CLONING AND EXPRESSION OF HUMAN LIVER UDP-GLUCURONOSYLTRANSFERASE CDNA, UDPCTH2

The human liver cDNA clone UDPGTh2, encoding a liver UDP-glucuronosylt ransferase (UDPGT) was isolated from a lambda gt 11 cDNA library by hy bridization to mouse transferase cDNA clone, UDPGTm1. UDPGTh2 encoded a 529 amino acid protein with an amino terminus membrane-insertion sig nal peptide and a carboxyl terminus membrane-spanning region. There we re three potential asparagine-linked glycosylation sites at residues 6 7, 68, and 315. In order to obtain UDPGTh2 protein encoded from cloned human liver UDP-glucuronosyltransferase cDNA, the clone was inserted into the pSVL vector (pUDPGTh2) and expressed in COS 1 cells. The pres ence of a transferase with Mr similar to 52,000 in transfected cells c ultured in the presence of [S-35]methionine was shown by immunocomplex ed products with goat antimouse transferase IgG and protein A-Sepharos e and analysis by sodium dodecyl sulfate-polyacrylamide gel electropho resis and autoradiography. The expressed UDPGT was a glycoprotein as i ndicated by electrophoretic mobility shift in Mr similar to 3,000-4,00 0 when expressed in the presence of tunicamycin. The extent of glycosy lation was difficult to assess, although one could assume that glycosy l structures incorporated at the level of endoplasmic reticulum were a lways the core oligosaccharides. Thus, it is likely that at least two moieties inserted can account for the shift of Mr similar to 3,000-4,0 00. This study demonstrates the cDNA and deduced amino acid sequence o f human liver UDP-glucuronosyltransferase cDNA, UDPGTh2.