Ultrafast Ultraviolet Photodissociation at 193 nm and its Applicability to Proteomic Workflows

摘要

The results in the present study illustrate the utility of using ultrafast UVPD for high-throughput proteomic workflows. UVPD at 193 nm results in ample production of a, b, c, x, y, and z sequence ions, in addition to immonium ions and v and w side-chain loss ions. Precursor dissociation efficiencies range from 50-98percent for doubly-charged peptides using a single laser pulse. Peptides containing one or more amino acids with aromatic side-chains exhibit higher dissociation efficiencies than peptides without aromatic guoups, and the peptide amide bond quantity has relatively little impact on UVPD efficiency. A background subtraction procedure to account for the formation of rather high abundances of ions due to photoionization of background organic species during the UVPD period was implemented and significantly improved subsequent SEQUEST scoring and peptide identifications for complex mixtures analyzed by LC-UVPD-MS. Comparable and often improved in silico searching results were achievable for biologically relevant samples, including a human HT-1080 cytosolic fibrosarcoma cell better than the slower CID approach even for complex biological samples based on this high-throughput, shotgun-style proteomic strategy.