL-5-substituted hydantoins are converted exclusively to the L-forms of the corresponding N-carbamyl-amino acids by hydantoinase in combination with hydantoin racemase. The N-carbamyl-l-amino acids are then converted to L-amino acids by N-carbamyl-L-amino acid amidohydrolase 5.2.1.1 maleate isomerase synthesis whole cells of Escherichia coli expressing the enzyme mutant MaiR-D48 catalyze the isomerization of maleic acid to fumaric acid at 1 M substrate concentration showing its potential for industrial use 5.2.1.5 linoleate isomerase synthesis the enzyme can be useful for biocatalysis of linoleic acid to conjugated linoleic acid 5.2.1.5 linoleate isomerase synthesis method for converting plant oil into a specific conjugated linoleic acid using a synergistic biocatalytic system based on immobilized Propionibacterium acnes linoleate isomerase and Rhizopus oryzae lipase. Using linoleate isomerase immobilized on D301R anion-exchange resin, under optimal conditions up to 109 g/l trans-10,cis-12-linoleic acid are obtained after incubation of 200 g/l sunflower oil. The conversion ratio of linoleic acid to conjugated linoleic acid is 90.5% 5.2.1.5 linoleate isomerase synthesis production of trans-10,cis-12-linoleic acid as a single isomer by expression of a fusion construct with enhanced green fluorescent protein on the surface of yeast cells. Upon optimization, a maximum yield of trans-10,cis-12-linoleic acid of 4 mg/ml is observed within 20 h using the strain as whole-cell biocatalyst 5.2.1.5 linoleate isomerase synthesis production of trans-10,cis-12-linoleic acid by codon-optimized expression in Mortierella alpina. In presence of 5 microM long-chain acyl-CoA synthetase inhibitor the trans-10,cis-12-linoleic acid content reaches up to 29 mg/l 5.2.1.5 linoleate isomerase synthesis strain Lactobacillus plantarum ZS2058 is able to convert over 50% of linoleic acid to cis-9,trans-11-linoleate and trans9,trans-linoleate. Some intermediates 10-hydroxy-cis-12-octadecenoic acid, 10-oxo-cis-12-octadecenoic acid and 10-oxo-trans-11-octadecenoic acid are found. Expression of the genes coding the multicomponent linoleate isomerase containing myosin-cross-reactive antigen, short-chain dehydrogenase/oxidoreductase and acetoacetate decarboxylase in Escherichia coli leads to the synthesis of cis9,trans-11-linoleate and three kinds of intermediates 5.3.1.1 triose-phosphate isomerase synthesis enzymatic synthesis of D-xylulose 5-phosphate using triosephosphate isomerase and transketolase. Comparison of use of glyceraldehyde 3-phosphate or dihydroxyacetone phosphate as starting substrate by process modeling via the use of windows of operation 5.3.1.4 L-arabinose isomerase synthesis an Escherichia coli galactose kinase gene knockout strain, which contains the L-arabinose isomerase gene to isomerize D-galactose to D-tagatose, shows a higher conversion yield of tagatose because galactose is not metabolized by endogenous galactose kinase. In whole cells of the galactose kinase knockout strain, the isomerase-catalyzed reaction exhibits an equilibrium shift towards tagatose, producing a tagatose fraction of 68% at 37°C, whereas the purified L-arabinose isomerase gives a tagatose equilibriumfraction of 36% 5.3.1.4 L-arabinose isomerase synthesis expression of the Escherichia coli genes araA, araB, and araD encoding L-arabinose isomerase, L-ribulokinase, and L-ribulose-5-phosphate 4-epimerase, respectively, in Corynebacterium glutamicum under the control of a constitutive promoter. The recombinant strain is able to grow on mineral salts medium containing L-arabinose as the sole carbon and energy source. Under oxygen deprivation and with L-arabinose as the sole carbon and energy source, carbon flow of the recombinant strain is redirected to produce up to 40, 37, and 11%, respectively, of the theoretical yields of succinic, lactic, and acetic acids. Usinga sugar mixture containing 5% D-glucose and 1% L-arabinose under oxygen deprivation, cells metabolize L-arabinose at a constant rate, resulting in combined organic acids yield based on the amount of sugar mixture consumed after D-glucose depletion of 83% that is comparable to that before D-glucose depletion, 89% 5.3.1.4 L-arabinose isomerase synthesis production of D-tagatose from D-galactose by L-arabinose isomerase immobilized on chitopearl beads. Half-lives of immobilized enzyme at 70°C, 75°C, 80°C, 85°C and 90°C are 388, 106, 54, 36, and 22 h, respectively. With pH control at 7.5, D-tagatose production at 70°C in a stirred tank reactor doubles compared to conditions without pH control 5.3.1.4 L-arabinose isomerase synthesis production of ethanol after improvement of a bacterial L-arabinose utilization pathway consisting of L-arabinose isomerase from Bacillus licheniformis and L-ribulokinase and L-ribulose-5-phosphate-4-epimerase from Escherichia coli after expression of the corresponding genes in Saccharomyces cerevisiae.Yeast transformants expressing the codon-optimized genes show strongly improved L-arabinose conversion rates. The ethanol production rate from L-arabinose can be increased more than 2.5fold from 0.014 g ethanol per h and g dry weight to 0.036 g ethanol per h and g dry weight and the ethanol yield can be increased from 0.24 g ethanol per g consumed L-arabinose to 0.39 g ethanol per g consumed L-arabinose 5.3.1.4 L-arabinose isomerase synthesis production of the intracellular enzymes L-arabinose isomerase and D-xylose isomerase in Lactobacillus bifermentans. After 9 h cultivation in optimized medium, Arabinose isomerase and xylose isomerase activities are 9.4 and 7.24 U/ml, respectively. For optimal growth, the strain requires Tween 80 at 1 g/l and a source of inorganic nitrogen, e.g., ammonium citrate. The bacterium has no requirement for sodium acetate for either growth or production of isomerases. The production rate of enzymes is increased when metal ions are added, primarily manganese 5.3.1.4 L-arabinose isomerase synthesis strain carrying mutant C450S/N475K is able to produce 95 g L-ribulose per l from 500 g L-arabinose per l under optimum conditions of pH 8, 70°C, and 10 units enzyme per ml with a conversion yield of 19% over 2 h. The half-lives of the mutated enzyme at 70 and 75°C are 35 and 4.5 h, respectively 5.3.1.4 L-arabinose isomerase synthesis alginate-immobilized Escherichia coli cells, recombinantly expressing the L-arabinose isomerase from Bacillus licheniformis, shows high stability and are suitable for L-ribulose production at low costs 5.3.1.4 L-arabinose isomerase synthesis engineered enzyme mutants are useful for production of D-tagatose 5.3.1.4 L-arabinose isomerase synthesis L-ribose production of the enzyme in a coupled assay system with mannose-6-phosphate isomerase, EC 5.3.1.8, in recombinant Escherichia coli ER2566, AI/MPI ratio, 1:2.5, method optimization. L-Ribose is a potential starting material for the synthesis of many L-nucleoside-based pharmaceutical compounds 5.3.1.4 L-arabinose isomerase synthesis the enzyme is useful in production of low-calorie sweetener D-tagatose. Tagatose also may potentially be useful as a prescription drug additive, to mask unpleasant tastes, and as a sweetener in toothpaste, mouthwash, and cosmetics such as flavored lipstick 5.3.1.4 L-arabinose isomerase synthesis the enzyme is a biocatalyst for the production of L-ribulose 5.3.1.4 L-arabinose isomerase synthesis the enzyme is used for the commercial production of D-tagatose 5.3.1.4 L-arabinose isomerase synthesis coexpression of beta-D-galactosidase gene and L-arabinose isomerase mutant Q299K for synthesis of D-tagatose. Recombinant cells exhibit maximum D-tagatose producing activity at 34°C and pH 6.5 and in the presence of borate, 10 mM Fe2+, and 1 mM Mn2+. Cells can hydrolyze more than 95% lactose and convert 43% D-galactose into D-tagatose 5.3.1.4 L-arabinose isomerase synthesis coexpression with a thermostable beta-galactosidase from Thermus thermophilus HB27 in Escherichia coli. The recombinant cells show optimal catalytic temperature and pH at 70°C and 7.0, respectively. With the addition of borate, D-tagatose is produced directly from lactose in 16 h in a concentration of 101 g/l, a yield of 20.2%, and a productivity of 6.3 g/l/h 5.3.1.4 L-arabinose isomerase synthesis enzyme is displayed on the spore surface of Bacillus subtilis DB403 by using an anchoring protein and a peptide linker. This displayed protein shows high specific activity and stability and is used as a immobilized biocatalyst for producing D-tagatose through batch and semi-continuous biotransformation. The conversion rate of D-tagatose from 125 g/l D-galactose achieved 79.7% at 28 h, and the volumetric productivity reaches 4.3 g/l/h