Cytochrome P-450-mediated activation of procarcinogens and promutagens to DNA-damaging products by measuring expression of umu gene in Salmonella typhimurium TA1535/pSK1002.
A simple and sensitive procedure for the determination of cytochrome P-450 (P-450)-mediated activation of chemical procarcinogens and promutagens to DNA-damaging products has been developed using a method measuring the expression of the umu gene in Salmonella typhimurium TA1535/pSK1002, which is based upon the initial procedures as described by Oda et al. [Mutation Res. 147, 219 (1985)]. The chemicals examined were a variety of potent carcinogenic and mutagenic compounds including heterocyclic aromatic amines, aromatic amines, polycyclic aromatic hydrocarbons and aflatoxin B1. These chemicals were incubated with rat liver microsomes or a reconstituted monooxygenase system containing three forms of purified P-450 in the presence of a bacterial tester strain, and the induced-umu gene expression was determined by measuring the beta-galactosidase activity produced by fusion gene in the cells. The activity was increased linearly for at least 2 hr with an initial lag time of 30 min and was dependent on the concentrations of P-450 in the reaction mixture. Thus, the metabolic activation of these compounds by P-450 could be compared on a basis of the specific beta-galactosidase activity/min/nmol P-450. Among three forms of P-450, two isozymes induced by 3-methylcholanthrene were found to be more active in catalyzing the metabolic activation of most of the chemicals examined than a form of P-450 which is induced by phenobarbital. Data also showed that a high spin form of P-450 isolated from 3-methylcholanthrene-treated rats had a profound role in the activation of procarcinogens and promutagens. This conclusion was based on the results of catalytic activities by three forms of P-450 in a reconstituted monooxygenase system, and on the effects of specific antibodies against these P-450s on the reactions catalyzed by liver microsomes.
Shimada
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Nakamura
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D000818:Animals; D002273:Carcinogens / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004247:DNA / Q000187:drug effects*; D004789:Enzyme Activation; D005668:Furylfuramide / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D009153:Mutagens / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D012486:Salmonella typhimurium / Q000235:genetics
D002273:Carcinogens; D007527:Isoenzymes; D009153:Mutagens; D005668:Furylfuramide; D004247:DNA; D003577:Cytochrome P-450 Enzyme System