Benzo(a)pyrene hydroxylase from Saccharomyces cerevisiae. Substrate binding, spectral and kinetic data.
Saccharomyces cerevisiae, brewer's yeast, produces a microsomal benzo(a)pyrene hydroxylase when grown at high glucose concentrations of which the haemoprotein, cytochrome P-450 (RH, reduced-flavoprotein:oxygen oxidoreductase (RH-hydroxylating) EC 1.14.14.1) is a component. We report here kinetic data derived from Lineweaver-Burk plots of benzo(a)pyrene hydroxylation. The Michaelis constant was decreased by growth of the yeast in the presence of benzo(a)pyrene showing the induction of a form of the enzyme more specific for this compound. NADPH or cumene hydroperoxide could be used as cofactors by this enzyme, although with different Km and V values for benzo(a)pyrene. A solubilised and a solubilised, immobilised enzyme preparation were capable of benzo(a)pyrene hydroxylation, using cumene hydroperoxide but not NADPH as the cofactor. Benzo(a)pyrene was found to produce a modified type I spectral change with yeast and rat liver microsomes. The interaction of benzo(a)pyrene with cytochrome P-450 was investigated further by means of an equilibrium gel filtration technique. There appeared to be 20 binding sites per mol ofcytochrome P-450 for benz(a)pyrene, in both yeast and rat liver microsomes.
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism*; D001555:Benzene Derivatives / Q000378:metabolism; D001579:Benzopyrene Hydroxylase / Q000378:metabolism*; D001580:Benzopyrenes / Q000378:metabolism; D002850:Chromatography, Gel; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D007700:Kinetics; D008862:Microsomes, Liver / Q000201:enzymology; D009249:NADP / Q000378:metabolism; D010545:Peroxides / Q000378:metabolism; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D012995:Solubility; D013057:Spectrum Analysis
D001555:Benzene Derivatives; D001580:Benzopyrenes; D010545:Peroxides; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D001579:Benzopyrene Hydroxylase; D001189:Aryl Hydrocarbon Hydroxylases; C007164:cumene hydroperoxide