at 20 h with good reusability 5.3.1.4 L-arabinose isomerase synthesis expression of L-arabinose isomerase in fusion with the signal peptide of usp45 leads to secretion of the enzyme in induced cultures. Secretion is imrpoved by use of Lactobacillus lactis strains deficient for major proteases, ClpP and HtrA, or by use of an enhancer of protein secretion in Lactobacillus lactis fused to the recombinant isomerase gene fused to the signal peptide 5.3.1.4 L-arabinose isomerase synthesis production of ribose by immobilized recombinant Escherichia coli cells expressing the L-arabinose isomerase gene and mannose-6-phosphate isomerase mutant W17Q/N90A/L129F. The immobilized cells produce 99 g/l L-ribose from 300 g/l L-arabinose in 3 h at pH 7.5 and 60 °C in the presence of 1 mM Co2+, with a conversion yield of 33 % (w/w) and a productivity of 33 g/l/h 5.3.1.4 L-arabinose isomerase synthesis under optimal conditions, recombinant Escherichia coli cells expressing AraA can convert 150 g/l and 250 g/l D-galactose to D-tagatose with conversion rates of 32% (32 h) and 27% (48 h) 5.3.1.5 xylose isomerase synthesis used in industry for the production of high-fructose corn syrups 5.3.1.5 xylose isomerase synthesis commercial importance in the production of high-fructose corn syrup, potential application in the production of ethanol from hemicelluloses 5.3.1.5 xylose isomerase synthesis production of high fructose corn syrup 5.3.1.5 xylose isomerase synthesis production of fructose, which is used as an alternate sugar to sucrose or invert sugar in the food and beverage industries, it is also used in the baking and dairy industry 5.3.1.5 xylose isomerase synthesis higher rates of xylose utilization by further improved strains make alcoholic fermentation of hemicellulose fractions of plant biomass a realistic enterprise 5.3.1.5 xylose isomerase synthesis alcohol fermentation of xylose and mixed sugars by Saccharomyces cerevisiae constitutively overexpressing of the Orpinomyces sp. xylose isomerase, the Saccharomyces cerevisiae xylulokinase, and the Pichia stipitis SUT1 sugar transporter genes. A strain adapted for enhanced growth on xylose by serial transfer in xylose-containing minimal medium under aerobic conditions can ferment 20 g per l of xylose to ethanol with a yield of 0.37 g per g and production rate of 0.026 g per l and h. Raising the fermentation temperature from 30°C to 35°C results in a substantial increase in the ethanol yield and production as well as a significant reduction in the xylitol yield. Ethanol production from xylose and a mixture of glucose and xylose is achieved in complex medium containing yeast extract, peptone, and borate with a considerably high yield of 0.48 g per g 5.3.1.5 xylose isomerase synthesis co-overexpression of xylose isomerase and endogenous xylulokinase in a Hansenula polymorpha strain lacking NAD(P)H-dependent xylose reductase and NAD-dependent xylitol dehydrogenases activities. Recombinant strain displays improved ethanol production during the fermentation of xylose 5.3.1.5 xylose isomerase synthesis expression of Escherichia coli xylose isomerase and xylulokinase in Pseudomonas putida S12 for efficient utilization of D-xylose and L-arabinose. After laboratory evolution of strains by repeated transfer to fresh minimal medium with xylose, a strain that efficiently utilizes xylose at a considerably improved growth rate can be obtained. The high yield can be attributed in part to glucose dehydrogenase inactivity, whereas the improved growth rate may be connected to alterations in the primary metabolism. The evolved D-xylose-utilizing strain metabolizes L-arabinose as efficiently as D-xylose, while its ability to utilize glucose is not affected 5.3.1.5 xylose isomerase synthesis the combination of TxyA, XloA, and XylA is useful tool for the D-xylulose production from beta-1,3-xylan 5.3.1.6 ribose-5-phosphate isomerase synthesis production of D-allose from D-psicose by enzyme. At pH 7.5 and 50°C, synthesis of 165 g D-allose per l within 6 h 5.3.1.6 ribose-5-phosphate isomerase synthesis RpiB is a potential producer of L-form monosaccharides 5.3.1.6 ribose-5-phosphate isomerase synthesis production of allose from fructose by a one-pot reaction using Flavonifractor plautii D-allulose 3-epimerase and Clostridium thermocellum ribose 5-phosphate isomerase. Under optimal reaction conditions of pH 7.5, 60°C, 0.1 g/l epimerase, 12 g/l isomerase, and 600 g/l fructose in the presence of 1 mM Co2+, 79 g/l allose can be produced within 2 h, with a conversion yield of 13% 5.3.1.7 mannose isomerase synthesis potential use of immobilized enzyme in a process for the production of fructose syrups containing a higher proportion of fructose 5.3.1.7 mannose isomerase synthesis expression in Bacillus subtilis leads to secretion of enzyme. Under optimum conditions, production of D-mannose using crude enzyme reaches about 150 g/l with approximately 25% turnover yield 5.3.1.7 mannose isomerase synthesis optimization of fermentation medium for producing intracellular D-mannose isomerase. The optimized medium containing 1.3% w/v D-mannose, and 0.055 % soybean meal results in 1.32fold enhanced D-mannose production of 1.638 Units/ml 5.3.1.7 mannose isomerase synthesis under optimum conditions, 101.6 g/l D-mannose is produced from 400 g/l D-fructose after reaction for 9 h, giving a conversion yield of 25.4% 5.3.1.8 mannose-6-phosphate isomerase synthesis inactivation of the manA gene encoding phosphomannose isomerase results in production of amphotericins and their aglycones, 8-deoxyamphoteronolides. A double mutant lacking the phosphomannose isomerase and phosphomannomutase genes produces 8-deoxyamphoteronolides in good yields along with trace levels of glycosylated amphotericins 5.3.1.8 mannose-6-phosphate isomerase synthesis L-ribose production of the enzyme in a coupled assay system with L-arabinose isomerase, EC 5.3.1.4, in recombinant Escherichia coli ER2566, AI/MPI ratio, 1:2.5, method optimization. L-Ribose is a potential starting material for the synthesis of many L-nucleoside-based pharmaceutical compounds 5.3.1.8 mannose-6-phosphate isomerase synthesis mannose-6-phosphate isomerase from Bacillus subtilis is applied for the production of L-ribose by direct isomerization of L-ribulose 5.3.1.8 mannose-6-phosphate isomerase synthesis development of a agrobacterium-mediated Jatropha curcas transformation procedure using mannose 6-phosphate isomerase as a selectable marker. The transformation efficiency with mannose selection is twice that of kanamycin selection 5.3.1.8 mannose-6-phosphate isomerase synthesis production of ribose by immobilized recombinant Escherichia coli cells expressing the L-arabinose isomerase gene and mannose-6-phosphate isomerase mutant W17Q/N90A/L129F. The immobilized cells produce 99 g/l L-ribose from 300 g/l L-arabinose in 3 h at pH 7.5 and 60°C in the presence of 1 mM Co2+, with a conversion yield of 33% (w/w) and a productivity of 33 g/l/h 5.3.1.8 mannose-6-phosphate isomerase synthesis use as a selectable marker gene for transformation of Oryza sativa. In both Indica and Japonica subspecies, PMI expression can select and produce transgenic plants in a pattern similar to that of Escherichia coli PMI. The transgenic plants exhibit an accumulation of plant PMI transcripts and enhancement of the in vivo PMI activity. A gene of interest is successfully transformed into rice using the plant PMI as selectable marker gene 5.3.1.8 mannose-6-phosphate isomerase synthesis use of PMI gene as a selctable marker for plant transformation. A codon optimized nucleotide sequence for use in Oryza sativa leads to higher transformation frequency (54.5%) and single copy rate (44.5%) 5.3.1.14 L-rhamnose isomerase synthesis large scale production of L-mannose from L-fructose by immobilized enzyme 5.3.1.14 L-rhamnose isomerase synthesis D-allose production from D-psicose by immobilized enzyme 5.3.1.14 L-rhamnose isomerase synthesis L-xylose and L-lyxose production from xylitol using Alcaligenes 701B strain and immobilized L-rhamnose isomerase enzyme 5.3.1.14 L-rhamnose isomerase synthesis the enzyme, cross-linked with glutaraldehyde and L-lysine, is used to produce D-allose from D-piscose, which is derived from D-fructose by a recombinant D-tagatose 3-epimerase, in a bioreactor plan, method optimization, overview 5.3.1.14 L-rhamnose isomerase synthesis 40 g/l L-lyxose is produced from 100 g/l L-xylulose by the enzyme during 60 min, while 25 g/l L-mannose is produced from 100 g/l L-fructose in 80 min 5.3.1.14 L-rhamnose isomerase synthesis under optimized conditions of pH 7, 70°C, 1 mM Mn2+, 27 U enzyme/l, and 600 g D-allulose/l, 199 g D-allose/l is