Charging behavior of highly basic peptides during electrospray ionization. A predilection for protons

Abstract

The extent of charging (or protonation) during the electrospray ionization process has been examined for a series of specifically constructed arginine-rich peptides, which differ in structure by the length of the peptide chain and the number and proximity of arginine residues. It has been found that although a small peptide of the series will protonate fully, supporting a charge on each arginine side-chain, the same charging behavior is not observed for larger peptides with the same repeating primary structure. Furthermore, no significant increase in the extent of charging was observed as the length of the peptide chain, or the distance between potential charge-bearing sites, was increased. The apparent sites of protonation in the [M + nH] n+ peptide ions have been examined for several representative peptides based on the extent of protonation compared to that of structurally related peptides, and their dissociation behavior. Despite the potential for proton migration during the collisional activation event, the fragmentation pattern of the peptide ions studied suggests that the charge-bearing protons are reasonably localized at the time of dissociation commensurate with our previous observations for singly and multiply charged peptide ions. The charging behavior of the model peptides is discussed in the context of a reported mechanism for the electrospray ionization process.