Cloned (Comment) | Organism |
---|---|
D249A, D249S, and D249K mutants are expressed in Escherichia coli BL21(DE3) cells | Comamonas testosteroni |
Protein Variants | Comment | Organism |
---|---|---|
D249A | the mutation leads to 2fold increased Km value compared to the wild type, the mutant shows increased retention time, suggesting a smaller molecule size than dimeric wild type enzyme | Comamonas testosteroni |
D249A | mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 30fold decrease in catalytic efficiency, decrease in melting temperature | Comamonas testosteroni |
D249K | he mutation leads to 4fold increased Km value compared to the wild type, the mutant shows increased retention time, suggesting a smaller molecule size than dimeric wild type enzyme | Comamonas testosteroni |
D249K | mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 1.4fold decrease in catalytic efficiency, decrease in melting temperature | Comamonas testosteroni |
D249S | the mutant has similar kinetic parameters to wild type enzyme | Comamonas testosteroni |
D249S | mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 1400fold decrease in catalytic efficiency, decrease in melting temperature | Comamonas testosteroni |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.00305 | - |
NAD+ | mutant D249S, pH 10.2, 25°C | Comamonas testosteroni | |
0.0038 | - |
NAD+ | wild type enzyme, in 0.1 M CAPS buffer, at pH 10.2, at 25°C | Comamonas testosteroni | |
0.0038 | - |
NAD+ | wild-type, pH 10.2, 25°C | Comamonas testosteroni | |
0.005 | - |
NAD+ | mutant enzyme D249S, in 0.1 M CAPS buffer, at pH 10.2, at 25°C | Comamonas testosteroni | |
0.009 | - |
NAD+ | mutant enzyme D249K, in 0.1 M CAPS buffer, at pH 10.2, at 25°C | Comamonas testosteroni | |
0.009 | - |
NAD+ | mutant D249K, pH 10.2, 25°C | Comamonas testosteroni | |
0.016 | - |
NAD+ | mutant enzyme D249A, in 0.1 M CAPS buffer, at pH 10.2, at 25°C | Comamonas testosteroni | |
0.016 | - |
NAD+ | mutant D249A, pH 10.2, 25°C | Comamonas testosteroni |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
52000 | - |
calculated from amino acid sequence | Comamonas testosteroni |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Comamonas testosteroni | P80702 | - |
- |
Purification (Comment) | Organism |
---|---|
metal-chelate chromatography | Comamonas testosteroni |
Renatured (Comment) | Organism |
---|---|
thermal and urea-induced unfolding profiles for wild-type and mutant enzymes appear as a two-state transition and three-state transition, respectively | Comamonas testosteroni |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
androsterone + NAD+ | - |
Comamonas testosteroni | 5alpha-androstan-3,17-dione + NADH + H+ | - |
? | |
androsterone + NAD+ | - |
Comamonas testosteroni | 5alpha-androstane-3,17-dione + NADH + H+ | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | x-ray crystallography | Comamonas testosteroni |
More | concentration-dependent dimerization. The calculated molecular masses through gel filtration chromatography for wild-type, mutants D249A, D249K, and D249S at the concentration of 1 mg/ml are 52, 42, 43, and 50 kDa, and change to 47, 25, 26, and 34 kDa at 0.01 mg/ml, respectively | Comamonas testosteroni |
Synonyms | Comment | Organism |
---|---|---|
3alpha-HSD/CR | - |
Comamonas testosteroni |
3alpha-hydroxysteroid dehydrogenase/carbonyl reductase | - |
Comamonas testosteroni |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
47.2 | - |
melting temperature, mutant D249S | Comamonas testosteroni |
47.9 | - |
melting temperature, mutant D249A | Comamonas testosteroni |
49.41 | - |
melting temperature, mutant D249K | Comamonas testosteroni |
54 | - |
the melting temperature of the wild type enzyme is at 54.76°C | Comamonas testosteroni |
54.8 | - |
melting temperature, wild-type | Comamonas testosteroni |