Cloning and Expression of a Novel, Tissue Specifically Expressed Member of the UDP-GalNAc:Polypeptide N-Acetylgalactosaminyltransferase Family

• Published in: The Journal of biological chemistry Vol. 273; no. 42; pp. 27749 - 27754
• Main Authors: Hagen, Kelly G. Ten, Hagen, Fred K., Balys, Marlene M., Beres, Thomas M., Van Wuyckhuyse, Brian, Tabak, Lawrence A.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 16.10.1998; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; Conserved Sequence; DNA, Complementary - genetics; Gene Library; Glycosylation; Isoenzymes - biosynthesis; Isoenzymes - genetics; Molecular Sequence Data; Multigene Family; N-Acetylgalactosaminyltransferases - biosynthesis; N-Acetylgalactosaminyltransferases - genetics; Polypeptide N-acetylgalactosaminyltransferase; Protein Processing, Post-Translational; Rats; Recombinant Proteins - biosynthesis; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Sublingual Gland - enzymology; Tissue Distribution
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.42.27749

We report the cloning and expression of the fifth member of the mammalian UDP-GalNAc:polypeptideN-acetylgalactosaminyltransferase (ppGaNTase) family. Degenerate polymerase chain reaction amplification and hybridization screening of a rat sublingual gland (RSLG) cDNA library were used to identify a novel isoform termed ppGaNTase-T5. Conceptual translation of the cDNA reveals a uniquely long stem region not observed for other members of this enzyme family. Recombinant proteins expressed transiently in COS7 cells displayed transferase activity in vitro. Relative activity and substrate preferences of ppGaNTase-T5 were compared with previously identified isoforms (ppGaNTase-T1, -T3, and -T4); ppGaNTase-T5 and -T4 glycosylated a restricted subset of peptides whereas ppGaNTase-T1 and -T3 glycosylated a broader range of substrates. Northern blot analysis revealed that ppGaNTase-T5 is expressed in a highly tissue-specific manner; abundant expression was seen in the RSLG, with lesser amounts of message in the stomach, small intestine, and colon. Therefore, the pattern of expression of ppGaNTase-T5 is the most restricted of all isoforms examined thus far. The identification of this novel isoform underscores the diversity and complexity of the family of genes controllingO-linked glycosylation.