cDNA Cloning and Expression of a Novel Human UDP- -acetyl-α-D-galactosamine

Abstract

The glycosylation of serine and threonine residues during mucin-type O-linked protein glycosylation is carried out by a family of UDP-GalNAc:polypeptide N- acetylgalactosaminyltransferases (GalNAc-transferase). Previously two members, GalNAc-T1 and -T2, have been isolated and the genes cloned and characterized. Here we report the cDNA cloning and expression of a novel GalNAc-transferase termed GalNAc-T3. The gene was isolated and cloned based on the identification of a Gal- NAc-transferase motif (61 amino acids) that is shared between GalNAc-T1 and -T2 as well as a homologous Caenorhabditis elegans gene. The cDNA sequence has a 633-amino acid coding region indicating a protein of 72.5 kDa with a type II domain structure. The overall amino acid sequence similarity with GalNAc-T1 and -T2 is ap- proximately 45%; 12 cysteine residues that are shared between GalNAc-T1 and -T2 are also found in GalNAc- T3. GalNAc-T3 was expressed as a soluble protein with- out the hydrophobic transmembrane domain in insect cells using a Baculo-virus vector, and the expressed Gal- NAc-transferase activity showed substrate specificity different from that previously reported for GalNAc-T1 and -T2. Northern analysis of human organs revealed a very restricted expression pattern of GalNAc-T3