Structural gene products of the Ah locus. Evidence for many unique P-450-mediated monooxygenase activities reconstituted from 3-methylcholanthrene-treated C57BL/6N mouse liver microsomes.
Cytochrome P-450 from cholate-solubilized liver microsomes prepared from 3-methylcholanthrene-treated genetically "responsive" C57BL/6N mice (Ahb/Ahb) was partially purified by aminooctyl-Sepharose 4B column chromatography with an elution buffer that included Emulgen 911, followed by hydroxylapatite column chromatography. The P-450 was separated into 16 fractions: 15 fractions principally associated with the Ahb allele and induction by 3-methylcholanthrene and 1 fraction not associated with the Ahb allele, i.e. predominantly constitutive form(s) of P-450. The metabolism of benzo[a]pyrene, 7-ethoxycoumarin, biphenyl, ethoxyresorufin, acetanilide, 2-acetylaminofluorene, phenacetin, estradiol-17 beta, and testosterone was investigated. With these nine substrates, we examined the rates of formation of 17 products, i.e. 17 P-450-mediated reconstituted monooxygenase "'activities," in each of the 16 fractions. Differing reductase/cytochrome and lipid/cytochrome requirements were observed for each activity; the reasons for these empirical differences are not known. By two-factor analysis of variance our data for 17 catalytic activities can be explained by a minimum of 19 unique groups of monooxygenase activities: 12 induced by 3-methylcholanthrene and 7 control (endogenous). At least one of these distinctly different 3-methylcholanthrene-induced groups and at least one of these constitutive groups is associated with a marked blue spectral shift (approximately 2.0 nm) in the Soret peak of the reduced hemoprotein.CO complex, suggesting multiple forms of inducible and control "P-448." Almost every reconstituted monooxygenase activity therefore appears to be unique and probably represents the aggregate activity from numerous forms of P-450.
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000302:isolation & purification; D005796:Genes / Q000187:drug effects*; D008748:Methylcholanthrene / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000235:genetics* / Q000302:isolation & purification; D013379:Substrate Specificity
D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases