Structure of [M + H − H2O]+ from Protonated Tetraglycine Revealed by Tandem Mass Spectrometry and IRMPD Spectroscopy

Multiple-stage tandem mass spectrometry and collision-induced dissociation were used to investigate loss of H2O or CH3OH from protonated versions of GGGX (where X = G, A, and V), GGGGG, and the methyl esters of these peptides. In addition, wavelength-selective infrared multiple photon dissociation was used to characterize the [M + H − H2O]+ product derived from protonated GGGG and the major MS3 fragment, [M + H − H2O − 29]+ of this peak. Consistent with the earlier work [ Ballard K. D. ; Gaskell S. J. J. Am. Soc. Mass Spectrom. 1993, 4, 477−481 ; Reid G. E. ; Simpson R. J. ; O’Hair R. A. J. Int. J. Mass Spectrom. 1999, 190/191, 209−230 ], CID experiments show that [M + H − H2O]+ is the dominant peak generated from both protonated GGGG and protonated GGGG−OMe. This strongly suggests that the loss of the H2O molecule occurs from a position other than the C-terminal free acid and that the product does not correspond to formation of the b4 ion. Subsequent CID of [M + H − H2O]+ supports this proposal by resulting in a major product that is 29 mass units less than the precursor ion. This is consistent with loss of HNCH2 rather than loss of carbon monoxide (28 mass units), which is characteristic of oxazolone-type b n ions. Comparison between experimental and theoretical infrared spectra for a group of possible structures confirms that the [M + H − H2O]+ peak is not a substituted oxazolone but instead suggests formation of an ion that features a five-membered ring along the peptide backbone, close to the amino terminus. Additionally, transition structure calculations and comparison of theoretical and experimental spectra of the [M + H − H2O − 29]+ peak also support this proposal.