Heterogeneity of hepatic microsomal UDP-glucuronosyltransferase(s) activities: a new kinetic approach for the study of induction and specificity.
UDP-glucuronosyltransferase (UGT) activities have been described as heterogeneous, i.e. supported by a family of isoenzymes, each of them being capable of conjugating a given chemical family of aglycons, including steroids, coumarines, and phenols. Some of these isoenzymes are specifically induced by xenobiotics. In order to discriminate between the different isoenzymes, we propose a new in situ approach that combines induction (gene regulation) and catalytic activities (specificity). The characterization of one isoenzyme is obtained by (i) increasing its amount by specific inductive stimulation and (ii) studying simultaneously the glucuronidation kinetics of a series of alternative substrates. Provided the substrates are of similar structure, a linear relationship can be established between their glucuronidation rates before versus after induction. We developed a simple mathematical model to analyze the kinetic behaviors observed. With this method, it is possible to know (i) the exact extent of induction of a given isoenzyme by a given inducer (induction factor, n) and (ii) its strict specificity. This in situ methodology is complementary to isolated protein or cDNAs, for the characterization of the real in situ substrate specificity of differentially regulated UGT isoenzymes.
Boutin
J A
JA
Institut de Recherches SERVIER, Suresnes, France.
Antoine
B
B
Siest
G
G
D000818:Animals; D004790:Enzyme Induction / Q000187:drug effects; D014453:Glucuronosyltransferase / Q000096:biosynthesis / Q000378:metabolism*; D006168:Guinea Pigs; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000096:biosynthesis / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D008954:Models, Biological; D010634:Phenobarbital / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013379:Substrate Specificity
D007527:Isoenzymes; D014453:Glucuronosyltransferase; D010634:Phenobarbital