Table 4.
Testosterone hydroxylation by 2B4dH/H226Y and mutants at 200 μM substrate.
| Turn over (nmol/min/nmol P450) | Stereoselectivity | Regioselectivity | |||
|---|---|---|---|---|---|
| P450 | 16α-OH | 16β-OH | 2α-OH | 16α-OH:16β-OH | 16-OH:2-OH |
| 2B4dH/H226Y | 0.43 | 0.06 | 0.17 | 7.2 | 2.9 |
| V292A | 0.26 | ND | 0.13 | ND | 2.0 |
| F296A | 0.44 | ND | 0.34 | ND | 1.3 |
| T302A | 0.01 | 0.03 | ND | 0.3 | ND |
| I363Aa | 3.7 | 1.4 | 0.51 | 2.7 | 10 |
| V367A | 0.17 | 0.31 | 0.07 | 0.6 | 6.9 |
| V367L | 0.07 | ND | 0.09 | ND | 0.80 |
| V477A | ND | ND | ND | ND | ND |
| V477F | 0.08 | 0.01 | 0.46 | 8.0 | 0.19 |
I363A also catalyzes P450 testosterone 15α-hydroxylation (12 nmol/min/nmol P450), whereas P450 2B4dH/H226Y showed non-determinable (ND) testosterone 15α-hydroxylase activity.
Results are the mean of at least three determinations. The standard error of mean (SEM) was approximately ± 25% of the mean values. SEM was not indicated in the Table to enhance the clarity.