MS/MS of Protonated Polyproline Peptides: The Influence of N-terminal Protonation on Dissociation

Abstract

A unique collision-induced dissociation pattern was observed for protonated polyproline peptides of length n in which y(n-2) and/or y(n-4) ions were formed in much higher abundance than any other product ions. Cleavage occurs only at every other amide bond, such that product ions are formed only from the losses of even numbers of proline residues. Exclusive losses of even numbers of proline residues were not observed from sodiated peptides. Further study of the tandem mass spectrometry (MS/MS) patterns of protonated proline-rich peptides showed that the substitution of alanine in the second position of polyproline peptides did not prevent the dominant formation of y(n-2) and y(n-4) ions. The loss of ProAla to form the y(8) ion from (ProAlaPro(8)NH(2)+H)(+) was as abundant as the loss of ProPro from (Pro(10)NH(2)+H)(+). However, modification of the peptides that presumably affected the location of the proton on the peptide did alter the MS/MS spectra. Pro(10) and Pro(5) with blocked N-termini or with arginine substituted for the first proline residue did not form abundant y(n-2) or y(n-4) ions. MS(3) and double resonance experiments showed that dissociation of intermediate y(n) product ions can produce y(n-2) ions, but are not necessary dissociation pathway intermediates. This analysis suggests that the ionizing proton must be located at the N-terminus for the peptide ion to dissociate in this manner.