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The neutral N ‐linked glycans of the ustilaginomycete yeast Sympodiomycopsis paphiopedili
Author(s) -
Flores Ronilo Jose D.,
Ohashi Takao,
Kawasaki Hiroko,
Fujiyama Kazuhito
Publication year - 2017
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/yea.3233
Subject(s) - glycan , biology , high performance liquid chromatography , mannose , extraction (chemistry) , chromatography , biochemistry , chemistry , glycoprotein
Abstract Sympodiomycopsis paphiopedili is a basidiomycetous yeast under the subphylum Ustilaginomycotina and is a commensal organism originally isolated from the nectar of a plant species in Japan. In this study, the neutral N ‐linked glycans of S. paphiopedili were prepared and structurally analysed using high‐performance liquid chromatography (HPLC) and mass spectrometry (MS). Glycosidase digestion analyses were also performed to verify certain glycan linkages. HPLC and MS analyses revealed the presence of neutral N ‐linked glycans ranging from Man 3 GlcNAc 2 ‐PA to Man 9 GlcNAc 2 ‐PA in length. The most abundant neutral N ‐linked glycan structure in this species was found to be the Man α 1–2Man α 1–6(Man α 1–3)Man α 1–6(Man α 1–2Man α 1–2Man α 1–3)Man β 1–4GlcNAc β 1–4GlcNAc (M8A). Moreover, the second and third most abundant neutral N ‐linked glycan in S. paphiopedili were the Man α 1–2Man α 1–6(Man α 1–2Man α 1–3)Man α 1–6(Man α 1–2Man α 1–2Man α 1–3)Man β 1–4GlcNAc β 1–4GlcNAc (M9A) and the Man α 1–6(Man α 1–3)Man β 1–4GlcNAc β 1–4GlcNAc (M3B). On the other hand, the effect of the combination of glycoprotein extraction methods (citrate buffer extraction or bead extraction) and the subsequent glycan release methods (hydrazinolysis or PNGase F digestion) on the detection of N ‐linked glycan peaks was also examined for S. paphiopedili and Saccharomyces cerevisiae in order to avoid under‐representation of N ‐linked glycan structures. High mannose and possible hypermannosylated glycan peaks were detected in all method combinations in S. cerevisiae with the citrate buffer extraction–hydrazinolysis method giving the highest peak yields as compared with the other methods. Here we report the first account of the structural analysis of the neutral N ‐linked glycan of S. paphiopedili and the comparison of the effect of combinations of glycoprotein extraction methods and glycan release methods with that of the glycan analysis in S. paphiopedili and S. cerevisiae . Copyright © 2017 John Wiley & Sons, Ltd.