Effect of proline position on symmetric versus asymmetric fragmentation of doubly-protonated tryptic-type peptides

Abstract

The fragmentation reactions of the doubly-protonated tryptic-type peptides APAAAAR, AAPAAAR, AAAPAAR and AAAAPAR have been studied using an electrospray/quadrupole/time-of-flight (QqTo F) mass spectrometer. The tendency to cleave amide bonds N-terminal to proline (P) is in competition with the tendency to cleave the second amide bond, counting from the N-terminus; the result of such competition depends on the position of the Pro residue. When the Pro residue is remote from the Arg (R) residue a strong proline effect is observed resulting in formation, to a large extent, of a doubly-charged y species and a neutral fragment, so-called asymmetric amide bond cleavage. By contrast, when the Pro residue approaches the C-terminal Arg residue the proline effect is reduced with respect to cleavage of the second amide bond; in both cases formation of singly-charged y and b ions, so-called symmetric bond cleavage, increases significantly in importance. The results are discussed in terms of relative energetics for symmetric and asymmetric bond cleavage as revealed by approximate proton affinities of the b species and the singly-charged y species.