Covalent binding of benzo[a]pyrene to cytochrome P-450 beta NF-B2 and other proteins in reconstituted mixed-function oxidase systems.
A reconstituted mixed-function oxidase system, containing the major beta-naphthoflavone-induced isozyme of rat liver cytochrome P-450 bound benzo[a]pyrene covalently in the presence of NADPH. NADPH-cytochrome P-450 reductase was required for binding and a maximum rate of adduct formation was obtained at 8 units of reductase per nmol cytochrome P-450. Phosphatidylcholine inhibited this reaction. Benzo[a]pyrene was bound to the cytochrome, but not to the reductase, as shown by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Approximately 6 molecules of benzo[a]pyrene bound to each molecule cytochrome P-450 during prolonged incubations. No binding occurred when the beta-naphthoflavone-induced isozyme of cytochrome P-450 was replaced by the major isozyme induced by phenobarbital, but both cytochromes incorporated benzo[a]pyrene to approximately the same extent when they were incubated together in the presence of the reductase and NADPH. Metabolically activated benzo[a]pyrene also bound covalently to purified epoxide hydrolase, when this enzyme was added to the reconstituted mixed-function oxidase system.
D000818:Animals; D001564:Benzo(a)pyrene; D001571:Benzoflavones / Q000494:pharmacology; D001580:Benzopyrenes / Q000378:metabolism*; D002273:Carcinogens / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D004790:Enzyme Induction; D004851:Epoxide Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase / Q000302:isolation & purification / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D019324:beta-Naphthoflavone
D001571:Benzoflavones; D001580:Benzopyrenes; D002273:Carcinogens; D011506:Proteins; D001564:Benzo(a)pyrene; D019324:beta-Naphthoflavone; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D009251:NADPH-Ferrihemoprotein Reductase; D004851:Epoxide Hydrolases