title: Identification and Validation of Atypical N-Glycosylation Sites.
abstract: It is well-known that N-linked glycans usually attach to asparagine residues in the N-X-S/T motifs of proteins. However, accumulating evidence indicates that N-glycosylation could also possibly occur at other atypical motifs. In this study, we tried to identify atypical N-glycosylation sites using our recently developed solid-phase extraction of the N-linked Glycans And Glycosite-containing peptides (NGAG) method. Peptides with deamidation sites at asparagine residues but lacking a typical asparagine-X-serine/threonine sequons (N-X-S/T, X is any amino acid except proline) motif were identified from deglycosylated peptide data as potentially atypical glycosite-containing peptides. These atypical glycosites were verified by the presence of glycans on their intact glycopeptides and further confirmed by specific inhibition of cells with an N-linked glycosylation inhibitor, tunicamycin. From this study, two atypical N-linked glycosylation sites with N-X-C and N-X-V motifs were identified and validated from an ovarian cancer cell line (OVCAR-3).
authors.affiliation: Department of Pathology, Johns Hopkins University , 400 North Broadway, Smith Building, Room 4001, Baltimore, Maryland 21287, United States.
authors.forename: Shisheng
authors.initials: S
authors.lastname: Sun
authors.affiliation: Department of Pathology, Johns Hopkins University , 400 North Broadway, Smith Building, Room 4001, Baltimore, Maryland 21287, United States.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D001216:Asparagine / Q000737:chemistry*; D001665:Binding Sites; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D006031:Glycosylation; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000737:chemistry*; D015203:Reproducibility of Results; D052616:Solid Phase Extraction
chemical_list: D011134:Polysaccharides; D001216:Asparagine