produced without byproducts over 2.5 h, with a conversion yield of 33% and a productivity of 79.6 g/l/h 5.3.1.15 D-lyxose ketol-isomerase synthesis about 60.0 g/l D-mannose is obtained from 400 g/l D-glucose in 8 h by coexpression of the D-glucose isomerase from Acidothermus cellulolyticus and D-lyxose isomerase from Thermosediminibacter oceani in Escherichia coli cells. The system exhibits maximum activity at pH 6.5 and 65°C with Co2+ supplement 5.3.1.15 D-lyxose ketol-isomerase synthesis bioproduction of D-tagatose from D-galactose via L-ribose synthesis performed by two step-isomerization using L-arabinose isomerase from shigella flexneri and D-lyxose/ribose isomerase. The overall 22.3% and 25% conversion rate are observed for D-tagatose and L-ribose production from D-galactose and L-arabinose, respectively 5.3.1.15 D-lyxose ketol-isomerase synthesis under optimum conditions, 101.6 g/l D-mannose is produced from 400 g/l D-fructose after reaction for 9 h, giving a conversion yield of 25.4% 5.3.1.25 L-fucose isomerase synthesis two-step enzymatic strategy for the synthesis of 6-deoxy-L-sorbose. First, the isomerization of L-fucose to L-fuculose,and the epimerization of L-fuculose to 6-deoxy-L-sorbose catalyzed by D-tagatose 3-epimerase are coupled with the targeted phosphorylation of 6-deoxy-L-sorbose by human fructose kinase in a one-pot reaction. In the second reaction step, the phosphate group of the 6-deoxy-L-sorbose 1-phosphate is hydrolyzed with acid phosphatase to produce 6-deoxy-L-sorbose in 81% yield with regard to L-fucose 5.3.1.27 6-phospho-3-hexuloisomerase synthesis production of D-[1-13C]fructose 6-phosphate from 13C-enriched formaldehyde and D-ribose 5-phosphate using 3-hexulose 6-phosphate synthase and 6-phospho-3-hexuloisomerase from Methylomonas aminofaciens and spinach ribose 5-phosphate isomerase 5.3.1.27 6-phospho-3-hexuloisomerase synthesis engineering of a strain of Corynebacterium glutamicum, which is able to produce the polyamide building block cadaverine as non-native product, for coutilization of methanol by expression of the gene encoding NAD+-dependent methanol dehydrogenase (Mdh) from Bacillus methanolicus, deletion of the endogenous aldehyde dehydrogenase genes Ald and FadH and expression of genes for hexulose-6-phosphate synthase and 6-phospho-3-hexuloisomerase of the ribulose monophosphate pathway. Growth with methanol as sole carbon source is not observed, but in vivo activity of the ribulose monophosphate pathway 5.3.1.27 6-phospho-3-hexuloisomerase synthesis engineering of Corynebacterium glutamicum toward the utilization of methanol as an auxiliary carbon source in a sugar-based medium by heterologous expression of a methanol dehydrogenase from Bacillus methanolicus, and implementation of 3-hexulose-6-phosphate synthase and 6-phospho-3-hexuloisomerasehe of Bacillus subtilis. The recombinant strain shows an average methanol consumption rate of 1.7 mM/h in a glucose-methanol medium, and the culture grows to a higher cell density than in medium without methanol 5.3.2.3 TDP-4-oxo-6-deoxy-alpha-D-glucose-3,4-oxoisomerase (dTDP-3-dehydro-6-deoxy-alpha-D-galactopyranose-forming) synthesis synthesis of TDP-D-ravidosamine, necessary for in vitro glycosylation assays. TDP-D-ravidosamine is the anticipated sugar donor substrate of RavGT (the glycosyltransferase that links D-ravidosamine to the polyketide derived backbone defuco-gilvocarcin V). Defuco-gilvocarcin V exhibits superior anticancer/antibacterial activities 5.3.3.2 isopentenyl-diphosphate DELTA-isomerase synthesis with the combination of the proper substrates and the enzymes farnesyl diphosphate synthetase and isopentenyl-diphosphate DELTA-isomerase, it is possible to use labeled derivatives of isopentenyl diphosphate and dimethylallyl diphosphate to selectively in 5.3.3.2 isopentenyl-diphosphate DELTA-isomerase synthesis expression of the genes in E. coli is a useful method for maximizing the production of industrially valuable isoprenoids 5.3.3.2 isopentenyl-diphosphate DELTA-isomerase synthesis synthesis of polyisoprenyl diphosphates in an organic-aeqous dual-liquid phase system by trans-isoprenyl diphosphate synthase mutant Y81S and isopentenyl diphosphate isomerase 5.3.4.1 protein disulfide-isomerase synthesis useful in facilitating the in vitro folding of proteins 5.3.4.1 protein disulfide-isomerase synthesis development of a versatile baculovirus expression and secretion system, using the enzyme as a fusion partner. Fusion improves the secretions and antibacterial activities of recombinant nuecin proteins and may be used for large-scale production of bioactive proteines 5.3.99.2 Prostaglandin-D synthase synthesis optimized expression protocol for recombinant enzyme in Escherichia coli and purification protocol yielding large amounts of isotopically labeled enzyme 5.3.99.7 styrene-oxide isomerase synthesis the highly stereospecific enzyme can be used in a biotransformation system producing (R)-styrene oxide 5.3.99.7 styrene-oxide isomerase synthesis enzyme disruption mutant is unable to grow on styrene or styrene oxide and accumulates enantiopure (S)-styrene when incubated with styrene. Upon growth on citrate as carbon source, mutant strain exhibits a high whole-cell activity of styrene oxide formation. In a 3 l bioreactor experiment, mutant can accumulate 150 mM (S)-styrene oxide at an enantiomerixc excess above 98% in 13 h 5.4.2.3 phosphoacetylglucosamine mutase synthesis enzymic synthesis of beta-UDP-N-acetylglucosamine using UTP:N-acetylglucosamine 1-phosphate phosphotransferase and N-acetyl-D-glucosamine 1,6-phosphomutase 5.4.3.2 lysine 2,3-aminomutase synthesis lysine 2,3-aminomutase from methanoarchaea can act as potential in vivo and in vitro biocatalyst for the synthesis of beta-lysine and related pharmacologically active compounds 5.4.3.11 phenylalanine aminomutase (D-beta-phenylalanine forming) synthesis the enzyme can be used for synthesis of beta-arylalanines with different groups at the benzene ring, 93% yield of 2-nitro-beta-phenylalanine. Potential prospectin application for the enzyme 5.4.3.11 phenylalanine aminomutase (D-beta-phenylalanine forming) synthesis the enzyme is an attractive alternative enzymatic route to obtain enantiomerically pure alpha- and beta-amino acids, via green synthetic routes to chiral amines 5.4.3.11 phenylalanine aminomutase (D-beta-phenylalanine forming) synthesis utility of the methylidene imidazolone-dependent Pantoea agglomerans phenylalanine aminomutase (PaPAM) for making non-natural beta-amino acids, substrate specificity of PaPAM with several commercially available (S)-arylalanine substrates, overview 5.4.4.1 (hydroxyamino)benzene mutase synthesis expression of enzyme plus nitrobenzene nitroreductase in Escherichia coli. Rapid and stoichiometric conversion of nitrobenzene to 2-aminophenol, of 2-nitroacetophenone to 2-amino-3-hydroxyacetophenone, and of 3-nitroacetophenone to 3-amino-2-hydroxyacetophenone, as well as further conversions. Final yields of aminophenols after extraction and recovery are over 64% 5.4.99.1 methylaspartate mutase synthesis induction strategy to enhance the level of protein expression in bacteriophage T7 expression system in Escherichia coli. Yield of purified glutamate mutase S component MutS protein is increased threefold by reducing the induction temperature to 20°C and using 0.2% lactose and 50 mg per l IPTG simultaneously as inducers 5.4.99.5 chorismate mutase synthesis synthesis of L-phenylalanine (an important amino acid that is widely used in the production of food flavors and pharmaceuticals) by engineered Escherichia coli. Coexpression of Vitreoscilla hemoglobin gene, driven by a tac promoter, with the genes encoding 3-deoxy-D-arabinoheptulosonate-7-phosphate synthetase (aroF) and feedback-resistant chorismate mutase/prephenate dehydratase (pheAfbr), leads to increased productivity of L-phenylalanine and decreased demand for aeration by Escherichia coli CICC10245 5.4.99.7 Lanosterol synthase synthesis engineering ERG7 for producing biological active agents is promising 5.4.99.9 UDP-galactopyranose mutase synthesis production of uridine 5'-(trihydrogen diphosphate) P'-alpha-D-galactofuranosyl ester, a precursor required for the formation of the lipopolysaccharide O-antigen of Klebsiella pneumoniae serotype O1 5.4.99.11 isomaltulose synthase synthesis formation of isomaltulose by immobilized Erwinia rhapontici cells 5.4.99.11 isomaltulose synthase synthesis production of palatinose, which is used as a sweetener and as a substitute for sucrose 5.4.99.11 isomaltulose synthase synthesis strain produces isomaltulose with maximum yield of 78-89% of supplied sucrose and 4% contaminating