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Literature summary for 1.1.1.4 extracted from

  • Cui, Z.; Zhang, J.; Fan, X.; Zheng, G.; Chang, H.; Wei, W.
    Highly efficient bioreduction of 2-hydroxyacetophenone to (S)- and (R)-1-phenyl-1,2-ethanediol by two substrate tolerance carbonyl reductases with cofactor regeneration (2017), J. Biotechnol., 243, 1-9 .
    View publication on PubMed

Application

Application Comment Organism
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 Bacillus subtilis subsp. subtilis

Cloned(Commentary)

Cloned (Comment) Organism
gene bdhA, recombinant expression of His-tagged enzyme BDHA in Escherichia coli strain BL21 (DE3), co-expression with polyol dehydrogenase (GoSCR) from Gluconobacter oxydans 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) Bacillus subtilis subsp. subtilis

Protein Variants

Protein Variants Comment Organism
additional information in vitro bioreduction of 2-hydroxyacetophenone (2-HAP) is catalyzed by BDHA coupled with glucose dehydrogenase (GDH) from Bacillus subtilis for cofactor regeneration. The two coexpressed enantiocomplementary carbonyl reductases, BDHA 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, method optimization, overview. Products (R)-PED and (S)-PED are obtained with 99% yield, over 99% enantiomeric excess and 18.0 g/l/h volumetric productivity. The reaction is carried out in 5 ml sodium phosphate buffer (pH 7.0, 100 mM) at 30┬░C, containing 10 U/ml BDHA (cell free extract of Escherichia coli (BDHA)), 15 U/ml GoSCR (cell free extract of Escherichia coli (GoSCR)), 10 U/ml GDH (cell free extract of Escherichia coli (GDH)), 50-200 mM 2-HAP (with 10% DMSO as co-solvent), 60-250 mM D-glucose. Strong tolerance of BDHA and GoSCR against high substrate concentration Bacillus subtilis subsp. subtilis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics Bacillus subtilis subsp. subtilis
1
-
2-hydroxyacetophenone pH 7.0, 25┬░C, recombinant His-tagged enzyme Bacillus subtilis subsp. subtilis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2-hydroxyacetophenone + NADH + H+ Bacillus subtilis subsp. subtilis
-
(R)-1-phenyl-1,2-ethanediol + NAD+
-
?
2-hydroxyacetophenone + NADH + H+ Bacillus subtilis subsp. subtilis 168
-
(R)-1-phenyl-1,2-ethanediol + NAD+
-
?

Organic Solvent Stability

Organic Solvent Comment Organism
additional information DMSO is selected as the co-solvent, it does not affect the activity at up 30% v/v, but is inhibitory above Bacillus subtilis subsp. subtilis

Organism

Organism UniProt Comment Textmining
Bacillus subtilis subsp. subtilis O34788
-
-
Bacillus subtilis subsp. subtilis 168 O34788
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged BDHA from Escherichia coli strain BL21 (DE3) by nickel affinity chromatography and dialysis Bacillus subtilis subsp. subtilis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2-hydroxyacetophenone + NADH + H+
-
Bacillus subtilis subsp. subtilis (R)-1-phenyl-1,2-ethanediol + NAD+
-
?
2-hydroxyacetophenone + NADH + H+ over 99% enantiomeric excess Bacillus subtilis subsp. subtilis (R)-1-phenyl-1,2-ethanediol + NAD+
-
?
2-hydroxyacetophenone + NADH + H+
-
Bacillus subtilis subsp. subtilis 168 (R)-1-phenyl-1,2-ethanediol + NAD+
-
?
2-hydroxyacetophenone + NADH + H+ over 99% enantiomeric excess Bacillus subtilis subsp. subtilis 168 (R)-1-phenyl-1,2-ethanediol + NAD+
-
?
additional information an enantiocomplementary carbonyl reductase, 2,3-butanediol dehydrogenase (BDHA) from Bacillus subtilis is discovered to convert 2-hydroxyacetophenone (2-HAP) to (R)-1-phenyl-1,2-ethanediol ((R)-PED) with excellent stereochemical selectivity. No activity with NADPH Bacillus subtilis subsp. subtilis ?
-
-
additional information an enantiocomplementary carbonyl reductase, 2,3-butanediol dehydrogenase (BDHA) from Bacillus subtilis is discovered to convert 2-hydroxyacetophenone (2-HAP) to (R)-1-phenyl-1,2-ethanediol ((R)-PED) with excellent stereochemical selectivity. No activity with NADPH Bacillus subtilis subsp. subtilis 168 ?
-
-

Synonyms

Synonyms Comment Organism
2,3-butanediol dehydrogenase
-
Bacillus subtilis subsp. subtilis
BdhA
-
Bacillus subtilis subsp. subtilis

Temperature Optimum [┬░C]

Temperature Optimum [┬░C] Temperature Optimum Maximum [┬░C] Comment Organism
40
-
reduction of 2-HAP Bacillus subtilis subsp. subtilis

Temperature Stability [┬░C]

Temperature Stability Minimum [┬░C] Temperature Stability Maximum [┬░C] Comment Organism
50
-
purified recombinant His-tagged enzyme, inactivation within 2 h Bacillus subtilis subsp. subtilis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6
-
reduction of 2-HAP Bacillus subtilis subsp. subtilis

pH Stability

pH Stability pH Stability Maximum Comment Organism
7
-
purified recombinant His-tagged enzyme, more than 80% residual activity after 18 h at pH 7.0. The activity decreases significantly in acidic circumstances Bacillus subtilis subsp. subtilis

Cofactor

Cofactor Comment Organism Structure
additional information no activity with NADPH Bacillus subtilis subsp. subtilis
NADH dependent on Bacillus subtilis subsp. subtilis