Journal
RAPID COMMUNICATIONS IN MASS SPECTROMETRY
Volume 20, Issue 23, Pages 3557-3565Publisher
WILEY
DOI: 10.1002/rcm.2761
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Mass spectrometric analyses of various N-glycans binding to proteins and peptides are highly desirable for elucidating their biological roles. An approach based on collision-induced dissociation (CID) MSn spectra acquired by electrospray ionization linear ion trap time-of-flight mass spectrometry (ESI-LIT-TOFMS) in the positive- and negative-ion modes has been proposed as a direct method of assigning N-glycans without releasing them from N-glycopeptides. In the positive-ion mode of this approach, the MS2 spectrum of N-glycopeptide was acquired so that a glycoside-bond cleavage occurs in the chitobiose residue (i.e., GlcNAc beta 1-4GlcNAc, GlcNAc: N-acetylglucosamine) attached to asparagine (N), and two charges on the [M+H+Na](2+) precursor ion are shared with both of the resulting fragments. These fragments are sodiated B-n-type fragment ions of oligosaccharide (N-glycan) and a protonated peptide ion retaining one GlcNAc residue on the asparagine (N) residue. The structure of N-glycan was assigned by comparing MS3 spectra derived from both the sodiated Bn-type fragment ions of N-glycopeptide and the PA (2-aminopyridine) N-glycan standard (i.e., MSn spectral matching). In a similar manner, the structural assignment of sialylated N-glycan was performed by employing the negative-ion CID MSn spectra of deprotonated B-n-type fragment ions of N-glycopeptide and the PA N-glycan standard. The efficacy of this approach was tested with chicken egg yolk glycopeptides with a neutral and a sialylated N-glycan, and human serum IgG glycopeptides with neutral N-glycan isomers. These results suggest that the approach based on MSn spectral matching is useful for the direct and simple structural assignment of neutral and sialylated N-glycans of glycopeptides. Copyright (c) 2006 John Wiley & Sons, Ltd.
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