CαC Backbone Fragmentation Dominates in Electron Detachment Dissociation of Gas-Phase Polypeptide Polyanions

Fragmentation of peptide polyanions by electron detachment dissociation (EDD) has been induced by electron irradiation of deprotonated polypeptides [M−nH]n− with >10 eV electrons. EDD has been found to lead preferentially to a. and x fragment ions (CαC backbone cleavage) arising from the dissociation of oxidized radical anions [M−nH](n−1)−.. We demonstrate that CαC cleavages, which are otherwise rarely observed in tandem mass spectrometry, can account for most of the backbone fragmentation, with even‐electron x fragments dominating over radical a. ions. Ab initio calculations at the B3 LYP level of theory with the 6‐311+G(2 p,2 d)//6‐31+G(d,p) basis set suggested a unidirectional mechanism for EDD (cleavage always N‐terminal to the radical site), with a., x formation being favored over a, x. fragmentation by 74.2 kJ mol−1. Thus, backbone CαC bonds N‐terminal to proline residues should be immune to EDD, in agreement with the observations. EDD may find application in mass spectrometry for such tasks as peptide sequencing and localization of labile post‐translational modifications, for example, those introduced by sulfation and phosphorylation. EDD can now be performed not only in Fourier transform mass spectrometry, but also in far more widely used quadrupole (Paul) ion traps. Rare type a. and x fragment ions are found to be the dominant products of electron‐induced excitation of polypeptide radical anions. Computational and experimental investigations suggest that a highly preferential cleavage mechanism is initiated at the amide nitrogen radical site and proceeds in the direction of the N‐terminus (see scheme). This mechanism is unique among gas‐phase fragmentation phenomena and provides new insight into the reaction chemistry of radical anions.