Microbial transformation of steroids--IX. Purification of progesterone hydroxylase cytochrome P-450 from Phycomyces blakesleeanus.

Progesterone hydroxylase cytochrome P-450 was purified to homogeneity from Phycomyces blakesleeanus microsomes by a four step procedure. An M(r) value of 60,000 was determined for this protein by SDS-PAGE. The DEAE-cellulose and Blue-1 MIMETIC affinity fractions gave major peaks at 452 nm in a dithionite-reduced, carbon monoxide, difference spectrum. NaIO4-dependent progesterone hydroxylation was obtained by the pure enzyme without NADPH and NADPH-cytochrome P-450 reductase. NADPH-dependent hydroxylation required the addition of other Phycomyces microsomal proteins present in the Blue-1 fraction.