trehalose 5.4.99.11 isomaltulose synthase synthesis strain FMB1 show more than 90% conversion of sucrose at 4 g per l into isomaltulose in 2 days, with small amounts of trehalulose, glucose, and fructose as by-products. Use of strain in industrial production of isomaltulose 5.4.99.11 isomaltulose synthase synthesis immobilized cells of Protaminobacter rubrum strain CBS 574.77 are commonly used for the synthesis of isomaltulose from sucrose on an industrial scale 5.4.99.11 isomaltulose synthase synthesis isomaltulose production by the enzyme roduced from immobilized of Escherichia coli cells recombinantly expressing the enzyme, method overview 5.4.99.11 isomaltulose synthase synthesis isomaltulose (alpha-D-glucopyranosyl-1,6-D fructofuranose) is a carbohydrate used as ideal sucrose substitute. Sucrose isomerase is widely used in industries for the production of isomaltulose. The enzyme catalyzes the isomerization of sucrose into isomaltulose and trehalulose and may hydrolyze sucrose to produce small amounts of glucose and fructose 5.4.99.13 isobutyryl-CoA mutase synthesis IcmF is potentially useful as a reagent for bioremediation of pivalic acid found in pharmaceutical wastewaters (pivalic acid esters are used as pro-drugs) and/or for the biosynthesis of a simple beta-nonfunctional alpha-quaternary carboxylic acid. Another potential application of IcmF is in metabolic engineering of pathways to produce branched C4 and C5 building blocks that can subsequently be converted to useful derivatives, for example, the corresponding isobutyl and pivalyl alcohols 5.4.99.13 isobutyryl-CoA mutase synthesis the production of isobutanol, a branched-chain alcohol that can be used as a gasoline substitute, using a CoA-dependent pathway in recombinant Ralstonia eutropha strain H16. The designed pathway involves isobutyryl-CoA mutase. The engineered strain produces about 30 mg/l isobutanol from fructose. The carbon skeleton rearrangement chemistry demonstrated may be used to expand the range of the chemicals accessible with CoA-dependent pathways 5.4.99.15 (1->4)-alpha-D-glucan 1-alpha-D-glucosylmutase synthesis overexpression of the enzyme treY gene and the treY/treZ synthetic operon encoding enzyme and maltooligosyltrehalose trehalohydrolase significantly increases maltooligosyltrehalose synthase activity, the rate-limiting step, and improves the specific productivity and the final titer of trehalose. Furthermore, a strong decrease is noted in glycogen accumulation. Expression of UDP-glucose pyrophosphorylase/maltooligosyltrehalose synthase and UDP-glucose pyrophosphorylase/maltooligosyltrehalose synthase/maltooligosyltrehalose trehalohydrolase synthetic operons shows a partial recovery in the intracellular glycogen levels and a significant improvement in both intra- and extracellular trehalose content 5.4.99.15 (1->4)-alpha-D-glucan 1-alpha-D-glucosylmutase synthesis production of trehalose by enzyme plus maltooligosyltrehalose trehalohydrolase is increased in presence of 4-alpha-glucanotransferase, converting 70% of maltopentaose into trehalose after 6 h, while 60% is converted without 4-alpha-glucanotransferase 5.4.99.15 (1->4)-alpha-D-glucan 1-alpha-D-glucosylmutase synthesis conditions for the production of trehalose from starch by thermostable maltooligosyl trehalose synthase and maltooligosyl trehalose trehalohydrolase from Sulfolubus acidocaldarius ATCC 33909 5.4.99.15 (1->4)-alpha-D-glucan 1-alpha-D-glucosylmutase synthesis trehalose production from starch 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis immobilization of recombinant enzyme on Eupergit C250L for the production of trehalose. Immobilization does not affect optimum pH value, optimum temperature is shifted from 45°C to 65°C. Immobilized enzyme is stable at 70°C for 16 days and can be used more than 10times in batch reaction. A maximum yield of 42% trehalose can be reached from 50 g/l maltose 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis the enzyme is a good candidate in the large-scale production of trehalose from maltose 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis the enzyme is useful in the biotransformation process for inexpensive production of trehalose from maltose 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis trehalose synthase from Pyrococcus horikoshii can be applied to the sugar nucleotide cycling process for the synthesis of functional alpha-galactosyl oligosaccharides, alpha-galactose epitopes and globotriose, using the effective regeneration of UDP-Gal, method development and evaluation, overview 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis good application potential of the recombinant enzyme in the efficient conversion of trehalose from maltose 5.4.99.16 maltose alpha-D-glucosyltransferase synthesis the enzyme (TreS) produced from Acidiplasma spMBA-1 can catalyze a considerable amount of maltose into trehalose in a single step reaction. Maltose is a relatively cheap substrate. Hence, TreS can be used as an alternative commercial enzyme to produce trehalose commercially. A significant amount of glucose is being produced as a byproduct which hinders the production of commercial trehalose. If it is possible to suppress glucose production by genetic modifications, it could enhance trehalose production 5.4.99.17 squalene-hopene cyclase synthesis production of unnatural polyprenoids and supranatural steroids by manipulation of the enzyme reaction by combination of substrate analogues 5.4.99.17 squalene-hopene cyclase synthesis cyclization of homofarnesol to ambroxan as well as the conversion of citronellal to 2-isopropenyl-5-methyl-cyclohexanol bythe isozyme SHC1 can be economically attractive, as both products are used in the flavour and fragrance industry 5.4.99.17 squalene-hopene cyclase synthesis it is possible to use the cyanobacterium Synechocystis to generate squalene, a hydrocarbon of commercial interest and a potential biofuel 5.4.99.39 beta-amyrin synthase synthesis engineering of the sterol pathway in Saccharomyces cerevisiae for production of beta-amyrin. Expression of enzyme gene plus manipulation of 3-hydroxy-3-methylglutaryl-CoA reductase and lanosterol synthase in Saccharomyces cerevisiae leads to levels of 6 mg beta-amyrin per l of culture 5.4.99.39 beta-amyrin synthase synthesis use of enzyme for production of unnatural tetracyclic sesterterpenes 5.4.99.64 2-hydroxyisobutanoyl-CoA mutase synthesis introduction of a 2-hydroxyisobutyric acid synthesis route in Escherichia coli by concomitantly expressing the RCM genes and phaA and phaB, encoding beta-ketothiolase and NADP-dependent (R)-3-hydroxybutyryl-CoA dehydrogenase. A concentration of 3-hydroxybutyric acid of up to 17.7 mM can be obtained within 8 days of feeding the recombinant strain with gluconic acid as the main carbon source 5.4.99.64 2-hydroxyisobutanoyl-CoA mutase synthesis synthesis of 2-hydroxyisobutanoate in Cupriavidus necator H16 expressing mutase genes HcmA and HcmB. Due to the enantiospecificity of the mutase, fructose is a weaker substrate for 2-hydroxyisobutanoate synthesis than butanoate. Production rates achieved with the poly-3-hydroxybutanoate-negative strain H16 PHB?4 on butanoate are higher than on fructose. Using the wild-type does not significantly improve the production rates, the latter shows a 34fold and a 5fold lower 2-hydroxyisobutanoate synthesis rate compared to H16 PHB?4 on fructose and butanoate, respectively. Both strains show concomitant excretion of undesired side products, such as pyruvate and 3-hydroxybutanoate 5.5.1.4 inositol-3-phosphate synthase synthesis production of myo-inositol from glucose by a novel trienzymatic cascade of polyphosphate glucokinase, inositol 1-phosphate synthase and inositol monophosphatase. myo-Inositol (inositol) is important in the cosmetics, pharmaceutical and functional food industries. The conversion ratio from glucose to inositol reaches 90%, which is promising for the enzymatic synthesis of inositol without ATP supplementation 5.5.1.6 chalcone isomerase synthesis coexpression of enzyme with a 4-coumarate:CoA ligase and chalcone synthase in Escherichia coli for synthesis of unnatural flavonoids and stilbenes. A variety of unnatural carboxylic acids such as fluorocinnamic acid, furyl acrylic acid, thienyl acylic acid, naphthyl acrylic acid are used as substrates to give the corresponding unnatural flavanones 5.5.1.6 chalcone isomerase synthesis combinatorial expression of 4-coumaroyl:Co A ligase, chalcone synthase, chalcone isomerase, and flavanone 3beta-hydrolase in Saccharomyces cerevisiae for production of unnatural flavanones when fed with aromatic acrylic acids 5.5.1.6 chalcone isomerase synthesis