Similarity between condensed phase and gas phase chemistry: Fragmentation of peptides containing oxidized cysteine residues and its implications for proteomics

Amino acid residues containing thioethers are easily oxidized during protein purification, derivatization, and/or digestion. For instance, oxidation of methionine residues in proteins during SDS-PAGE is commonly observed. Under low energy collision induced dissociation this gives rise to a second series of fragment ion of lower abundance that are shifted by -64 Da when compared to the oxidized methionine-containing fragments. We report here that alkylated cysteine residues can be found in their oxidized form too, indicating that the oxidation of thioethers can occur during and following protein digestion and not only during SDS-PAGE or reduction and alkylation. Collision induced dissociation experiments on the singly- and multiply-charged species reveals that these peptides preferentially undergo elimination reactions that forms a dehydroalanine from the oxidized, alkylated cysteine residue. This contrasts to the less abundant elimination reaction of peptides containing oxidized methionines which cannot form an alpha,beta-unsaturated compound, but parallels the condensed phased chemistry of sulfoxides. The masses of both precursor and product ions are shifted such that these peptides cannot be identified in database searches with current algorithms. Incorporation of this fragmentation pattern is important for the isotope-coded affinity tag approach since this method is based on peptides containing cysteine residues.