S1.41 Human UDP-Galnac: PolypeptideN-acetylgalactosaminyl transferase: Identification of two distinct threonine transferase activities

Abstract

Purification of UDP-GalNAc: polypeptide N-acetylgalactosaminyl transferase from human placenta and ovine and porcine submaxillary glands by affinity chromatography on a defined synthetic peptide containing multiple threonine acceptor substrate sites resulted in separation of at least two distinct threonine transferase activities. A panel of synthetic peptides was utilized as acceptor substrates on transferase preparations before and after affinity purification on the synthetic peptide in order to evaluate the substrate specificities. Only a fraction of the transferase activity available was bound to the affinity column evaluated by a number of acceptor substrate peptides including the peptide used for the affinity chromatography column. Interestingly, one peptide containing a single threonine glycosylation site was not glycosylated by the affinity purified transferase and the transferase activity to this peptide passed quantatively through the affinity column. Furthermore, kinetic analysis of crude and purified transferase preparations revealed significant differences between the two transferase preparations. The results suggest that more than one threonine transferase activity exist, and it is likely to be due to different structural proteins.