Specific rearrangement reactions of acetylated lysine containing peptide b n ( n  = 4–7) ion series

Characterization of ε‐N ‐acetylated lysine containing peptides, one of the most prominent post‐translational modifications of proteins, is an important goal for tandem mass spectrometry experiments. A systematic study for the fragmentation reactions of b ions derived from ε‐N ‐acetyllysine containing model octapeptides (K Ac YAGFLVG and YAK Ac GFLVG) has been examined in detail. Collision‐induced dissociation (CID) mass spectra of b n ( n = 4–7) fragments of ε‐N ‐acetylated lysine containing peptides are compared with those of N ‐terminal acetylated and doubly acetylated (both ε‐N and N ‐terminal) peptides, as well as acetyl‐free peptides. Both direct and nondirect fragments are observed for acetyl‐free and singly acetylated ( ε‐N or N ‐terminal) peptides. In the case of ε‐N ‐acetylated lysine containing peptides, however, specific fragment ions ( m / z 309, 456, 569 and 668) are observed in CID mass spectra of b n ( n = 4–7) ions. The CID mass spectra of these four ions are shown to be identical to those of selected protonated C ‐terminal amidated peptides. On this basis, a new type of rearrangement chemistry is proposed to account for the formation of these fragment ions, which are specific for ε‐N ‐acetylated lysine containing peptides. Consistent with the observation of nondirect fragments, it is proposed that the b ions undergo head‐to‐tail macrocyclization followed by ring opening. The proposed reaction pathway assumes that b n ( n = 4–7) of ε‐N ‐acetylated lysine containing peptides has a tendency to place the K Ac residue at the C ‐terminal position after macrocyclization/reopening mechanism. Then, following the loss of CO, it is proposed that the marker ions are the result of the loss of an acetyllysine imine as a neutral fragment. Copyright © 2014 John Wiley & Sons, Ltd.