Differential cumene hydroperoxide sensitivity of cytochrome P-450 enzymes IA1 and IIB1 determined by their way of membrane incorporation

Abstract

The cytochrome P-450-dependent O-dealkylation of all alkoxyresorufins was used to study the effect of cumene hydroperoxide on cytochrome P-450 IIB1 and IA1 in microsomal and reconstituted systems. In liver microsomal systems from respectively phenobarbital and 3-methylcholanthrene pretreated male Wistar rats, cytochrome P-450 IIB1-dependent pentoxyresorufin- O-dealkylation appeared to be more sensitive to cumene hydroperoxide treatment than cytochrome P-450 IA1-dependent ethoxyresorufin- O-dealkylation. This phenomenon was also observed when the cumene hydroperoxide sensitivity of P-450 IIB1 and IA1 was studied in an isosafrole pretreated rat liver microsomal system. The decrease in alkoxy- O-dealkylating activities appeared to proceed by destruction of the cytochrome P-450 component of the enzyme system. Purification and reconstitution of the enzyme system components in a system in which the isolated proteins were not incorporated into a membrane resulted in the disappearance of the difference in sensitivity between the two P-450 enzymes. However, in a reconstituted system with membrane incorporated proteins, again cytochrome P-450 IIB1 expressed a higher sensitivity towards cumene hydroperoxide than cytochrome P-450 IA1. From this it was concluded tha the differential cumene hydroperoxide sensitivity of cytochrome P-450 IIB1 and IA1 is not caused by an intrinsic difference in their sensitivity but by a differential effect of membrane incorporated on their cumene hydropeoxide sensitivity.