Cloned (Comment) | Organism |
---|---|
mutant enzymes expressed in Escherichia coli BL21(DE3)-RIL as His-tag fusion proteins | Rattus norvegicus |
Crystallization (Comment) | Organism |
---|---|
sitting drop method, complete data set collected for the D239T mutant enzyme | Rattus norvegicus |
Protein Variants | Comment | Organism |
---|---|---|
D239E | decreased activity with NADH and NADPH | Rattus norvegicus |
D239S | significantly increased activity with NADPH | Rattus norvegicus |
D239S/F251R | specific for NADPH | Rattus norvegicus |
D239S/F251Y | bispecific for NADH and NADPH | Rattus norvegicus |
D239T | specific for NADPH, 11fold preference for NADPH over NADH | Rattus norvegicus |
D239T/F251R | specific for NADPH | Rattus norvegicus |
F251R | minor effects on activity | Rattus norvegicus |
F251Y | minor effects on activity | Rattus norvegicus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.003 | - |
NADH | F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.005 | - |
NADH | D239S/F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.006 | - |
NADH | D239E mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.006 | - |
NADH | wild type enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.009 | - |
NADPH | D239T mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.012 | - |
NADH | F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.017 | - |
NADH | D239S mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.022 | - |
NADPH | D239S/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.025 | - |
NADPH | D239S/F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.044 | - |
NADPH | D239T/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.094 | - |
NADPH | D239E mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.119 | - |
NADH | D239T mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.138 | - |
NADPH | F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.224 | - |
NADH | D239S/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.268 | - |
NADPH | D239S mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.512 | - |
NADH | D239T/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.617 | - |
NADPH | F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
0.924 | - |
NADPH | wild type enzyme, pH 7.0, 25°C | Rattus norvegicus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
2 ferricytochrome b5 + NADH | Rattus norvegicus | involved in the synthesis of fatty acids and cholesterol, and in the oxidation of xenobiotics | 2 ferrocytochrome b5 + NAD+ + H+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Rattus norvegicus | P20070 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant enzymes | Rattus norvegicus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2 ferricyanide + NADPH | - |
Rattus norvegicus | 2 ferrocyanide + NADP+ + H+ | - |
? | |
2 ferricytochrome b5 + NADH | - |
Rattus norvegicus | 2 ferrocytochrome b5 + NAD+ + H+ | - |
? | |
2 ferricytochrome b5 + NADH | involved in the synthesis of fatty acids and cholesterol, and in the oxidation of xenobiotics | Rattus norvegicus | 2 ferrocytochrome b5 + NAD+ + H+ | - |
? |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5.2 | - |
NADPH | D239E mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
33 | - |
NADPH | wild type enzyme, pH 7.0, 25°C | Rattus norvegicus | |
50 | - |
NADPH | F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
50 | - |
NADPH | F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
200 | - |
NADPH | D239S/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
217 | - |
NADPH | D239S mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
250 | - |
NADH | D239S/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
267 | - |
NADPH | D239T mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
333 | - |
NADH | D239T mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
367 | - |
NADH | F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
417 | - |
NADPH | D239S/F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
433 | - |
NADH | D239S mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
467 | - |
NADH | D239S/F251Y mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
500 | - |
NADH | F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
517 | - |
NADH | D239E mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
550 | - |
NADPH | D239T/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
717 | - |
NADH | D239T/F251R mutant enzyme, pH 7.0, 25°C | Rattus norvegicus | |
800 | - |
NADH | wild type enzyme, pH 7.0, 25°C | Rattus norvegicus |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
FAD | - |
Rattus norvegicus | |
NADH | native enzyme shows 3700fold preference for NADH over NADPH | Rattus norvegicus | |
NADPH | preferred cofactor for some engineered enzymes | Rattus norvegicus |