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Multistep Fractionation and Mass Spectrometry Reveal Zwitterionic and Anionic Modifications of the N- and O-glycans of a Marine Snail

Journal article
Authors B. Eckmair
Chunsheng S. Jin
D. Abed-Navandi
K. Paschinger
Published in Molecular & Cellular Proteomics
Volume 15
Issue 2
Pages 573-597
ISSN 1535-9476
Publication year 2016
Published at Institute of Biomedicine
Pages 573-597
Language en
Links dx.doi.org/10.1074/mcp.M115.051573
Keywords linked carbohydrate chains, performance liquid-chromatography, oyster, crassostrea-virginica, 2-dimensional sugar map, lymnaea-stagnalis, schistosoma-mansoni, acetylglucosamine residue, dictyostelium-discoideum, caenorhabditis-elegans, biomphalaria-glabrata, Biochemistry & Molecular Biology, iences, v88, p509, hachter h, 1982, advances in experimental medicine and biology, v144, p3, hneider p, 1995, lipid modifications of proteins, v250, p614
Subject categories Cell and Molecular Biology

Abstract

Various studies in the past have revealed that molluscs can produce a wide range of rather complex N-glycan structures, which vary from those occurring in other invertebrate animals; particularly methylated glycans have been found in gastropods, and there are some reports of anionic glycans in bivalves. Due to the high variability in terms of previously described structures and methodologies, it is a major challenge to establish glycomic workflows that yield the maximum amount of detailed structural information from relatively low quantities of sample. In this study, we apply differential release with peptide: N-glycosidases F and A followed by solid-phase extraction on graphitized carbon and reversed-phase materials to examine the glycome of Volvarina rubella (C. B. Adams, 1845), a margin snail of the clade Neogastropoda. The resulting four pools of N-glycans were fractionated on a fused core RP-HPLC column and subject to MALDI-TOF MS and MS/MS in conjunction with chemical and enzymatic treatments. In addition, selected N-glycan fractions, as well as O-glycans released by beta-elimination, were analyzed by porous graphitized carbon-LC-MS and MSn. This comprehensive approach enabled us to determine a number of novel modifications of protein-linked glycans, including N-methyl-2-aminoethylphosphonate on mannose and N-acetylhexosamine residues, core beta 1,3-linked mannose, zwitterionic moieties on core Gal beta 1,4Fuc motifs, additional mannose residues on oligomannosidic glycans, and bisubstituted antennal fucose; furthermore, typical invertebrate N-glycans with sulfate and core fucose residues are present in this gastropod.

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