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Literature summary extracted from
- Purification and identification of an Escherichia coli beta-keto ester reductase as 2,5-diketo-D-gluconate reductase YqhE (2002), Biotechnol. Prog., 18, 257-261.
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 1.1.1.274 | gne yqhE, DNA and amino acid sequence determination and analysis | Escherichia coli |
KM Value [mM]
| EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 1.1.1.274 | additional information | - |
additional information | - |
Escherichia coli | |
| 1.1.1.274 | 3.1 | - |
ethyl-2-methylacetoacetate | - |
Escherichia coli |  |
| 1.1.1.274 | 3.1 | - |
ethyl 2-methylacetoacetate | pH 7.0, 30°C, with a cofactor level 20fold higher than the Km value | Escherichia coli | |
| 1.1.1.346 | 3.1 | - |
ethyl 2-methylacetoacetate | at pH 7.0 and 30°C | Escherichia coli | |
Molecular Weight [Da]
| EC Number | Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|---|
| 1.1.1.274 | 31800 | - |
1 * 31800, SDS-PAGE | Escherichia coli |
| 1.1.1.346 | 31800 | - |
x * 31800, estimated from SDS-PAGE | Escherichia coli |
Natural Substrates/ Products (Substrates)
| EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 1.1.1.274 | ethyl 2-ethylacetoacetate + NADPH | Escherichia coli | stereospecific reaction | ethyl (2R)-ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl 2-methylacetoacetate + NADPH | Escherichia coli | stereospecific reaction | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl acetoacetate + NADPH | Escherichia coli | stereospecific reaction | ethyl (3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | additional information | Escherichia coli | substrate specificity, no activity with ethyl propionyl acetate and ethyl butyryl acetate | ? | - |
? | |
| 1.1.1.274 | additional information | Escherichia coli BL21-(DE3) | substrate specificity, no activity with ethyl propionyl acetate and ethyl butyryl acetate | ? | - |
? | |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 1.1.1.274 | Escherichia coli | - |
- |
- |
| 1.1.1.346 | Escherichia coli | - |
- |
- |
| 1.1.1.346 | Escherichia coli BL21-(DE3) | - |
- |
- |
Purification (Commentary)
| EC Number | Purification (Comment) | Organism |
|---|---|---|
| 1.1.1.274 | ammonium sulfate, DEAE-Sepharose, Matrix Red-A, Sephacryl S-200, hypatite C | Escherichia coli |
| 1.1.1.274 | native enzyme from strain BL21(DE3), 730fold to homogeneity by ammonium sulfate fractionation, DEAE ion exchange and Red-A affinity chromatography, ultrafiltration, hypatite C-resin chromatography, and a second ultrafiltration step | Escherichia coli |
| 1.1.1.346 | ammonium sulfate precipitation, DEAE-Sepharose column chromatography, Matrex Red-A column chromatography, Sephacryl S-200 gel filtration, and hypatite C cholumn chromatography | Escherichia coli |
Reaction
| EC Number | Reaction | Comment | Organism | Reaction ID |
|---|---|---|---|---|
| 1.1.1.274 | 2-dehydro-D-gluconate + NADP+ = 2,5-didehydro-D-gluconate + NADPH + H+ | enzyme also possesses beta-keto ester reductase activity | Escherichia coli |
Specific Activity [micromol/min/mg]
| EC Number | Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|---|
| 1.1.1.274 | 6.3 | - |
- |
Escherichia coli |
| 1.1.1.346 | 6.3 | - |
purified enzyme, at pH 7.0 and 30°C | Escherichia coli |
Storage Stability
| EC Number | Storage Stability | Organism |
|---|---|---|
| 1.1.1.274 | 4°C, purified enzyme, 400 mM potassium phoshate buffer, pH 7.0, several weeks without appreciable loss of activity | Escherichia coli |
| 1.1.1.346 | 4°C, potassium buffer, several weeks, no loss of activity | Escherichia coli |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 1.1.1.274 | 2,5-didehydro-D-gluconate + NADPH | - |
Escherichia coli | 2-keto-L-gulonate + NADP+ | - |
? | |
| 1.1.1.274 | 2,5-didehydro-D-gluconate + NADPH | - |
Escherichia coli BL21-(DE3) | 2-keto-L-gulonate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl 2-ethylacetoacetate + NADPH | stereospecific reaction | Escherichia coli | ethyl (2R)-ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl 2-methylacetoacetate + NADPH | stereospecific reaction | Escherichia coli | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl 2-methylacetoacetate + NADPH + H+ | 7% activity with NADH | Escherichia coli | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl 2-methylacetoacetate + NADPH + H+ | 7% activity with NADH | Escherichia coli BL21-(DE3) | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl acetoacetate + NADPH | stereospecific reaction | Escherichia coli | ethyl (3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl-(2R)-allylacetoacetate + NADPH | 250% of activity with ethyl-2-methylacetoacetate | Escherichia coli | ? | - |
