Purification and characterization of the major hepatic cannabinoid hydroxylase in the mouse: a possible member of the cytochrome P-450IIC subfamily.
Acute cannabidiol treatment of mice inactivated hepatic microsomal cytochrome P-450IIIA (P-450IIIA) and markedly inhibited in vitro cannabinoid metabolism. Antibodies raised against purified P-450IIIA inhibited the microsomal formation of quantitatively minor cannabinoid metabolites but had no effect on the major metabolites. Cannabinoid hydroxylation to the major metabolites was used as a functional probe to isolate and purify a P-450 (termed P-450THC) from hepatic microsomes of untreated mice. The purified protein had an apparent molecular weight of 47,000 and a specific content of 15.4 nmol/mg and exhibited an absorbance maximum at 452 nm for the reduced carbon monoxide complex. NH2-terminal sequence analysis of the first 16 residues of P-450THC suggests that it is a member of the P-450IIC subfamily, because its sequence is 85 and 69% identical to published sequences of rat hepatic P-450IIC7 and P-450IIC6, respectively. P-450THC exhibited high activity for cannabinoid hydroxylation and specifically produced 6 alpha- and 7-hydroxy-delta 1-tetrahydrocannabinol, as well as 6 alpha-, 7-, and 4"-hydroxycannabidiol. Unlike anti-P-450IIIA antibody, antibody raised against purified P-450THC markedly inhibited the microsomal formation of all major cannabinoid metabolites. Similar immunoinhibition studies also revealed the existence of orthologs of mouse P-450THC and P-450IIIA in human liver microsomes. Thus, cannabidiol treatment of mice resulted in the inactivation of at least two constitutive P-450 isozymes, which together account for the majority of the detected cannabinoid metabolites.
Bornheim
L M
LM
Department of Pharmacology, University of California, San Francisco 94143.
Correia
M A
MA
D000595:Amino Acid Sequence; D000818:Animals; D002185:Cannabidiol / Q000494:pharmacology; D002186:Cannabinoids / Q000378:metabolism*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000302:isolation & purification*; D007527:Isoenzymes / Q000302:isolation & purification*; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D051381:Rats
D002186:Cannabinoids; D065607:Cytochrome P-450 Enzyme Inhibitors; D007527:Isoenzymes; D002185:Cannabidiol; D003577:Cytochrome P-450 Enzyme System