Application | Comment | Organism |
---|---|---|
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 | Geobacillus stearothermophilus |
Protein Variants | Comment | Organism |
---|---|---|
D123N | mutation abolishes the oxidative activity. The carboxylate of D123 is the base needed for abstracting the proton to form the alkoxy intermediate that precedes the hydride transfer to the nicotinamide cofactor | Geobacillus stearothermophilus |
L252A | mutation reduces the enzyme activity to less than 10% of wild-type activity and sculpts the active site to accommodate a productive configuration of 3-monoalkyl glycerols | Geobacillus stearothermophilus |
L252A | the mutation sculpts the active site to accommodate a productive configuration of 3-monoalkyl glycerols. This mutation enhances the kcat 163-fold towards 3-ethoxypropan-1,2-diol, resulting in a specific activity similar to the one found for the wild-type towards glycerol. No activity with 3-phenoxypropane-1,2-diol | Geobacillus stearothermophilus |
V131A | about 70% of wild-type activity with glycerol | Geobacillus stearothermophilus |
Y142A | mutation reduces the enzyme activity to less than 10% of wild-type activity | Geobacillus stearothermophilus |
General Stability | Organism |
---|---|
the immobilised L252A shows good reusability and increased the operational stability | Geobacillus stearothermophilus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
7 | - |
3-phenoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
13.4 | - |
3-ethoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
30 | - |
3-propoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
31.2 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
45 | - |
3-methoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
50 | - |
glycerol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
78 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
162 | - |
3-butoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
281 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
371 | - |
3-ethoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
608 | - |
3-propoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
707 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
977 | - |
3-methoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
1880 | - |
glycerol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Geobacillus stearothermophilus | P32816 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
3-(2,2,2-trifluoroethoxy)propane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-(2,2,2-trifluoroethoxy)propan-2-one + NADH + H+ | - |
? | |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-butoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-butoxypropan-2-one + NADH + H+ | - |
? | |
3-butoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-ethoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-ethoxypropan-2-one + NADH + H+ | - |
? | |
3-ethoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-methoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-methoxypropan-2-one + NADH + H+ | - |
? | |
3-methoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-phenoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-phenoxypropan-2-one + NADH + H+ | - |
? | |
3-phenoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-propoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-propoxypropan-2-one + NADH + H+ | - |
? | |
3-propoxypropane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
3-[(propan-2-yl)oxy]propane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | 1-hydroxy-3-[(propan-2-yl)oxy]propan-2-one + NADH + H+ | - |
? | |
3-[(propan-2-yl)oxy]propane-1,2-diol + NAD+ | - |
Geobacillus stearothermophilus | ? + NADH + H+ | - |
? | |
glycerol + NAD+ | - |
Geobacillus stearothermophilus | glycerone + NADH + H+ | - |
? | |
glycerol + NAD+ | - |
Geobacillus stearothermophilus | D-glyceraldehyde + NADH + H+ | - |
? | |
additional information | enzyme is highly enantioselective towards S-isomers of substituted glycerol derivatives | Geobacillus stearothermophilus | ? | - |
- |
Synonyms | Comment | Organism |
---|---|---|
GldA | - |
Geobacillus stearothermophilus |
GlyDH | - |
Geobacillus stearothermophilus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.19 | - |
3-ethoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.27 | - |
3-propoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.3 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.32 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.7 | - |
3-phenoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.84 | - |
3-methoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
1.3 | - |
3-butoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
6.6 | - |
3-methoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
7.4 | - |
glycerol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
9 | - |
glycerol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
12 | - |
3-propoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
13 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
14 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
31 | - |
3-ethoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus |
General Information | Comment | Organism |
---|---|---|
metabolism | reaction mechanism leads to a proton transfer between the substrate and the acid residue before the hydride transfer from glycerol to NAD+. The mechanism takes place in a stepwise manner, consisting in the proton abstraction of the alcohol by D123 residue followed by the hydride transfer from glycerol to NAD+. This second step is the rate-limiting one | Geobacillus stearothermophilus |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.004 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.005 | - |
glycerol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.007 | - |
3-methoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.008 | - |
3-butoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.009 | - |
3-propoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.01 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.014 | - |
3-ethoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.018 | - |
3-[(propan-2-yl)oxy]propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.019 | - |
3-methoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.019 | - |
3-propoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.051 | - |
3-(2,2,2-trifluoroethoxy)propane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.085 | - |
3-ethoxypropane-1,2-diol | mutant L252A, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.112 | - |
3-phenoxypropane-1,2-diol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus | |
0.148 | - |
glycerol | wild-type, pH 7.0, 30°C | Geobacillus stearothermophilus |