The cDNA sequence and expression of a variant 17 beta-hydroxysteroid UDP-glucuronosyltransferase.
The cDNA encoding a member of a family of steroid UDP-glucuronosyltransferases has been cloned and sequenced. The 1871-base pair (bp) cDNA, designated UDPGTr-5, contains an open reading frame of 1590 bp flanked by 43 and 238 bp of 5'- and 3'-untranslated regions, respectively. The 530-amino acid protein encoded by the open reading frame is 93% and 84% similar in sequence to UDPGTr-3 and UDPGTr-4, which are other members of this multigene family. In common with these forms, the encoded protein contains regions characteristic of a signal peptide and a transmembrane-anchoring domain at the carboxyl terminus. Potential asparagine-linked glycosylation sites are not present in the encoded protein. Expression of UDPGTr-5 cDNA in COS cells demonstrated that the encoded enzyme has a subunit size of 50 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The expressed enzyme glucuronidates testosterone and dihydrotestosterone, although its activity towards these two substrates is about 30-fold less than that of UDPGTr-3. This low activity appeared to be a bone fide property of the enzyme and was not due to lower amounts of protein synthesized or increased rates of protein degradation. Northern analysis and cDNA sequencing demonstrated that UDPGTr-5 is encoded by two mRNA species which differed in the lengths of their 3'-untranslated regions. The cDNA to the longer transcript has a 1768-bp 3'-untranslated region which contains a segment that is 80% similar to the rat identifier sequence (ID) family of repetitive DNA elements. These data indicate that UDPGTr-5 is another 17 beta-hydroxysteroid UDP-glucuronosyltransferase that is encoded by two mRNAs transcribed from a single gene.
Mackenzie
P I
PI
Department of Clinical Pharmacology, Flinders Medical Centre, Bedford Park, South Australia.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D015723:Gene Library; D014644:Genetic Variation; D014453:Glucuronosyltransferase / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids; D011061:Poly A / Q000235:genetics; D021382:Protein Sorting Signals / Q000235:genetics; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D021382:Protein Sorting Signals; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D014453:Glucuronosyltransferase