Resistance to deglycosylation by ammonia of IgA1 O-glycopeptides: implications for the β-elimination of O-glycans linked to serine and threonine

Pools of O-glycopeptides (and their deglycosylated analogues) derived from trypsin-digested normal human serum IgA1 have been treated with ammonia under conditions reported to result in complete liberation of O-glycans linked to serine and threonine residues in glycopeptides and glycoproteins. MALDI-TOF MS analysis has revealed that only one of the six glycosylated sites is susceptible to β-elimination under these conditions. It is likely that resistance to β-elimination is due to very close proximity of proline to the glycosylated serine or threonine residues. Preliminary results using 0.1 M NaOH (instead of ammonia) to perform β-elimination indicated that there was also selective de-O-glycosylation with this reagent, however, these results were complicated by the concomitant hydrolysis of the peptide bonds. These findings may have implications for similarly O-glycosylated peptides and proteins and possibly for other chemical methods that are used to carry out β-eliminations of O-glycans.