Investigation of Product Ions Generated by 193 nm Ultraviolet Photodissociation of Peptides and Proteins Containing Disulfide Bonds

Disulfide bridges are unique post-translational modifications (PTM) that contribute to protein architecture and modulate function. This PTM, however, challenges top-down mass spectrometry by cyclizing stretches of the protein sequence. In order to produce and release detectable product ions that contribute to the assignment of proteoforms, regions of a protein encapsulated by disulfide bonds require two fragmentation events: cleavage of the protein backbone and cleavage of the disulfide bond. Traditional collisional activation methods do not cleave disulfide bonds efficiently, often leading to low sequence coverage of proteins that incorporate this feature. To address this challenge, we have evaluated the fragmentation pathways enabled by 193 nm ultraviolet photodissociation (UVPD) and UVPD coupled to electron transfer dissociation for the characterization of protein structures incorporating disulfide bonds. Cleavage of disulfide bonds by either approach results in S-S and C-S dissociation products that result from a combination of homolytic cleavage and hydrogen-transfer processes. Characterization of these product ions elevates interpretation of complex top-down spectra of proteins that incorporate disulfide bonds.

Automated summary

Disulfide bridges are post-translational modifications that impact the structure and function of proteins. Traditional collisional activation methods are not efficient in cleaving disulfide bonds, making it difficult to analyze proteins that incorporate this feature. Researchers have evaluated the fragmentation pathways enabled by 193 nm ultraviolet photodissociation and photodissociation coupled to electron transfer for the characterization of protein structures incorporating disulfide bonds.