Does Electron Capture Dissociation Cleave Protein Disulfide Bonds?

Peptide and protein characterization by mass spectrometry (MS) relies on their dissociation in the gas phase into specific fragments whose mass values can be aligned as ‘mass ladders’ to provide sequence information and to localize possible posttranslational modifications. The most common dissociation method involves slow heating of even-electron (M+n H)(n+) ions from electrospray ionization by energetic collisions with inert gas, and cleavage of amide backbone bonds. More recently, dissociation methods based on electron capture or transfer were found to provide far more extensive sequence coverage through unselective cleavage of backbone N–Cα bonds. As another important feature of electron capture dissociation (ECD) and electron transfer dissociation (ETD), their unique unimolecular radical ion chemistry generally preserves labile posttranslational modifications such as glycosylation and phosphorylation. Moreover, it was postulated that disulfide bond cleavage is preferred over backbone cleavage, and that capture of a single electron can break both a backbone and a disulfide bond, or even two disulfide bonds between two peptide chains. However, the proposal of preferential disulfide bond cleavage in ECD or ETD has recently been debated. The experimental data presented here reveal that the mechanism of protein disulfide bond cleavage is much more intricate than previously anticipated.