Characterization of the high mannose asparagine-linked oligosaccharides synthesized by microfilariae of Dirofilaria immitis

ABSTRACT

this report describes the structures of high-mannose-type N-linked oligosaccharides in glycoproteins synthesized by the microfilariae of Dirofioria immitis. Microfilariae of D. immitis were incubated in vitro in media contaning 2-[(3)H] mannose to allow metabolic radiolabeling of the oligosaccharide moieties of newly synthesized glycoproteins. Glycopeptides were prepared from the radiolabeled glycoproteins by digestion with pronase and fractionation by chromatography on concanavalin A-Sepharose. Thirty eight percent of 2-[(3)H] mannose incorporated into the microfilariae of D. immitis glycopeptides was recovered in high mannose-type asparagine-linked oligosaccharides whech were bound to the immobilized lectin. Upon treatment of 2-[(3)H] mannose labeled glycopeptides with endo-beta-N-acetylglucosaminidase H, the high mannose-type chains were released and their structures were determined by high performance liquid chromatography and exoglycosidase digestion. The major species of high mannose-type chains synthesized by microfilariae of D. immitis have the composition Man(5) GlcNAc(2), Man(6) ClcNAc(2), Man(8) GlcNA(2), and Man(8) GlcNAc(2). Structural analyses indicate that these oligosaccharides are similar to high mannose-type chains synthesized by vertebrates.