Improving Performance Metrics of Ultraviolet Photodissociation Mass Spectrometry by Selective Precursor Ejection

Confident protein identifications derived from high throughput bottom-up and top-down proteomics workflows.depend on acquisition of thousands of tandem mass spectrometry (MS/M$) Spectra with adequate signal-to-noise and accurate mass assignments of the fragment ions. Ultraviolet photodissociation (UVPD);using 198 nm photons has proven to be well-suited for activation and fragmentation of peptides and proteins in ion trap mass spectrometers, but the spectral signal-to-noise ratio (S/N) is typically lower than that obtain eaftorir collisional activation methods. The lower S/N is attributed to the dispersion of ion current among numerous fragment Chan' noels (a,b,c,x,y,z ions). In addition, frequently UVPD is performed such that a relatively large population of precursor imiS remains unaissociated after the UV photoactivation period in order to prevent nverdissociation into small uninformative or internal fragment ions. Here we report a method to improve spectral S/N and increase the accuracy of mass assignments of UVPD mass spectra via resonance ejection of undissociated precursor ions after photoactivation. This strategy, termed precursor ejection UVPD or PE-UVPD, allows the ion trap to be filled with more ions prior to UVPD while at the same time alleviating thespaCe,Charge problems that would' otherwise contrilmite to the skewing of mass assignments and reduction of S/N. Here we.report the performance gains by implementation of PE-UVPD for peptide analysis in an ion trap mass spectrometer.