CHARACTERIZATION OF A CLONED HUMAN DIHYDROTESTOSTERONE ANDROSTANEDIOLUDP-GLUCURONOSYLTRANSFERASE AND ITS COMPARISON TO OTHER STEROID ISOFORMS

A human cDNA, UDPGTh-3, encoding a otestosterone/5alpha-androstane-3al pha,17beta-diol UDP-glucuronosyltransferase (transferase) has been iso lated and characterized. The nucleotide sequence of UDPGTh-3 encodes a 530 amino acid protein with a typical membrane insertion-signal pepti de, a membrane-anchoring domain, and three potential asparagine-linked glycosylation sites. Alignment shows that this encoded isozyme is 96% identical to an apparent estriol-metabolizing isoform, HLUG4 [Coffman , B. L., et al., (1990) Arch. Biochem. Biophys. 281, 170-175]. The udp gth-3 isozyme is 78% identical to two other steroid isoforms, HLUG25 ( udpgth-1) [Jackson, M. R., et al. (1987) Biochem. J. 242, 581-588; Rit ter, J. K., et al. (1992) Biochemistry 31, 3409-3414] and udpgth-2 [Ri tter, J. K., et al. (1990) J. Biol. Chem. 265, 7900-7906]. udpgth-2 an d udpgth-1 metabolized parallel substrates (stereospecific estriols, 3 ,4-catechol estrogens, and the bile salt hyodeoxycholate), except that udpgth-2 was 100-fold more effective than udpgth-1. The mRNA encoding udpgth-3 is 2.4 kb in size and is present in liver, prostate, and tes tis; the mRNA encoding udpgth-2 is located in liver and kidney, wherea s that for udpgth-1 is liver-specific. Each of the liver mRNA species encoding udpgth-3, udpgth-2, or udpgth-1 was induced 2.5-3-fold by phe nobarbital treatment of the Erythrocebus patas monkey. In 16 human liv er mRNA samples, the message encoding udpgth-3 was generally uniformly expressed and that for udpgth-1 exhibited wide variations in its leve l, whereas that for udpgth-2 was barely detectable in nine samples and not detectable in the others. Three samples contained no message for either isoform. Substrate turnover by udpgth-3 is ranked as follows: p henolphthalein > 5alpha-androstane-3alpha,17,beta-diol > 5alpha-dihydr otestosterone = 4-hydroxybiphenyl > phenolsulfonphthalein (phenol red) > phenolphthalin. Genes encoding udpgth-3, udpgth-2, and udpgth-1 map ped to human chromosome 4 with genomic DNA from human/mouse and human/ hamster somatic cell hybrids; the genes encoding udpgth-1 and udpgth-2 mapped specifically to band 4q28. udpgth-3 exhibited similar K(m) val ues both for 5alpha-dihydrotestosterone (10 muM) and for its metabolit e, 5alpha-androstane-3alpha-17beta-diol (12.5 muM). Although the role of glucuronidation in the regulation of 5alpha-dihydrotestosterone lev els is not known, the location of the message for this isoform in targ et tissues, testis and prostate, indicates that the isoform is, most l ikely, important in the control of hormonal levels and, thus, in 5alph a-dihydrotestosterone action. Furthermore, a critical role for udpgth- 3 is suggested in light of the absence of its messenger RNA but the pr esence of that for four other transferase isoforms examined in the liv er of a patient with benign prostate hyperplasia, a condition associat ed with depressed glucuronidation of dihydrotestosterone.