transgenic tobacco overexpressing Saussurea medusa enzyme SmCHI produces up to fivefold more total flavonoids than wild-type, mainly due to accumulation of rutin. Transgenic tobacco treated with antisense SmCHI accumulates smaller amounts of flavonoids 5.5.1.12 copalyl diphosphate synthase synthesis the development of yeast strains carrying the engineered Erg20p, which support efficient isoprenoid production, e.g. by abietadiene synthase, and can be used as a dedicated chassis for diterpene production or biosynthetic pathway elucidation. The design developed can be applied to the production of any GGPP-derived isoprenoid and is compatible with other yeast terpene production platforms, method overview 6.1.1.2 tryptophan-tRNA ligase synthesis a functional, orthogonal suppressor tRNA pair can prove useful for the incorporation of bulky, unnatural amino acids into the genetic code. Role of tRNA flexibility in molecular recognition and the engineering and evolution of tRNA specificity 6.1.1.6 lysine-tRNA ligase synthesis recombinant Escherichia coli lysyl-tRNA synthase (LysU) has been previously utilised in the production of stabile, synthetic diadenosine polyphosphate (ApnA) analogues. LysU is also useful as a tool for highly controlled phosphate-phosphate bond formation between nucleotides, avoiding the need for complex protecting group chemistries. Resulting high yielding tandem LysU-based biosynthetic-synthetic/synthetic-biosynthetic strategies emerge for the preparation of varieties of ApnA analogues directly from inexpensive natural nucleotides and nucleosides. Analogues so formed make a useful small library with which to probe ApnA activities in vitro and in vivo leading to the discovery of potentially potent biopharmaceuticals active against chronic pain and other chronic, high-burden disease states 6.1.1.26 pyrrolysine-tRNAPyl ligase synthesis synthesis and genetic incorporation of aliphatic azides and alkynes into proteins using the natural pyrrolysyl tRNA synthetase/tRNACUA pair and the efficient bio-orthogonal labeling of these amino acids using [3+2] cycloaddition, i.e. click chemistry. Escherichia coli transformed with pBKPylS10 encoding pyrrolysyl tRNA synthetase and pMyo4TAGPylT-his610 encoding tRNACUA and a C-terminally hexahistidine tagged myoglobin gene with an amber codon at position 4 incorporates (2S)-2-amino-6-[[(prop-2-yn-1-yloxy)carbonyl]amino]hexanoic acid. The yield of protein containing (2S)-2-amino-6-[[(prop-2-yn-1-yloxy)carbonyl]amino]hexanoic acid is not improved by efforts to evolve the enzyme but is increased 5fold by increasing the concentration of (2S)-2-amino-6-[[(prop-2-yn-1-yloxy)carbonyl]amino]hexanoic acid 7.5fold 6.1.2.1 D-alanine-(R)-lactate ligase synthesis structure-based modification of D-alanine-D-alanine ligase from strain ATCC 43589 for D-alanyl-D-lactate and other depsipeptide synthesis by mutant S137G/Y207F 6.2.1.11 biotin-CoA ligase synthesis production of alpha-dehydrobiotin, an antibiotic, from biotinyl-CoA with biotinyl-CoA synthetase and acyl-CoA oxidase 6.2.1.12 4-coumarate-CoA ligase synthesis crude aspen CoA ligase can be effectively and economically used to synthesize caffeoyl CoA 6.2.1.12 4-coumarate-CoA ligase synthesis viability of a flavonoid network to utilize acrylic acid analogues and describe the combinatorial mutasynthesis of novel unnatural flavonoids using recombinant Saccharomyces cerevisiae, overview 6.2.1.12 4-coumarate-CoA ligase synthesis heterologous expression of biosynthetic genes encoding 4-coumarate:coenzyme A ligase and tyramine N-hydroxycinnamoyltransferase cloned from Arabidopsis thaliana and pepper, respectively. Simultaneous supplementation of substrates to the recombinant Escherichia coli results in the secretion of multiple tyramine derivatives into the medium at high yield: 4-coumaroyltyramine at 189 mg/l, feruloyltyramine at 135 mg/l, caffeoyltyramine at 40 mg/l. The recombinant Escherichia coli also produces, albeit at low concentration, a range of dopamine derivatives such as feruloyldopamine 6.2.1.12 4-coumarate-CoA ligase synthesis a de novo pathway for the production of raspberry ketone is assembled using four heterologous genes, encoding phenylalanine/tyrosine ammonia lyase, cinnamate-4-hydroxlase, coumarate-CoA ligase and benzalacetone synthase, in an industrial strain of Saccharomyces cerevisiae. It is possible to produce sensorially-relevant quantities of raspberry ketone in the industrial heterologous host 6.2.1.12 4-coumarate-CoA ligase synthesis a heterologous pathway to produce raspberry ketone from p-coumaric acid, including 4-coumarate:CoA ligase (4CL), benzalacetone synthase (BAS), and raspberry ketone/zingerone synthase (RZS1) from plants, is assembled in Escherichia coli. Raspberry ketone is an important ingredient in the flavor and fragrance industries. Engineered strain CZ-8 which cooverexpresses At4CL1, RpBAS, and RiRZS1 achieves levels of 90.97 mg/l of raspberry ketone 6.2.1.14 6-carboxyhexanoate-CoA ligase synthesis production of biotin 6.2.1.17 propionate-CoA ligase synthesis used for incorporation of unusual acyl groups into biopolymers and polyketide antibiotics 6.2.1.20 long-chain-fatty-acid-[acyl-carrier-protein] ligase synthesis the enzyme is a useful tool for the synthesis of native acyl-[acyl-carrier-protein] for use in examining other lipid biosynthetic enzymes in E. coli 6.2.1.20 long-chain-fatty-acid-[acyl-carrier-protein] ligase synthesis attractive alternative to current chemical and enzymatic methods of acyl-[acyl-carrier protein] preparation and analysis 6.2.1.30 phenylacetate-CoA ligase synthesis the expression of the heterologous enzyme phenylacetyl-CoA ligase, involved in catabolism of a penicillin precursor, is a useful strategy for improving the biosynthetic machinery of this fungus 6.2.1.30 phenylacetate-CoA ligase synthesis engineering of Hansenula polymorpha for secretion of penicillin. Introduction of the Penicilliumchrysogenum gene encoding the non-ribosomal peptide synthetase delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase in Hansenula polymorpha, results in the production of active enzyme, when co-expressed with the Bacillus subtilis sfp gene encoding a phosphopantetheinyl transferase that activates ACVS. Co-expression with the Penicillium chrysogenum genes encoding the cytosolic enzyme isopenicillin N synthase as well as the two peroxisomal enzymes isopenicillin N acyl transferase and phenylacetyl CoA ligase results in production of biologically active penicillin, which is efficiently secreted. The amount of secreted penicillin is approximately 1 mg/l. Penicillin production is decreased over 2fold in a yeast strain lacking peroxisomes 6.2.1.32 anthranilate-CoA ligase synthesis three anthranilate derivatives, N-methylanthranilate, methyl anthranilate, and methyl N-methylanthranilate are synthesized using metabolically engineered stains of Escherichia coli. NMT encoding N-methyltransferase from Ruta graveolens, AMAT encoding anthraniloyl-coenzyme A (CoA):methanol acyltransferase from Vitis labrusca, and pqsA encoding anthranilate coenzyme A ligase from Pseudomonas aeruginosa are cloned and Eschetrichia coli strains harboring these genes were used to synthesize the three desired compounds. Escherichia coli mutants (metJ, trpD, tyrR mutants), which provide more anthranilate and/or S-adenosyl methionine, are used to increase the production of the synthesized compounds. 