? | |
| 1.1.1.274 | ethyl-(2R)-ethylacetoacetate + NADPH | 120% of activity with ethyl-2-methylacetoacetate | Escherichia coli | ethyl-(2R)-ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethyl-(2R)-ethylacetoacetate + NADPH | 120% of activity with ethyl-2-methylacetoacetate | Escherichia coli BL21-(DE3) | ethyl-(2R)-ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethylacetoacetate + NADPH | 53% of activity with ethyl-2-methylacetoacetate | Escherichia coli | ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | ethylacetoacetate + NADPH | 53% of activity with ethyl-2-methylacetoacetate | Escherichia coli BL21-(DE3) | ethyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.274 | additional information | substrate specificity, no activity with ethyl propionyl acetate and ethyl butyryl acetate | Escherichia coli | ? | - |
? | |
| 1.1.1.274 | additional information | substrate specificity, no activity with ethyl propionyl acetate and ethyl butyryl acetate | Escherichia coli BL21-(DE3) | ? | - |
? | |
| 1.1.1.346 | ethyl 2-acetylpent-4-enoate + NADPH + H+ | 250% activity compared to ethyl 2-methylacetoacetate | Escherichia coli | ethyl (2R)-2-[(1S)-1-hydroxyethyl]pent-4-enoate + NADP+ | - |
? | |
| 1.1.1.346 | ethyl 2-ethyl-3-oxobutanoate + NADPH + H+ | 120% activity compared to ethyl 2-methylacetoacetate | Escherichia coli | ethyl (2R,3S)-2-ethyl-3-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.346 | ethyl 2-methylacetoacetate + NADH + H+ | 7% activity with NADH compared to NADPH | Escherichia coli | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NAD+ | - |
? | |
| 1.1.1.346 | ethyl 2-methylacetoacetate + NADPH + H+ | - |
Escherichia coli | ethyl (2R)-methyl-(3S)-hydroxybutanoate + NADP+ | - |
? | |
| 1.1.1.346 | ethyl acetoacetate + NADPH + H+ | 53% activity compared to ethyl 2-methylacetoacetate | Escherichia coli | ethyl (3S)-3-hydroxybutanoate + NADP+ | - |
? | |
Subunits
| EC Number | Subunits | Comment | Organism |
|---|---|---|---|
| 1.1.1.274 | monomer | 1 * 31800, SDS-PAGE | Escherichia coli |
| 1.1.1.346 | ? | x * 31800, estimated from SDS-PAGE | Escherichia coli |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 1.1.1.274 | 2,5-diketo-D-gluconate reductase | - |
Escherichia coli |
| 1.1.1.274 | 2,5-DKG | - |
Escherichia coli |
| 1.1.1.274 | beta-keto ester reductase | - |
Escherichia coli |
| 1.1.1.274 | YqhE | - |
Escherichia coli |
| 1.1.1.346 | 2,5-diketo-D-gluconate reductase | - |
Escherichia coli |
| 1.1.1.346 | 2,5-DKG reductase | - |
Escherichia coli |
| 1.1.1.346 | YqhE | - |
Escherichia coli |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 1.1.1.274 | 30 | - |
assay at | Escherichia coli |
Turnover Number [1/s]
| EC Number | Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 1.1.1.274 | 2.6 | - |
ethyl-2-methylacetoacetate | - |
Escherichia coli | |
| 1.1.1.274 | 2.6 | - |
ethyl 2-methylacetoacetate | pH 7.0, 30°C | Escherichia coli | |
| 1.1.1.346 | 2.6 | - |
ethyl 2-methylacetoacetate | at pH 7.0 and 30°C | Escherichia coli | |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 1.1.1.274 | 7 | - |
assay at | Escherichia coli |
Cofactor
| EC Number | Cofactor | Comment | Organism | Structure |
|---|---|---|---|---|
| 1.1.1.274 | NADH | 7% of the activity with NADPH | Escherichia coli | |
| 1.1.1.274 | NADP+ | - |
Escherichia coli | |
| 1.1.1.274 | NADPH | - |
Escherichia coli | |
| 1.1.1.274 | NADPH | dependent on | Escherichia coli | |
| 1.1.1.346 | NADPH | dependent on | Escherichia coli | |
pI Value
| EC Number | Organism | Comment | pI Value Maximum | pI Value |
|---|---|---|---|---|
| 1.1.1.346 | Escherichia coli | isoelectric focusing | - |
5.9 |
General Information
| EC Number | General Information | Comment | Organism |
|---|---|---|---|
| 1.1.1.346 | metabolism | the enzyme reduces 2,5-didehydro-D-gluconate, a key step in the microbial synthesis of vitamin C | Escherichia coli |
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