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PubMed · 1718996

Molecular basis for a functionally unique cytochrome P450IIB1 variant.

K M Kedzie·C A Balfour·G Y Escobar·S W Grimm·Y A He·D J Pepperl·J W Regan·J C Stevens·J R Halpert
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Abstract

Liver microsomes from phenobarbital-treated rats of four inbred strains expressing distinct allelic variants of cytochrome P450IIB1 were analyzed. The Wistar Munich (WM) strain exhibited 5- to 10-fold lower androstenedione 16 beta-hydroxylase activity (a specific P450IIB1 marker) than the Lewis, Wistar Kyoto, and Wistar Furth strains. The androstenedione 16 beta-hydroxylase in the WM liver microsomes was refractory to inactivation by N-(2-p-nitrophenethyl)chlorofluoroacetamide, a selective P450IIB1 inactivator in the other three strains. Purified P450IIB1-WM was insensitive to the inactivator and exhibited 5-fold lower androstenedione 16 beta-hydroxylase, testosterone 16-hydroxylase, and 7-ethoxycoumarin deethylase activities but the same benzphetamine demethylase activity and slightly higher androstenedione 16 alpha-hydroxylase activity than a P450IIB1 purified from outbred Sprague-Dawley rats, which appears to correspond to the form in Lewis rats. The stereoselectivity of androstenedione 16-hydroxylation catalyzed by P450IIB1-WM (16 beta-OH:16 alpha-OH = 1.4) is thus distinct from that (16 beta-OH:16 alpha-OH = 12-15) of other P450IIB1 preparations described. A cDNA encoding P450IIB1-WM was cloned and sequenced, revealing a single amino acid substitution (Gly-478----Ala) compared with the published sequence (Fujii-Kuriyama, Y., Mizukami, Y., Kawajiri, K., Sogawa, K., and Muramatsu, M. (1982) Proc. Natl. Acad. Sci. U. S. A. 79, 2793-2797). Heterologous expression of P450IIB1 and P450IIB1-WM confirmed the striking difference in androstenedione metabolite profiles, strongly implicating the involvement of Ala-478 in defining the distinctive catalytic properties of P450IIB1-WM.

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AllelesAnimalsCloning, MolecularCytochrome P-450 CYP2B1Cytochrome P-450 Enzyme SystemDNAGenetic VariationKineticsLiverMaleMicrosomes, LiverOxidoreductasesPhenobarbitalPlasmidsRNARatsRats, Inbred LewRats, Inbred StrainsRats, Inbred WFRats, Inbred WKYRecombinant ProteinsSequence Homology, Nucleic AcidSpecies SpecificitySteroid 16-alpha-HydroxylaseSubstrate SpecificityComparative StudyJournal ArticleResearch Support, Non-U.S. Gov'tResearch Support, U.S. Gov't, P.H.S.

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K M Kedzie, C A Balfour, G Y Escobar, S W Grimm, Y A He, D J Pepperl, J W Regan, J C Stevens, J R Halpert. 1991-11-25. Molecular basis for a functionally unique cytochrome P450IIB1 variant.. https://pubmed.ncbi.nlm.nih.gov/1718996/

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