0.1853 mM N-methylanthranilate and 0.0952 mM methyl N-methylanthranilate are synthesized 6.2.1.45 E1 ubiquitin-activating enzyme synthesis expression of the full length of human UBE1 in Escherichia coli and purification by Ni-NTA superflow sepharose and strep-tactin sepharose which is based on UB-UBE1 high-energy thioester bonded intermediate complex. Purified UBE1 can activate and conjugate UB to ubiquitin-conjugating enzyme E2s 6.2.1.45 E1 ubiquitin-activating enzyme synthesis 5–10 mg of histidine-tagged mouse ubiquitin-activating enzyme E1 can be easily obtained from a 1 l Escherichia coli culture. A low temperature during the protein induction step is critical to obtain an active enzyme 6.2.1.60 marinolic acid-CoA ligase synthesis the pair acetyltransferase HolE/TmlU can readily ligate pseudomonic acid C to a variety of primary amines including 3-aminocoumarins, but is less effective with a series of aryl amines. An adjacent substrate carbonyl appears useful or important for recognition of the amine donor 6.2.1.74 3-amino-5-hydroxybenzoate-[acyl-carrier protein] ligase synthesis preparation of several potential substrates believed to be early biosynthetic intermediates to FR900482 and the mitomycins to reveal the biochemical production of mitomycin C and FR900482 6.3.1.2 glutamine synthetase synthesis isolated glutamine synthetase forms theanine in the mixture with the yeast fermentation system, production in high concentrations can be achieved 6.3.1.2 glutamine synthetase synthesis the organism is used for industrial production of L-glutamine 6.3.1.2 glutamine synthetase synthesis high efficiency L-Gln production by coupling genetically engineered Bacillus subtilis glutamine synthetase with yeast alcoholic fermentation system using small ubiquitin-related modifier fusion technology SUMO and 0.1% lactose as inducer. Fusion protein is expressed in totally soluble form in Escherichia coli and can be purified to 90% purity by nickel nitrilo-triacetic acid resin chromatography with a yield of 625 mg per liter fermentation culture. About 121 mg recombinant GS is obtained from 1 l fermentation culture with 96% purity and 23 U/mg activity 6.3.1.2 glutamine synthetase synthesis rapid and scalable two-step protocol for expression and purification of glutamine synthetase in an auxotrophic Escherichia coli strain utilizing differential precipitation by divalent cations followed by affinity chromatography to produce suitable quantities of homogenous material for structural characterization 6.3.1.9 trypanothione synthase synthesis production of trypanothione and trypanothione disulfide in >200 mg quantities by mutant C59A lacking the amidase activity. The protocol also allows the synthesis of related glutathione conjugates 6.3.1.20 lipoate-protein ligase synthesis enzyme-mediated two-step labeling protocol suitable for live-cell labeling using lipoic acid ligase with norbornene substrates and subsequent inverse-electron demand Diels-Alder reaction 6.3.1.20 lipoate-protein ligase synthesis the enzyme can be used for a two-step labeling strategy that combines high selectivity of enzyme-mediated labeling with the chemoselectivity of palladium-catalyzed Sonogashira cross-coupling, overview 6.3.2.2 glutamate-cysteine ligase synthesis resting cells of Escherichia coli expressing gamma-glutamylcysteine synthetase, with ATP regeneration through glycolysis, synthesize 12.1 mM theanine in 18 h from 429 mM ethylamine 6.3.2.2 glutamate-cysteine ligase synthesis at elevated concentrations of the precursor amino acids and ATP, Escherichia coli JM109 pTrc99A-gshF produces 36 mM GSH with a molar yield of 0.9 mol/mol based on added cysteine and of 0.45 mol/mol based on added ATP. When ATP is replaced with glucose, the strain produces 7 mM in 3 h. In the presence of glucose and the pmr1 mutant of Saccharomyces cerevisiae BY4742 for ATP generation, Escherichia coli JM109 pTrc99A-gshF produces 33.9 mM GSH in 12 h with a yield of 0.85 mol/mol based on added l-cysteine 6.3.2.2 glutamate-cysteine ligase synthesis Rhodosporidium diobovatum with its GSH1 and GSH2 genes can be useful for industrial GSH production 6.3.2.3 glutathione synthase synthesis at elevated concentrations of the precursor amino acids and ATP, Escherichia coli JM109 pTrc99A-gshF produces 36 mM GSH with a molar yield of 0.9 mol/mol based on added cysteine and of 0.45 mol/mol based on added ATP. When ATP is replaced with glucose, the strain produces 7 mM in 3 h. In the presence of glucose and the pmr1 mutant of Saccharomyces cerevisiae BY4742 for ATP generation, Escherichia coli JM109 pTrc99A-gshF produces 33.9 mM GSH in 12 h with a yield of 0.85 mol/mol based on added l-cysteine 6.3.2.3 glutathione synthase synthesis overexpression of the first gene of GSH biosynthesis, gamma-glutamylcysteine synthetase, or of the central regulatory gene of sulfur metabolism, leads to an elevated pool of intracellular GSH, and the strains additionally accumulate almost twice as much ethanol as the wild-type strain during glucose fermentation. Transformants with a high GSH pool prove more sensitive to exogenous ethanol than the corresponding wild-type strains 6.3.2.3 glutathione synthase synthesis Rhodosporidium diobovatum with its GSH1 and GSH2 genes can be useful for industrial GSH production 6.3.2.4 D-Alanine-D-alanine ligase synthesis D-alanine-D-alanine ligase is a biocatalyst for synthesizing D-amino acid dipeptides, use of a thermostable ATP regeneration system 6.3.2.25 tubulin-tyrosine ligase synthesis restoration of the excitability of the giant axon of Doryteuthis bleekeri by tubulin-tyrosine ligase and microtubule proteins 6.3.2.29 cyanophycin synthase (L-aspartate-adding) synthesis expression of cyanophycin synthase in wild-type Sinorhizobium meliloti 1021 and in a phbC-negative mutant. Yeast mannitol broth yields the highest cyanophycin contents in both Sinorhizobium meliloti 1021 strains. Supplying the medium with isopropyl-beta-D-thiogalactopyranoside, aspartic acid, and arginine enhances cyanophycin contents about 2.5- and 2.8fold. Varying the nitrogen-to-carbon ratio in the medium enhanced the cyanophycin content further to 43.8% w/w of cell dry weight. Cyanophycin from the Sinorhizobium meliloti strains consists of equimolar amounts of aspartic acid and arginine and contains no other amino acids even if the medium is supplemented with glutamic acid, citrulline, ornithine, or lysine. Cyanophycin isolated from Sinorhizobium meliloti exhibits average molecular weights between 20 and 25 kDa. Cyanophycin isolated after expression in Escherichia coli S17-1 exhibits average molecular weight between 22 and 30 kDa. Co-expression of cyanophycinase from Anabaena sp. PCC7120 encoded by cphB17120 in cphA17120-positive Escherichia coli S17-1, Sinorhizobium meliloti 1021, and its phbC-negative mutant gives cyanophycinase activities in crude extracts, and no CGP granules occur 6.3.2.29 cyanophycin synthase (L-aspartate-adding) synthesis expression of the cyanophycin synthetase of Synechocystis sp. PCC 6308 in Pseudomonas putida ATCC 4359 using an optimised medium for cultivation, results in synthesis of insoluble cyanophycin up to 14.7 w/w and soluble cyanophycin amounting up to 28.7 w/w of the cell dry matter. The soluble CGP is composed of 50.4 mol% aspartic acid, 32.7 mol% arginine, 8.7 mol% citrulline and 8.3 mol% lysine. The insoluble cyanophycin contains less than 1 mol% of citrulline. Using a mineral salt medium with 1.25 or 2% w/v sodium succinate, respectively, plus 23.7 mM L-arginine, the cells synthesise insoluble cyanophycin amounting up to 25% to 29% of the CDM with only a very low citrulline content 6.3.2.29 cyanophycin synthase (L-aspartate-adding) synthesis restriction of cyanophycin accumulation to the potato tubers by using the cyanophycin synthetase gene from Thermosynechococcus elongatus BP-1, under the control of the tuber-specific class 1 promoter. Tuber-specific cytosolic expression by pB33-cphATe as well as tuber-specific plastidic expression by pB33-PsbYcphATe results in significant polymer accumulation solely in the tubers. In plants transformed with pB33-cphATe, both cyanophycin synthetase and cyanophycin are detected in the cytoplasm leading to an increase up to 2.3% cyanophycin of dry weight and resulting in small and deformed tubers. In B33-PsbY-cphATe tubers, cyanophycin synthetase and cyanophycin are exclusively found in amyloplasts leading to a cyanophycin accumulation up to 7.5% of dry weight. These tubers are normal in size, some clones show reduced tuber yield and sometimes exhibit brown sunken staining starting at tubers navel. During a storage period over of 32 weeks of one selected clone, the cyanophycin content was stable in B33-PsbYcphATe tubers but the stress symptoms increased. Nitrogen fertilization in the greenhouse does not lead not to an increased cyanophycin yield, slightly reduced protein content, decreased starch content, and changes in the amounts of bound and free arginine and aspartate 6.3.2.29 cyanophycin synthase (L-aspartate-adding) synthesis synthesis of cyanophycin using an anabolism-based media-dependent plasmid addiction system to enhance plasmid stability, and a process based on a modified mineral salts medium yielding a cyanophycin content of 42% w/w at the maximum without the addition of amino acids to the medium. This plasmid addiction system is based on different lysine biosynthesis pathways and consists of a knockout of the chromosomal dapE that disrupts the native succinylase pathway in Escherichia coli and the complementation by the plasmid-encoded artificial aminotransferase pathway mediated by the dapL gene from Synechocystis sp. PCC 6308, which allows the synthesis of the essential lysine precursor L,L-2,6-diaminopimelate. This plasmid also harbors an engineered cyanophycin synthetase gene responsible for cyanophycin production. Cultivation experiments reveal an at least 4.5fold enhanced production of