Exploiting differential dissociation chemistries of O-linked glycopeptide ions for the localization of mucin-type protein glycosylation

Richard R. Seipert, Eric D. Dodds, Carlito B Lebrilla

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

From a glycoproteomic perspective, the unambiguous localization of O-linked oligosaccharide attachment sites is fraught with analytical obstacles. Because no consensus protein sequence exists for O-glycosylation, there is potential for glycan attachment at numerous serine and threonine residues of a given protein. The well-established tendency for O-glycan attachment to occur within serine and threonine rich domains adds further complication to site-specific assignment of mucin-type glycosylation. In addition to the complexities contributed by the polypeptide chain, the O-linked carbohydrate modifications themselves are exceedingly diverse in both compositional and structural terms. This work is aimed at contributing an improved fundamental understanding of the chemistry that dictates dissociation of O-glycopeptide ions during tandem mass spectrometry (MS/MS). Infrared multiphoton dissociation (IRMPD) has been applied to an assortment of O-linked glycopeptide ions encompassing various compositions and charge states. Protonated O-glycopeptides were found to undergo a combination of glycosidic bond cleavage (complete coverage) and peptide bond cleavage (partial coverage). In contrast to previous observations of N-linked glycopeptide dissociation, the sodiated O-glycopeptides did not yield significantly different information as compared to the corresponding protonated ions. IRMPD of deprotonated O-glycosylated peptides provided informative side chain losses from nonglycosylated serine and threonine residues, which indirectly implicated sites of glycan attachment. In this manner, the combination of positive mode and negative mode MS/MS was found to provide conclusive assignment of O-glycosites.

Original languageEnglish (US)
Pages (from-to)493-501
Number of pages9
JournalJournal of Proteome Research
Volume8
Issue number2
DOIs
StatePublished - Feb 2009

Fingerprint

Glycosylation
Glycopeptides
Mucins
Ions
Threonine
Serine
Polysaccharides
Proteins
Peptides
Infrared radiation
Consensus Sequence
Tandem Mass Spectrometry
Oligosaccharides
Mass spectrometry
Carbohydrates
Chemical analysis

Keywords

  • ESI
  • FT-ICR MS
  • Glycopeptides
  • Glycoproteomics
  • IRMPD
  • MS/MS
  • O-linked glycosylation
  • Protein glycosylation
  • Site-specific glycosylation analysis

ASJC Scopus subject areas

  • Biochemistry
  • Chemistry(all)

Cite this

Exploiting differential dissociation chemistries of O-linked glycopeptide ions for the localization of mucin-type protein glycosylation. / Seipert, Richard R.; Dodds, Eric D.; Lebrilla, Carlito B.

In: Journal of Proteome Research, Vol. 8, No. 2, 02.2009, p. 493-501.

Research output: Contribution to journalArticle

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