Demonstration of a Novel UDPGalNAc:GlcNAc ß1-4 N-Acetylgalactosaminyltransferase in Extracts of Schistosoma mansoni

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It was recently demonstrated that complex-type N-linked oligosaccharides in glycoproteins synthesized by Schistosoma mansoni contain the unusual terminal sequence GalNAc beta 1-4GlcNAc beta 1-2Man alpha 1-2R. This structure suggests that the parasite might contain a novel glycosyltransferase that can add GalNAc to terminal GlcNAc residues in N-linked oligosaccharides. This report describes an assay for this enzyme, designated UDPGalNAc:GlcNAc beta 1-4 N-acetylgalactosaminyltransferase (beta 1-4GalNAcT), and demonstrates the presence of the enzyme activity in schistosome extracts. As an acceptor for the beta 1-4GalNAcT, we prepared from human fibrinogen a truncated biantennary glycopeptide that contained terminal GlcNAc residues. When this acceptor was incubated with schistosome extracts in the presence of UDP-[3H]GalNAc, Mn2+, and detergent, [3H]GalNAc was transferred to the glycopeptide acceptor. Approximately 75% of the radioactivity in the product isolated by lectin affinity chromatography was recovered as [3H]GalNAc following hydrolysis; likewise, a majority of the isolated product was bound by immobilized Wisteria floribunda agglutinin, a lectin that binds to schistosome-derived oligosaccharides containing terminal beta 1-4-linked GalNAc residues. The activity of the beta 1-4GalNAcT in schistosome extracts was dependent on time, protein, and UDPGalNAc.