cyanophycin in comparison to control cultivations 6.3.2.29 cyanophycin synthase (L-aspartate-adding) synthesis cyanophycin produced in Escherichia coli is composed of 50% of aspartic acid, 45% of arginine, and 3.5% of lysine, and exhibits a homogenous molecular mass of 35 kDa. Cultivation in presence of arginine, aspartic acid, lysine and glucose with the minimal resource leads to 1.72 g/l soluble cyanophycin 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis establishment of cyanophycin, i.e. multi-L-arginylpoly-L-aspartic acid or CGP, synthesis and applicability of this industrially widely used microorganism for the production of this polyamide, overview 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis production of biodegradable polymers that can be used to substitute petrochemical compounds in commercial products by the recombinant enzyme in transgenic plants 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis expression of cyanophycin synthase in wild-type Sinorhizobium meliloti 1021 and in a phbC-negative mutant. Yeast mannitol broth yields the highest cyanophycin contents in both Sinorhizobium meliloti 1021 strains. Supplying the medium with isopropyl-beta-D-thiogalactopyranoside, aspartic acid, and arginine enhances cyanophycin contents about 2.5- and 2.8fold. Varying the nitrogen-to-carbon ratio in the medium enhanced the cyanophycin content further to 43.8% w/w of cell dry weight. Cyanophycin from the Sinorhizobium meliloti strains consists of equimolar amounts of aspartic acid and arginine and contains no other amino acids even if the medium is supplemented with glutamic acid, citrulline, ornithine, or lysine. Cyanophycin isolated from Sinorhizobium meliloti exhibits average molecular weights between 20 and 25 kDa. Cyanophycin isolated after expression in Escherichia coli S17-1 exhibits average molecular weight between 22 and 30 kDa. Co-expression of cyanophycinase from Anabaena sp. PCC7120 encoded by cphB17120 in cphA17120-positive Escherichia coli S17-1, Sinorhizobium meliloti 1021, and its phbC-negative mutant gives cyanophycinase activities in crude extracts, and no CGP granules occur 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis expression of the cyanophycin synthetase of Synechocystis sp. PCC 6308 in Pseudomonas putida ATCC 4359 using an optimised medium for cultivation, results in synthesis of insoluble cyanophycin up to 14.7 w/w and soluble cyanophycin amounting up to 28.7 w/w of the cell dry matter. The soluble CGP is composed of 50.4 mol% aspartic acid, 32.7 mol% arginine, 8.7 mol% citrulline and 8.3 mol% lysine. The insoluble cyanophycin contains less than 1 mol% of citrulline. Using a mineral salt medium with 1.25 or 2% w/v sodium succinate, respectively, plus 23.7 mM L-arginine, the cells synthesise insoluble cyanophycin amounting up to 25% to 29% of the CDM with only a very low citrulline content 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis restriction of cyanophycin accumulation to the potato tubers by using the cyanophycin synthetase gene from Thermosynechococcus elongatus BP-1, under the control of the tuber-specific class 1 promoter. Tuber-specific cytosolic expression by pB33-cphATe as well as tuber-specific plastidic expression by pB33-PsbYcphATe results in significant polymer accumulation solely in the tubers. In plants transformed with pB33-cphATe, both cyanophycin synthetase and cyanophycin are detected in the cytoplasm leading to an increase up to 2.3% cyanophycin of dry weight and resulting in small and deformed tubers. In B33-PsbY-cphATe tubers, cyanophycin synthetase and cyanophycin are exclusively found in amyloplasts leading to a cyanophycin accumulation up to 7.5% of dry weight. These tubers are normal in size, some clones show reduced tuber yield and sometimes exhibit brown sunken staining starting at tubers navel. During a storage period over of 32 weeks of one selected clone, the cyanophycin content was stable in B33-PsbYcphATe tubers but the stress symptoms increased. Nitrogen fertilization in the greenhouse does not lead not to an increased cyanophycin yield, slightly reduced protein content, decreased starch content, and changes in the amounts of bound and free arginine and aspartate 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis synthesis of cyanophycin using an anabolism-based media-dependent plasmid addiction system to enhance plasmid stability, and a process based on a modified mineral salts medium yielding a cyanophycin content of 42% w/w at the maximum without the addition of amino acids to the medium. This plasmid addiction system is based on different lysine biosynthesis pathways and consists of a knockout of the chromosomal dapE that disrupts the native succinylase pathway in Escherichia coli and the complementation by the plasmid-encoded artificial aminotransferase pathway mediated by the dapL gene from Synechocystis sp. PCC 6308, which allows the synthesis of the essential lysine precursor L,L-2,6-diaminopimelate. This plasmid also harbors an engineered cyanophycin synthetase gene responsible for cyanophycin production.Cultivation experiments reveal an at least 4.5fold enhanced production of cyanophycin in comparison to control cultivations 6.3.2.30 cyanophycin synthase (L-arginine-adding) synthesis cyanophycin produced in Escherichia coli is composed of 50% of aspartic acid, 45% of arginine, and 3.5% of lysine, and exhibits a homogenous molecular mass of 35 kDa. Cultivation in presence of arginine, aspartic acid, lysine and glucose with the minimal resource leads to 1.72 g/l soluble cyanophycin 6.3.2.49 L-alanine-L-anticapsin ligase synthesis fermentative production of L-alanyl-L-glutamine by a metabolically engineered Escherichia coli strain expressing L-amino acid alpha-ligase 6.3.2.49 L-alanine-L-anticapsin ligase synthesis Lal is a promising enzyme to achieve cost-effective synthesis 6.3.2.49 L-alanine-L-anticapsin ligase synthesis enzyme Trp332 mutants can alter the substrate specificity and activity depending on the size and shape of substituted amino acids, scope for the rational design of the enzyme to produce desirable dipeptides, the positioning of the conserved Arg residue in L-aminoa cid ligase is important for enantioselective recognition of L-amino acids 6.3.2.54 L-2,3-diaminopropanoate-citrate ligase synthesis the SbnCEF synthetases and decarboxylase SbnH are necessary and sufficient to produce staphyloferrin B in vitro in reactions containing component substrates L-2,3-diaminopropionic acid, citric acid and 2-oxoglutaric acid 6.3.2.56 staphyloferrin B synthase synthesis the SbnCEF synthetases and decarboxylase SbnH are necessary and sufficient to produce staphyloferrin B in vitro in reactions containing component substrates L-2,3-diaminopropionic acid, citric acid and 2-oxoglutaric acid 6.3.3.4 (carboxyethyl)arginine beta-lactam-synthase synthesis clavulanic acid intermediates: deoxygaunidinoproclavaminic acid, guanidinoproclavaminic acid, and dihydroclavaminic acid are heterologously produced in Streptomyces venezuelae recombinant using four sets of early genes from the clavulanic acid biosynthetic pathway 6.3.3.5 O-ureido-D-serine cyclo-ligase synthesis expression of four D-cycloserine biosynthetic genes (dcsC, dcsD, dcsE, and dcsG) in tandem in Escherichia coli. In presence of L-serine and hydroxyurea, D-cycloserine is produced. Coexpression of gene DcsJ and optimization of conditions results in production of 0.98 mM D-cycloserine 6.3.3.6 carbapenam-3-carboxylate synthase synthesis stereoselctive synthesis of dimethylsubstituted carbapenams using engineered 5-carboxymethylproline synthase and carbapenam-3-carboxylate synthase 6.3.4.4 adenylosuccinate synthase synthesis expression of mutant P242N in an inosine-producing strain results in an approximately 4.66fold increase in inosine production, to 0.41 g/L, in minimal medium without hypoxanthine accumulation 6.3.4.10 biotin-[propionyl-CoA-carboxylase (ATP-hydrolysing)] ligase synthesis site-directed antibody immobilization by fusing biotin carboxyl carrier protein BCCP to the IgG-binding domain of Protein A, and the resulting fusion protein is immobilized on the biotin protein ligase-modified gold surface of the sensor chip for quartz crystal microbalance through complexation between BCCP and biotin protein ligase. The layer of the IgG-binding domain is successfully captured the antibody, and the antibody retains high antigen-binding capability 6.3.4.12 glutamate-methylamine ligase synthesis the enzyme is usable in theanine formation by coupling with the alcoholic fermentation system of bakers yeast, Saccharomyces cerevisiae 6.3.4.12 glutamate-methylamine ligase synthesis the enzyme is