Hydroxylations in biosynthesis of bile acids. Cytochrome P-450 LM4 and 12alpha-hydroxylation of 5beta-cholestane-3alpha, 7alpha-diol.
12alpha-Hydroxylation of 5beta-cholestane-3alpha, 7alpha-diol was studied in reconstituted systems consisting of electrophoretically homogeneous cytochrome P-450 LM4 fractions and NADPH-cytochrome P-450 reductase from rabbit liver microsomes. Cytochrome P-450 LM4 fractions were prepared from untreated, phenobarbital-treated, beta-naphthoflavone-treated and starved rabbits. The purified cytochromes catalyzed 12alpha-hydroxylation more efficiently than the corresponding microsomes. In the reconstituted systems, carbon monoxide inhibited 12alpha-hydroxylation by 50-80%. The rate of 12alpha-hydroxylation was three to four times higher with cytochrome P-450 LM4 fractions from starved rabbits than with cytochrome P-450 LM4 fractions from untreated, phenobarbital-treated and beta-naphthoflavone-treated animals. Amino acid analyses, peptide mapping experiments as well as absorption spectral and circular dichroism spectral analyses revealed physical differences between cytochrome P-450 LM4 fractions from starved animals and preparations from phenobarbital-treated animals. The results indicate the presence of a cytochrome P-450 species in the cytochrome P-450 LM4 fraction specific for 12alpha-hydroxylation.
D000818:Animals; D001571:Benzoflavones / Q000378:metabolism* / Q000494:pharmacology; D001647:Bile Acids and Salts / Q000096:biosynthesis*; D002384:Catalysis; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D005419:Flavonoids / Q000378:metabolism*; D006900:Hydroxylation; D008862:Microsomes, Liver / Q000201:enzymology*; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism*; D010634:Phenobarbital / Q000494:pharmacology; D011817:Rabbits; D013217:Starvation / Q000378:metabolism; D019324:beta-Naphthoflavone
D001571:Benzoflavones; D001647:Bile Acids and Salts; D005419:Flavonoids; D019324:beta-Naphthoflavone; D003577:Cytochrome P-450 Enzyme System; D009251:NADPH-Ferrihemoprotein Reductase; D010634:Phenobarbital