2 HUMAN LIVER CDNAS ENCODE UDP-GLUCURONOSYLTRANSFERASES WITH 2 LOG DIFFERENCES IN ACTIVITY TOWARD PARALLEL SUBSTRATES INCLUDING HYODEOXYCHOLIC ACID AND CERTAIN ESTROGEN DERIVATIVES

Two human liver UDP-glucuronosyltransferase cDNA clones, HLUG25 [Jackson, M. R., et al. (1987) Biochem. J. 242, 581-588] and UDPGT(h)-2 [Ritter, J. K., et al. (1990) J. Biol. Chem. 266, 7900-7906] have previously been shown to encode isozymes active in the glucuronidation of hyodeoxycholic acid (HDCA) and certain estrogen derivatives (estriols and 3,4-catechol estrogens), respectively. Here we report that the UDPGT(h)-2-encoded isoform (udpgt(h)-2) and the HLUG25-encoded isoform (udpgt(h)-1) have parallel aglycon specificities. Following expression in COS-1 cells, each isoform metabolized three types of dihydroxy- or trihydroxy-substituted ring structures, including the 3,4-catechol estrogen (4-hydroxyestrone), estriol and 17-epiestriol, and HDCA, but the udpgt(h)-2 isozyme is 100-fold more efficient than udpgt(h)-1. udPgt(h)-1 and udpgt(h)-2 are 86% identical overall (76 differences out of 528 amino acids), including 55 differences in the first 300 amino acids of the amino terminus, a domain which confers isoform substrate specificity. The data indicate that a high level of conservation in the amino terminus is not required for the preservation of substrate selectivity. Analysis of glucuronidation activity encoded by UDPGT(h)-1/UDPGT(h)-2 chimeric cDNAs constructed at their common restriction sites, SacI (codon 297), NcoI (codon 385), and HhaI (codon 469), showed that nine amino acids between residues 385 and 469 are important for catalytic efficiency, suggesting that this region represents a domain which is critical for catalysis but distinct from that responsible for aglycon selection. In parallel with the existence of liver and kidney microsomal HDCA glucuronosyl transferase activity, mRNA coding for udpgt(h)-2 is expressed in liver and kidney, whereas that for udpgt(h)-1 is expressed only in the liver. These data indicate that udpgt(h)-2 is a primary isoform responsible for the detoxification of the bile salt intermediate as well as the active estrogen in termediates.