enclosed with dried Saccharomyces cerevisiae cells in a dialysis membrane tube to produce theanine from glutamic acid and ethylamine to 100% conversion rate. Six repeats of the reaction are possible in presence of NAD+, overview 6.3.4.12 glutamate-methylamine ligase synthesis efficient synthesis of gamma-glutamyl compounds by co-expression of gamma-glutamylmethylamide synthetase and polyphosphate kinase in engineered Escherichia coli 6.3.4.15 biotin-[biotin carboxyl-carrier protein] ligase synthesis the enzyme is utilized to synthesize site-specific biotinylated proteins via a biotin acceptor peptide tag 6.3.4.15 biotin-[biotin carboxyl-carrier protein] ligase synthesis in vivo biotinylation of bacterial magnetic particles synthesized by Magnetospirillum magneticum AMB-1 by heterologous expression of Escherichia coli biotin ligase. To biotinylate bacterial magnetic particles in vivo, biotin acceptor peptide is fused to a bacterial magnetic particles surface protein, Mms13, and Escherichia coli biotin ligase is simultaneously expressed in the truncated form lacking the DNA-binding domain. The biotinylated biotin acceptor protein-displaying bacterial magnetic particles are then exposed to streptavidin by simple mixing. The streptavidin-binding capacity of bacterial magnetic particles biotinylated in vivo is 35fold greater than that of bacterial magnetic particles biotinylated in vitro 6.3.4.15 biotin-[biotin carboxyl-carrier protein] ligase synthesis production of [35S]-biotin from Na-35SO4 and desthiobiotin with a specific activity of 30.7 Ci/mmol by expression of the biotinylation domain from the Plasmodium falciparum acetyl-CoA carboxylase in Escherichia coli as a biotinylation substrate. Overexpression of the biotin synthase, BioB, and biotin ligase, BirA, increases biotinylation of the biotinylation domain 160fold over basal levels. Biotinylated biotinylation domain is purified by affinity chromatography, and free biotin is liberated using acid hydrolysis 6.3.4.15 biotin-[biotin carboxyl-carrier protein] ligase synthesis site-directed antibody immobilization by fusing biotin carboxyl carrier protein BCCP to the IgG-binding domain of Protein A, and the resulting fusion protein is immobilized on the biotin protein ligase-modified gold surface of the sensor chip for quartz crystal microbalance through complexation between BCCP and biotin protein ligase. The layer of the IgG-binding domain is successfully captured the antibody, and the antibody retains high antigen-binding capability 6.3.5.1 NAD+ synthase (glutamine-hydrolysing) synthesis enzymatic method for efficient synthesis of NAD+ of high purity with 3H, 14C, or other labels at any nonexchangeable position of the NMN+ or AMP portions of the NAD+ molecule 6.3.5.2 GMP synthase (glutamine-hydrolysing) synthesis industrial production of 5-guanylic acid 6.3.5.2 GMP synthase (glutamine-hydrolysing) synthesis importance of guanine synthesis in immune cell function, GMP synthetase is a potential target for immunosuppressive therapy 6.3.5.11 cobyrinate a,c-diamide synthase synthesis co-expression of the cobA gene from Propionibacterium freudenreichii and the cbiA, -C, -D, -E, -T, -F, -G, -H, -J, -K, -L, and -P genes from Salmonella enterica serovar typhimurium in Escherichia coli result in the production of cobyrinic acid a,c-diamide. Strains that have neither cbiP nor cbiA synthesized 1-desmethylcobyrinic acid even in the presence of cbiD, suggesting that CbiA and CbiP are necessary for CbiD activity 6.4.1.1 pyruvate carboxylase synthesis super-transfection of CHO cells with a mammalian construct bearing codon optimized yeast cytosolic pyruvate carboxylase PYC2 and a strong fusion promoter for optimal expression of PYC2 enzyme in order to control the lactate metabolism of mAb (IgG1-kappa) producing CHO clones. Presence of Pyc2 results in an improved mAb titer up to 5%, galactosylation up to 2.5folds, and mannosylation up to twofold 6.4.1.1 pyruvate carboxylase synthesis the presence of 10 mM L-aspartate in the production medium directly represses pyruvate carboxylase expression. This leads to limited malate flux which in turn regulates the malate/citrate antiporters resulting in increasing cis-aconitate decarboxylase activity to simultaneously convert cis-aconitate, citrate isomer, into itaconic acid 6.4.1.2 acetyl-CoA carboxylase synthesis expression in Bombyx mori using the silkworm NPV bacmid system. Recombinant protein shows biotinylation capacity,and exhibits posttranslational biotinylation and phosphorylation 6.4.1.2 acetyl-CoA carboxylase synthesis in yeast engineered to produce the polyketide 6-methylsalisylic acid (6-MSA), both 6-MSA and native fatty acid levels increase by 3fold in presence of mutant S1157A which is not deactivated when glucose is depleted 6.4.1.3 propionyl-CoA carboxylase synthesis expression of the propionyl-CoA carboxylase complex from Streptomyces coelicolor supports the highest levels of heterologous polyketide production in Escherichia coli. The molar yield of 6-deoxyerythronolide B of Escherichia coli, harboring the wild-type Streptomyces coelicolor propionyl-CoA carboxylase, is 1.2% 6.5.1.1 DNA ligase (ATP) synthesis plays an important role for recombination of DNA fragments in vitro 6.5.1.2 DNA ligase (NAD+) synthesis DNA ligase is an indispensible reagent in the chemical synthesis of double-stranded DNA of specific nucleotide sequence 6.5.1.2 DNA ligase (NAD+) synthesis an important use of DNA ligase is the preparation of recombinant DNA molecules for use in the cloning of DNA 6.5.1.2 DNA ligase (NAD+) synthesis DNA ligase is used for cDNA cloning by replacement synthesis 6.5.1.3 RNA ligase (ATP) synthesis synthesis of oligoribonucleotides with defined sequence 6.5.1.3 RNA ligase (ATP) synthesis RNA ligase-mediated method for the efficient creation of large, synthetic RNAs 6.5.1.3 RNA ligase (ATP) synthesis selective isotope labeling of RNA by introducing a labeled RNA segment via a two-step protocol, which uses T4 DNA ligase and T4 RNA ligase 1, and a one-pot protocol, which uses T4 RNA ligase 1 alone. Protection of termini is not required, provided segmentation sites can be chosen such that the segments fold into the target structure or target-like structures and thus are not trapped into stable alternate structures. Labeling protocols are generally applicable to large RNA molecules and can be extended to more than three segments 6.5.1.3 RNA ligase (ATP) synthesis efficient synthesis of 5' pre-adenylated DNA 7.1.1.1 proton-translocating NAD(P)+ transhydrogenase synthesis overexpression of pyridine nucleotide transhydrogenase, PntAB, results in a significant increase in biomass and glycolic acid titer and yield. Improved redox homeostasis resulting from PntAB overexpression positively affects the anabolic rate of the cell. PntAB overexpression result in 154 and 37% increase in NAD+/NADH ratio, at 48 and 72 h, respectively, while the the NADP+ concentrations are 70 and 30% lower at 24 and 72 h. Expression of PntAB in an optimized glycolic acid-producing strain improves the growth and product titer significantly 7.1.1.3 ubiquinol oxidase (H+-transporting) synthesis inactivating the cytochrome bd-II oxidase of the aerobic respiratory chain is a simple and effective strategy to improve poly(3-hydroxybutyrate) biosynthesis in Escherichia coli 7.1.2.2 H+-transporting two-sector ATPase synthesis concomitant increase in shoot Na+ accumulation and leaf succulence of plants, at optimal salinity increased CO2 assimilation, stomatal conductance, sub-stomatal CO2 concentration, transpiration rate, Rubisco specific activity and both high nitrogen-use efficiency and photosynthetic nitrogen-use efficiency, higher salt levels impair photosynthetic capacity via a stomatal limitation 7.2.2.14 P-type Mg2+ transporter synthesis by overexpression of mgtA in Escherichia coli, a concentration of 32.41 g succinic acid per liter with a yield of 0.81 g per g glucose, can be obtained in a batch fermentation by using the low-cost mixture of Mg(OH)2 and NH3-H2O to replace MgCO3 as the alkaline neutralizer. The effect of the inhibitory compounds in lignocellulosic hydrolyzates on cell growth and succinic acid production can be relieved. In a 3-liter bioreactor, the overall productivity and yield of succinic acid in the whole anaerobic stage are 2.15 g per liter and h and 0.86 g per g total sugar, respectively 7.5.2.1 ABC-type maltose transporter synthesis overexpression of malEFG-a improves the utilization rate of starch, and thereby enhances avermectin production, which is useful for the improvement of commercial antibiotic production using starch as the main carbon source in the fermentation process