title: Complementary structural information of positive- and negative-ion MSn spectra of glycopeptides with neutral and sialylated N-glycans.
abstract: Positive- and negative-ion MSn spectra of chicken egg yolk glycopeptides binding a neutral and a sialylated N-glycan were acquired by using electrospray ionization linear ion trap time-of-flight mass spectrometry (ESI-LIT-TOFMS) and collision-induced dissociation (CID) with helium as collision gas. Several characteristic differences were observed between the positive- and negative-ion CID MSn (n = 2, 3) spectra. In the positive-ion MS2 spectra, the peptide moiety was presumably stable, but the neutral N-glycan moiety caused several B-type fragmentations and the sialylated N-glycan almost lost sialic acid(s). In contrast, in the negative-ion MS2 spectra, the peptide moiety caused several side-chain and N-glycan residue (e.g., N-acetylglucosamine (GlcNAc) residue) fragmentations in addition to backbone cleavages, but the N-glycan moieties were relatively stable. The positive-ion MS3 spectra derived from the protonated peptide ion containing a GlcNAc residue (203.1 Da) provided enough information to determine the peptide amino-acid sequence including the glycosylation site, while the negative-ion MS3 spectra derived from the deprotonated peptide containing a 0,2X1-type cross-ring cleavage (83.1 Da) complicated the peptide sequence analysis due to side-chain and 0,2X1 residue related fragmentations. However, for the structural information of the N-glycan moiety of the glycopeptides, the negative-ion CID MS3 spectra derived from the deprotonated 2,4A6-type cross-ring cleavage ion (neutral N-glycan) or the doubly deprotonated B6-type fragment ion (sialylated N-glycan) are more informative than are those of the corresponding positive-ion CID MS3 spectra. Thus, the positive-ion mode of CID is useful for the analyses of peptide amino-acid sequences including the glycosylation site. The negative-ion mode of CID is especially useful for sialylated N-glycan structural analysis. Therefore, in the structural analysis of N-glycopeptides, their roles are complementary.
authors.affiliation: Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo 001-0021, Japan. deguchi@glyco.sci.hokudai.ac.jp
authors.forename: Kisaburo
authors.initials: K
authors.lastname: Deguchi
authors.forename: Hiroki
authors.initials: H
authors.lastname: Ito
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Takegawa
authors.forename: Nagai
authors.initials: N
authors.lastname: Shinji
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Nakagawa
authors.forename: Shin-Ichiro
authors.initials: S
authors.lastname: Nishimura
mesh_terms: D000818:Animals; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D004527:Egg Proteins / Q000737:chemistry; D006020:Glycopeptides / Q000737:chemistry*; D006031:Glycosylation; D015394:Molecular Structure; D011134:Polysaccharides / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D004527:Egg Proteins; D006020:Glycopeptides; D011134:Polysaccharides