Adrenodoxin-cytochrome P450scc interaction as revealed by EPR spectroscopy: Comparison with the putidaredoxin-cytochrome P450cam system

The cholesterol side-chain cleavage reaction catalyzed by cytochrome P450sc c comprises three consecutive monooxygenase reactions (22R-hydroxylation, 2 0S-hydroxylation, and C-20-C-22 bond scission) that produces pregnenolone. The electron equivalents necessary for the oxygen activation are supplied f rom a 2Fe-2S type ferredoxin, adrenodoxin. We found that 1:1 stoichiometric binding of oxidized adrenodoxin to oxidized cytochrome P450scc complexed w ith cholesterol or 25-hydroxycholesterol caused shifts of the high-spin EPR signals of the heme moiety at 5 K. Such shifts were not observed for the l ow-spin EPR signals. Ligation of CO or NO to the reduced heme of cytochrome P450scc complexed with reduced adrenodoxin and various steroid substrates did not cause any change in the axial EPR spectrum of the reduced iron-sulf ur center at 77 K. These results are in remarkable contrast to those obtain ed for the cytochrome P450cam-d-camphor-putidaredoxin ternary complex, sugg esting that the mode of cross talk between adrenodoxin and cytochrome P450s cc is very different from that in the Pseudomonas system. The difference ma y be primarily due to the location of the charged amino acid residues of th e ferredoxins important for the interaction with the partner cytochrome P45 0.