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Literature summary extracted from

  • McQualter, R.B.; Petrasovits, L.A.; Gebbie, L.K.; Schweitzer, D.; Blackman, D.M.; Chrysanthopoulos, P.; Hodson, M.P.; Plan, M.R.; Riches, J.D.; Snell, K.D.; Brumbley, S.M.; Nielsen, L.K.
    The use of an acetoacetyl-CoA synthase in place of a beta-ketothiolase enhances poly-3-hydroxybutyrate production in sugarcane mesophyll cells (2015), Plant Biotechnol. J., 13, 700-707 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.3.1.194 gene nphT7, functional recombinant expression in sugarcane (Saccharum sp.) cultivar Q117, coexpression of gene nphT7 with genes phaB and phaC Streptomyces sp. CL190
6.2.1.16 recombinant enzyme expression Streptomyces lividans

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
6.2.1.16 purified recombinant enzyme SlAacS complexed with AMP and acetoacetate, hanging drop vapor diffusion method, mixing of 11 mg/ml protein and 1.5fold molar excess of buffered AMP and acetoacetate with a reservoir solution containing 0.7-1.2M K3citrate and 50 mM BTP-HCl, pH 7.0, 14°C, 1 week, X-ray diffraction structure determination and analysis, molecular replacement using full-length Salmonella enterica acetyl-CoA synthetase, PDB ID1PG4, stripped of waters and cofactors/substrates as search model Streptomyces lividans

Protein Variants

EC Number Protein Variants Comment Organism
2.3.1.194 additional information engineering the production of polyhydroxyalkanoates (PHAs) into high biomass bioenergy crops has the potential to provide a sustainable supply of bioplastics and energy from a single plant feedstock. One of the major challenges in engineering C4 plants for the production of poly[(R)-3-hydroxybutyrate] (PHB) is the significantly lower level of polymer produced in the chloroplasts of mesophyll cells compared to bundle sheath cells, thereby limiting the full PHB yield potential of the plant. The access to substrate for PHB synthesis may limit polymer production in mesophyll cell chloroplasts. The use of an acetoacetyl-CoA synthase, that catalyses the conversion of acetyl-CoA and malonyl-CoA to acetoacetyl-CoA with the release of carbon dioxide, in place of a beta-ketothiolase, the first enzyme in the bacterial PHA pathway, enhances poly-3-hydroxybutyrate production in sugarcane mesophyll cells. The engineered cells shows increased production of PHB and increased polymer molecular weight. PhaA is the main contributor to low PHB yield in mesophyll cells. Phenotypes of transgenic sugarcane leafs, overview Streptomyces sp. CL190

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.2.1.16 Mg2+ required Streptomyces lividans

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.3.1.194 acetyl-CoA + malonyl-CoA Streptomyces sp. CL190
-
 acetoacetyl-CoA + CoA + CO2
-
 ir
6.2.1.16 ATP + acetoacetate + CoA Streptomyces lividans
-
 AMP + diphosphate + acetoacetyl-CoA
-
 ?
6.2.1.16 ATP + acetoacetate + CoA Streptomyces lividans TK24
-
 AMP + diphosphate + acetoacetyl-CoA
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.3.1.194 Streptomyces sp. CL190 D7URV0
-
-
6.2.1.16 Streptomyces lividans D6EQU8
-
-
6.2.1.16 Streptomyces lividans TK24 D6EQU8
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
6.2.1.16 recombinant enzyme Streptomyces lividans

Source Tissue

EC Number Source Tissue Comment Organism Textmining

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
6.2.1.16 4.91 6.8 purified recombinant enzyme, pH 7.5, 30°C Streptomyces lividans

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.3.1.194 acetyl-CoA + malonyl-CoA
-
 Streptomyces sp. CL190 acetoacetyl-CoA + CoA + CO2
-
 ir
6.2.1.16 ATP + acetoacetate + CoA
-
 Streptomyces lividans AMP + diphosphate + acetoacetyl-CoA
-
 ?
6.2.1.16 ATP + acetoacetate + CoA
-
 Streptomyces lividans TK24 AMP + diphosphate + acetoacetyl-CoA
-
 ?

Synonyms

EC Number Synonyms Comment Organism
2.3.1.194 AACS
-
 Streptomyces sp. CL190
2.3.1.194 Acetoacetyl-CoA synthase
-
 Streptomyces sp. CL190
2.3.1.194 NphT7
-
 Streptomyces sp. CL190
6.2.1.16 AACS
-
 Streptomyces lividans
6.2.1.16 Acetoacetyl-CoA synthetase
-
 Streptomyces lividans
6.2.1.16 acsA1
-
 Streptomyces lividans
6.2.1.16 SlAacS
-
 Streptomyces lividans

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
6.2.1.16 30
-
 assay at Streptomyces lividans

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.2.1.16 5.89
-
 acetoacetate pH 7.5, 30°C, recombinant enzyme Streptomyces lividans
6.2.1.16 6.43
-
 ATP pH 7.5, 30°C, recombinant enzyme Streptomyces lividans
6.2.1.16 8.16
-
 CoA pH 7.5, 30°C, recombinant enzyme Streptomyces lividans

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
6.2.1.16 7.5
-
 assay at Streptomyces lividans

Cofactor

EC Number Cofactor Comment Organism Structure
6.2.1.16 ATP
-
 Streptomyces lividans

General Information

EC Number General Information Comment Organism
6.2.1.16 additional information in addition to the two catalytic states, additional conformations are observed crystallographically that likely play a role in allowing access and egress of substrates and products from the relatively buried active site. The enzyme shows conformational flexibility. Structure-function analysis, overview. The C-terminal domain undergoes a large conformational change in the catalytic mechanism of acyl-CoA synthetases, the C-terminal extension is important for catalytic activity, structure comparisons. One region from the N-terminal domain interacts is the so-called P-loop, a glycine-, serine-, and threonine-rich region that interacts with the phosphates of ATP. This P-loop adopts multiple conformations in the different crystal structures and may play an important role in the release of PPi and trigger the conformational change. Specifically, the main chain carbonyls of Ser272, Gly274, and Gly277 form direct or water-mediated hydrogen bonds with Asn637 and Ser640. Asn637 also interacts directly with Arg183 and Asp187, while the carbonyl of Gly639 and the carbonyl and side chain oxygens of Ser640 interact with Ser184, Asp187, and Arg188. Streptomyces lividans

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
6.2.1.16 23.6
-
 CoA pH 7.5, 30°C, recombinant enzyme Streptomyces lividans
6.2.1.16 26.6
-
 acetoacetate pH 7.5, 30°C, recombinant enzyme Streptomyces lividans
6.2.1.16 32.4
-
 ATP pH 7.5, 30°C, recombinant enzyme Streptomyces lividans