Glycosylation sites of influenza viral glycoproteins Tryptic glycopeptides from the A/WSN (H 0N 1) hemagglutinin glycoprotein

The glycosylation sites of the hemagglutinin glycoprotein of influenza A/WSN virus were investigated by analysis of tryptic glycopeptides separated by ion exchange chromatography and gel filtration. Five major classes of glycosylated tryptic peptides were obtained from the HA 1 subunit, and a single major glycosylated peptide was obtained from the HA 2 subunit. Amino acid sequence analyses indicated that three of the tryptic glycopeptides were each obtained from a distinct glycosylation site on the HA glycoprotein molecule. The oligosaccharides linked to each glycopeptide class were characterized by radiolabeling with various sugar precursors, gel filtration analysis of Pronase digests, and determination of their susceptibility to cleavage by endoglycosidase H. The results indicate that complex oligosaccharides are present in three of the tryptic glycopeptides obtained from HA 1 as well as the glycopeptide obtained from HA 2. One of the tryptic glycopeptides of HA 1 contains a typical high-mannose oligosaccharide, whereas another HA, glycopeptide contains oligosaccharides that have characteristics intermediate between those of typical complex and high-mannose chains. The various tryptic glycopeptides also exhibit differences in extent of sulfation, and nonsulfated and nonsulfated forms of the same glycopeptides appear to be separable by ion exchange chromatography. Affinity chromatography on immobilized plant lectins indicated that each tryptic glycopeptide contained oligosaccharides that were heterogeneous in their exact structures.