Characteristic Fragmentation Behavior of Linear and Cyclic O-Linked Glycopeptides and Their Peptide Skeletons in MALDI-TOF/TOF MS

Understanding characteristic post-source decay (PSD) fragmentation patterns in tandem mass spectrometry (MS/MS) is important for the identification of target molecules. In this study, we explored the characteristic PSD patterns associated with <i>O</i>-linked glycopeptides and their cyclization using the MALDI-TOF/TOF MS analysis of linear and cyclic antifreeze glycoproteins. We performed a comparative analysis of the proton and sodium adduct ions of the peptide backbones of antifreeze glycoproteins, which have a simple repeating sequence, shedding light on the characteristics of the fragmentation of the threonine side chain and that of its cyclized form. Furthermore, the presence or absence of a glycan on the threonine side chain and its substitution with serine caused changes in its fragmentation. These findings are expected to contribute to the prediction of three-dimensional peptide structures and the search for physiologically active <i>O</i>-linked glycopeptides and cyclic (glyco)peptides.