title: The neutral N-linked glycans of the ustilaginomycete yeast Sympodiomycopsis paphiopedili.
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 Man3GlcNAc2-PA to Man9GlcNAc2-PA in length. The most abundant neutral N-linked glycan structure in this species was found to be the Manα1-2Manα1-6(Manα1-3)Manα1-6(Manα1-2Manα1-2Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAc (M8A). Moreover, the second and third most abundant neutral N-linked glycan in S. paphiopedili were the Manα1-2Manα1-6(Manα1-2Manα1-3)Manα1-6(Manα1-2Manα1-2Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAc (M9A) and the Manα1-6(Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAc (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.
authors.affiliation: International Center for Biotechnology, Osaka University, Japan.
authors.forename: Ronilo Jose D
authors.initials: RJD
authors.lastname: Flores
authors.affiliation: International Center for Biotechnology, Osaka University, Japan.
authors.forename: Takao
authors.identifier: 0000-0002-5737-2230
authors.initials: T
authors.lastname: Ohashi
authors.affiliation: NITE Biological Resource Center, National Institute of Technology and Evaluation, Chiba, Japan.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kawasaki
authors.affiliation: International Center for Biotechnology, Osaka University, Japan.
authors.forename: Kazuhito
authors.identifier: 0000-0001-5801-2774
authors.initials: K
authors.lastname: Fujiyama
mesh_terms: D001487:Basidiomycota / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D013058:Mass Spectrometry; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000737:chemistry
chemical_list: D011134:Polysaccharides
keywords: N-glycan; Sympodiomycopsis; basidiomycetous yeasts; cell wall