ISOLATION AND CHARACTERIZATION OF UGT2B15(Y-85) - A UDP-GLUCURONOSYLTRANSFERASE ENCODED BY A POLYMORPHIC GENE

Genetic polymorphisms occur in many of the drug metabolizing enzymes, However, the effect of polymorphisms in the genes encoding phase II dr ug metabolizing UDP-glucuronosyltransferases is still undescribed, des pite the many reported cases of variations in glucuronidation activiti es. Characterization of the UGT2B15(Y-85) cDNA, which was isolated fro m human prostate and LNCaP cell cDNA libraries, revealed 20 nucleotide differences between UGT2B15(Y-85) and the previously characterized UG T2B15 protein UGT2B15(D-85), However only one of the two variations in the coding region leads to an amino acid change from aspartic acid to a tyrosine residue at position 85, The genomic DNA of 27 subjects wer e analysed by direct sequencing of polymerase chain reaction (PCR) pro ducts and demonstrated that UGT2B15(D-85) and UGT2B15(Y-85) are encode d by variant alleles prevalent in the Caucasian population, Expression of UGT2B15(D-85) and UGT2B15(Y-85) in HK293 cells demonstrated simila r substrate specificities, Of the 65 potential substrates tested for a ctivity, the proteins were active on phenolic compounds, coumarins, fl avonoids, drugs and steroid hormones, Both proteins displayed similar K-m values of 2.2 and 2.4 mu M for androstane-3 alpha,17 beta-diol and dihydrotestosterone, respectively. However, results suggest that UGT2 B15(Y-85) has a higher V-max than UGT2B15(D-85), Specific reverse tran scriptase (RT)-PCR analysis revealed expression of the UGT2B15 gene in a wide range of extrahepatic tissues including the human liver, kidne y, testis, mammary gland, placenta, adipose, shin, uterus, prostate an d lung, The wide expression of UGT2B15 in many tissues indicates that it is a major glucuronidation enzyme in humans.