Table 2.
Kinetic analysis of the POR mutant p.Y607C according to yeast microsomal coexpression assays of either WT or mutant POR with human CYP17A1, CYP21A2, and CYP19A1
| CYP17A1
|
CYP21A2
|
CYP19A1
|
|||
|---|---|---|---|---|---|
| 17α-Hydroxylase Prog → 17OHP | 17,20-Lyase (classic) 17Preg → DHEA | 17,20-Lyase (alternative) 5pdiolone → An | 21-Hydroxylase Prog → DOC | Aromatase A → E1 | |
| Vmax (pmol/μg · min) | |||||
| WT | 0.24 ± 0.02 | 0.22 ± 0.00 | 0.27 ± 0.01 | 0.30 ± 0.02 | 0.014 ± 0.001 |
| p.Y607C | 0.21 ± 0.02 | 0.13 ± 0.01 | 0.16 ± 0.00 | 0.27 ± 0.02 | 0.010 ± 0.001 |
| Km (μm) | |||||
| WT | 0.26 ± 0.10 | 0.71 ± 0.04 | 0.85 ± 0.14 | 0.24 ± 0.08 | 180 ± 44 |
| p.Y607C | 0.41 ± 0.13 | 0.95 ± 0.11 | 0.75 ± 0.06 | 0.27 ± 0.10 | 262 ± 54 |
| Catalytic efficiency Vmax/Km (% WT) | |||||
| p.Y607C | 56 ± 3 | 44 ± 4 | 66 ± 3 | 79 ± 2 | 56 ± 3 |
All assays were carried out in three independent triplicate experiments; results are presented as means ± sem. For CYP17A1, both 17α-hdyroxylase and 17,20-lyase activities within the classic and alternative androgen pathway were determined. Prog, Progesterone; 17Preg, 17-hydroxypregnenolone; 5pdiolone, 5-pregnanediolone; DOC, 11-deoxycortiocosterone; A, androstenedione; E1, estrone.