Electron capture dissociation for structural characterization of multiply charged protein cations

For proteins of < 20 kDa, this new radical site dissociation method-cleaves different and many more backbone bonds than the conventional MS/MS methods (e.g, collisionally activated dissociation, CAD) that add energy directly to the even-electron ions. A minimum kinetic energy difference between the electron and ion maximizes capture; a 1 eV difference reduces capture by 10 (3). Thus, in an FTMS ion cell with added electron trapping electrodes, cap ture appears to be achieved best at the boundary between the potential well s that trap the electrons and ions, now providing 80 +/- 15% precursor ion conversion efficiency. Capture cross section is dependent on the ionic char ge squared (z(2)), minimizing the secondary dissociation of lower charge fr agment ions. Electron capture is postulated to occur initially at a protona ted site to release an energetic (similar to 6 eV) H-. atom that is capture d ata high-affinity site such as -S-S- or backbone amide to cause nonergodi c (before energy randomization) dissociation. Cleavages between every pair of amino acids in mellitin (2.8 kDa) and ubiquitin (8.6 kDa) are represente d in their ECD and CAD spectra, providing complete data for their de novo s equencing. Because posttranslational modifications-such as carboxylation, g lycosylation, and sulfation are less easily lost in ECD than in CAD, ECD as signments of their sequence positions are far more specific.