SUBSTRATE INFLUENCE ON INTERACTION BETWEEN CYTOCHROME-P450 AND CYTOCHROME B(5) IN MICROSOMES

We have been able to demonstrate that cytochrome bg interacts closely with cytochrome P450 in the microsomal membrane, and that substrates c an serve to order the interaction: Using the water-soluble carbodiimid e EDC we could crosslink cytochrome b(5) and CYP2B4 in microsomes from phenobarbital-treated rabbits and cytochrome b(5) and CYP1A2 in micro somes from beta-naphthoflavone-treated animals. The substrate benzphet amine increased the specific interaction between cytochrome b(5) and C YP2B4, decreasing the formation of higher molecular weight oligomeric complexes with cytochrome b(5). In contrast, no ordering of the intera ctions were obtained on addition of 7-ethoxycoumarin, a substrate of C YP1A2, or of benzphetamine to microsomes of beta-naphthoflavone-treate d animals in the presence of EDC. Of interest, although evidence could be shown for complementary charge-pairing between cytochrome b(5) and a number of other microsomal proteins in the membranes, and while the extent of CYP1A2 and CYP2B4 interaction with cytochrome b(5) each exc eeded 30% in the presence of substrate, no significant complexation of the P450s was obtained with any other microsomal proteins. (C) 1996 A cademic Press, Inc.