This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Mass spectrometric identification of cross-linked peptides can provide valuable information about the structure of protein complexes. We describe a straightforward database search scheme that identifies and assigns statistical confidence estimates to spectra from cross-linked peptides. The method is well suited to targeted analysis of a single protein complex, without requiring an isotope labeling strategy. Our approach uses a SEQUEST-style search procedure in which the database is comprised of a mixture of single peptides with and without linkers attached and cross-linked products. In contrast to several previous approaches, we generate theoretical spectra that account for all of the expected peaks from a cross-linked product, and we employ an empirical curve-fitting procedure to estimate statistical confidence measures. We show that our fully automated procedure successfully reidentifies spectra from a previous study, and we provide evidence that our statistical confidence estimates are accurate.
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