Aminopyrine-N-demethylase. II. Characterization of a unique monooxygenase isoform P-450ap

A previously unidentified cytochrome P-450ap possessing the highest aminopyrine-N-demethylase activity has been isolated from liver microsomes of 4-isopropylaminoantipyrine-induced rats, using affinity chromatography in combination with ion-exchange chromatography with subsequent separation on a hydroxyapatite column. The isolated cytochrome P-450ap has the following characteristics: M r=49 kD, CO-peak maximum at 450.5 nm, rate of demethylation in a reconstituted system for aminopyrine of 25.5 nmoles of HCHO/min per nmole of P-450, and for benzphetamine a rate of 17.0nmoles of HCHO/min per nmole of P-450. The hemoprotein synthesis is paralleled by the synthesis of a protein with M r of 51 kD. Immunochemical analysis permitted the identification of the latter protein as cytochrome P-450_b. It was, demonstrated that cytochrome P-450ap does not interact with the antibodies to the major phenobarbital induced form, i.e. with cytochrome P-450_b.