Evidence for functional and structural multiplicity of pregnenolone-16 alpha-carbonitrile-inducible cytochrome P-450 isozymes in rat liver microsomes.
Administration of pregnenolone-16 alpha-carbonitrile (PCN) to adult female rats caused a 2-fold increase in total liver microsomal cytochrome P-450 along with 5-7-fold increases in four in vitro monooxygenase activities considered diagnostic for the major PCN-inducible cytochrome P-450 isozyme. However, upon administration of chloramphenicol to PCN-treated rats, these monooxygenase activities could be resolved into three groups. Thus, the ability of the microsomes to convert triacetyloleandomycin to a metabolite that forms a spectral complex with the reduced heme iron was decreased by 80% by chloramphenicol, whereas only a 50% decrease was observed in the rate of conversion of (R)-warfarin to its 9,10-dehydro metabolite and in the rate of 6 beta-hydroxylation of androstenedione. More strikingly, the 10-hydroxylation of (R)-warfarin was actually enhanced 2-fold by the chloramphenicol treatment. Fractionation studies were carried out on liver microsomes from PCN-treated adult male rats, and two highly purified cytochromes P-450, referred to as PCNa and PCNb, were recovered. PCNb was found to be identical in the sequence of the first 15 amino acid residues with a PCN-inducible isozyme, the complete amino acid sequence of which has recently been deduced in another laboratory [Gonzalez, F. J., Nebert, D. W., Hardwick, J. P., & Kasper, C. B. (1985) J. Biol. Chem. 260, 7435-7441]. The other isozyme, PCNa, differed in amino acid sequence in three of the first 15 positions from PCNb. Upon immunoblot analysis, polyclonal antibodies raised to PCNb also recognized PCNa. Thus, the PCN-inducible family of rat liver cytochrome P-450 comprises at least two separate proteins.
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D002701:Chloramphenicol / Q000494:pharmacology; D003101:Collodion; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D007527:Isoenzymes / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D010105:Oxygenases / Q000378:metabolism; D011285:Pregnenolone Carbonitrile / Q000494:pharmacology*; D011487:Protein Conformation; D051381:Rats; D011919:Rats, Inbred Strains; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors / Q000378:metabolism
D007527:Isoenzymes; D011285:Pregnenolone Carbonitrile; D002701:Chloramphenicol; D003101:Collodion; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C029009:steroid hormone 7-alpha-hydroxylase