Potential crosslinked peptides were checked for the possibility that the data fit better to a linear peptide than to a crosslinked peptide. One indication that a linear peptide is a better fit is seeing that one peptide is matched with all b-ions and the other with all y-ions. A second indication is the absence of fragments specific for the shorter peptide. Searches for linear peptides were made with the MS-Tag feature of Protein Prospector. Access to MS-Tag is made via the Search Compare Search Results page in Protein Prospector. This page lists the retention time for each potential crosslink product. A click on the retention time (RT) brings up the MS-Tag page. MS-Tag provides an efficient means for searching a selected MS/MS data set against a large database. We searched each spectrum 3 times. The variables were 1) database NCBInr 2013.6.17 and Taxonomy All species, 2) database NCBInr 2013.6.17 and Taxonomy Sus scrofa, and 3) 6-user protein database and Taxonomy Sus Scrofa. A constant parameter in each search was Crosslinking> User Defined Link: No Link. The No Link setting instructed the software to search for linear peptides that fit the MS/MS data. Searching All species provided a means of finding unexpected linear fits such as human keratin.

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