Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21 (DE3) cells | Comamonas testosteroni |
Protein Variants | Comment | Organism |
---|---|---|
S114A | the mutation eliminates the hydrogen bonding interaction with P185, causing a conformational change in a nonproductive binding of NADH and a significant loss of activity, the mutant enzyme decreases 3100fold in V/Et value with no apparent change in Km value for substrates | Comamonas testosteroni |
S114A | mutant enzyme exhibits a pronounced increase in the magnitude of ellipticity at 222 nm. S114A mutant enzyme decreases 3100fold in catalytic efficiency with no apparent change in Km for substrates. Addition of NADH to S114A mutant enzyme induces a secondary structural change | Comamonas testosteroni |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
NADH | - |
Comamonas testosteroni |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0012 | - |
5alpha-androstane-3,17-dione | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.0012 | - |
5alpha-androstan-3,17-dione | wild type enzyme, in 0.1 M HEPES at pH 7.5 | Comamonas testosteroni | |
0.0031 | - |
5alpha-androstan-3,17-dione | mutant enzyme S114A, in 0.1 M HEPES at pH 7.5 | Comamonas testosteroni | |
0.0031 | - |
5alpha-androstane-3,17-dione | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni | |
0.004 | - |
NADH | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.004 | - |
NADH | wild type enzyme, in 0.1 M HEPES at pH 7.5 | Comamonas testosteroni | |
0.0065 | - |
NADH | mutant enzyme S114A, in 0.1 M HEPES at pH 7.5 | Comamonas testosteroni | |
0.0065 | - |
NADH | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Comamonas testosteroni | P80702 | - |
- |
Purification (Comment) | Organism |
---|---|
Ni-NTA column chromatography | Comamonas testosteroni |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
5alpha-androstane-3,17-dione + NADH + H+ | - |
Comamonas testosteroni | androsterone + NAD+ | - |
? | |
androsterone + NAD+ | - |
Comamonas testosteroni | 5alpha-androstan-3,17-dione + NADH + H+ | - |
? |
Synonyms | Comment | Organism |
---|---|---|
3alpha-hydroxysteroid dehydrogenase/carbonyl reductase | - |
Comamonas testosteroni |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.02 | - |
NADH | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni | |
63 | - |
NADH | wild-type, pH 7.5, 25°C | Comamonas testosteroni |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD+ | - |
Comamonas testosteroni | |
NADH | NADH association to enzyme involves a bimolecular binding step and isomerization. The binding of NADH into a hydrophobic pocket in the active site restricts its motion and shields the fluorescence quenching from solvent | Comamonas testosteroni |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0034 | - |
NADH | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.006 | - |
NADH | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
3.8 | - |
NADH | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni | |
6.5 | - |
5alpha-androstane-3,17-dione | mutant S114A, pH 7.5, 25°C | Comamonas testosteroni | |
15000 | - |
NADH | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
54000 | - |
5alpha-androstane-3,17-dione | wild-type, pH 7.5, 25°C | Comamonas testosteroni |