Loss of 45 Da from a2 Ions and Preferential Loss of 48 Da from a2 Ions Containing Methionine in Peptide Ion Tandem Mass Spectra
Yamil Simon, Pedatsur Neta, Xiaoyu Yang, Stephen E. Stein
While analyzing tandem mass spectra of tryptic tripeptides, intense unassigned peaks were observed, which correspond to a neutral loss of 45 Da from a2 ions. From exact mass analysis this loss is ascribed to formamide (or NH3+CO). Elimination of formamide from a2 ions requires that the ions cyclize. A mechanism for this elimination is proposed on the basis of theoretical calculations. The structure of the a2-45 ions is confirmed by their fragmentation in MS3 experiments. Loss of formamide from a2 ions occurs in competition with other neutral losses, such as H2O. However, if the a2 ion contains methionine, a neutral loss of 48 Da, ascribed to CH3SH, predominates, and is followed by loss of formamide. The intensity of the a2-48 peak formed from XaaMet has a maximum value of 42% (of the total intensities of all ions) for Xaa = Gly, varies between 15% and 40% for most other Xaa residues, is lower for residues that can undergo loss of water or ammonia, and very low for Lys or Arg. When the order of the residues is reversed to MetXaa the loss of 48 Da is much smaller. This effect can be used to determine the sequence of b2 ions containing Met in proteomic studies. Direct loss of CH3SH from b2 ions is much less favorable than loss of CO to form a2 ions. Considerable loss of CH3SH is observed from doubly protonated tryptic tripeptides with N-terminal Met, but much less when singly protonated or when Met is in the center position.
Journal of the American Society for Mass Spectrometry