Cloned (Comment) | Organism |
---|---|
expression in Escherichia coli | Comamonas testosteroni |
Protein Variants | Comment | Organism |
---|---|---|
K159A | decrease in the catalytic constant by 56fold and increase in the dissociation constant by 75fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis | Comamonas testosteroni |
K159A | decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by residues K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis. Residue N86 is important for stabilizing the position of K159 | Comamonas testosteroni |
N86A | decrease in the catalytic constant by 37fold and increase in the dissociation constant by 8fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Residue N86 is important for stabilizing the position of K159 | Comamonas testosteroni |
N86A | decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by residues K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis. Residue N86 is important for stabilizing the position of K159 | Comamonas testosteroni |
Y155F | decrease in the catalytic constant by 220fold and increase in the dissociation constant by 3fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis | Comamonas testosteroni |
Y155F | decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants | Comamonas testosteroni |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
NADH | - |
Comamonas testosteroni |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0012 | - |
5alpha-androstan-3,17-dione | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.0012 | - |
5beta-androstane-3,17-dione | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.0022 | - |
5alpha-androstan-3,17-dione | mutant N86A, pH 7.5, 25°C | Comamonas testosteroni | |
0.0022 | - |
5beta-androstane-3,17-dione | mutant N86A, pH 7.5, 25°C | Comamonas testosteroni | |
0.004 | - |
NADH | wild-type, pH 7.5, 25°C | Comamonas testosteroni | |
0.005 | - |
5alpha-androstan-3,17-dione | mutant Y155F, pH 7.5, 25°C | Comamonas testosteroni | |
0.005 | - |
5beta-androstane-3,17-dione | mutant Y155F, pH 7.5, 25°C | Comamonas testosteroni | |
0.007 | - |
5alpha-androstan-3,17-dione | mutant K159A, pH 7.5, 25°C | Comamonas testosteroni | |
0.007 | - |
5beta-androstane-3,17-dione | mutant K159A, pH 7.5, 25°C | Comamonas testosteroni | |
0.01 | - |
NADH | mutant N86A, pH 7.5, 25°C | Comamonas testosteroni | |
0.029 | - |
NADH | mutant Y155F, pH 7.5, 25°C | Comamonas testosteroni |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Comamonas testosteroni | P80702 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
5alpha-androstan-3,17-dione + NADH + H+ | - |
Comamonas testosteroni | androsterone + NAD+ | - |
? | |
5beta-androstane-3,17-dione + NADH + H+ | - |
Comamonas testosteroni | 5beta-androstan-3alpha-ol-17-one + NAD+ | - |
? |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NADH | - |
Comamonas testosteroni |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0034 | - |
NADH | wild-type, pH 7.5, 25°C, steady-state kinetic study | Comamonas testosteroni | |
0.004 | - |
NADH | mutant Y155F, pH 7.5, 25°C, steady-state kinetic study | Comamonas testosteroni | |
0.0057 | - |
NADH | mutant Y155F, pH 7.5, 25°C, product inhibition study | Comamonas testosteroni | |
0.006 | - |
NADH | wild-type, pH 7.5, 25°C, product inhibition study | Comamonas testosteroni | |
0.01 | - |
NADH | mutant N86A, pH 7.5, 25°C, steady-state kinetic study | Comamonas testosteroni | |
0.033 | - |
NADH | mutant N86A, pH 7.5, 25°C, product inhibition study | Comamonas testosteroni | |
0.161 | - |
NADH | mutant K159A, pH 7.5, 25°C, product inhibition study | Comamonas testosteroni | |
0.585 | - |
NADH | mutant K159A, pH 7.5, 25°C, steady-state kinetic study | Comamonas testosteroni |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.0034 | - |
wild-type, pH 7.5, 25°C | Comamonas testosteroni | NADH | |
0.004 | - |
mutant Y155F, pH 7.5, 25°C | Comamonas testosteroni | NADH | |
0.01 | - |
mutant N86A, pH 7.5, 25°C | Comamonas testosteroni | NADH | |
0.585 | - |
mutant K159A, pH 7.5, 25°C | Comamonas testosteroni | NADH |