Table 1

P-450 content and reductase activity in membranes isolated fromE. coli-expressing P-450s and CPR

ConstructP-450 Yield and ContentCPR ActivityCPR Content1-a
nmol/L culture nmol/mg membrane protein nmol cyt. c reduced/min/mg membrane protein nmol/mg membrane protein
CYP3A4/pJR7250  ± 430.43  ± 0.18450  ± 580.15
CYP2D6/pJR7235  ± 480.37  ± 0.12423  ± 490.14
CYP3A4/CYP2D6/pJR7222  ± 190.26  ± 0.07420  ± 360.14
CYP3A4/CYP2D6/pDRlacZ167  ± 240.42  ± 0.1299  ± 290.033

Membranes were isolated from E. coli-expressing CYP2D6 or CYP3A4 or a combination of these P-450s with either high (pJR7) or low levels (pDRlacZ) of CPR. Expression was carried out as described inExperimental Procedures. P-450 contents were measured by Fe2+ versus Fe2+-CO difference spectroscopy. CPR activity was assayed by determining reduction of cytochromec. Values are given as means of six experiments ±S.D.

  • 1-a Calculated from cytochrome c reductase activity and known activity of purified CPR towards cytochromec (3000 nmol cytochrome c reduced/nmol CPR/min) (Nadler and Strobel, 1991).