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Origin of Disagreements in Tandem Mass Spectra Interpretation by Search Engines.

Several proteomic database search engines that interpret LC-MS/MS data do not identify the same set of peptides. These disagreements occur even when the scores of the peptide-to-spectrum matches suggest good confidence in the interpretation. Our study shows that these disagreements observed for the interpretations of a given spectrum are almost exclusively due to the variation of what we call the "peptide space", i.e., the set of peptides that are actually compared to the experimental spectra. We discuss the potential difficulties of precisely defining the "peptide space." Indeed, although several parameters that are generally reported in publications can easily be set to the same values, many additional parameters-with much less straightforward user access-might impact the "peptide space" used by each program. Moreover, in a configuration where each search engine identifies the same candidates for each spectrum, the inference of the proteins may remain quite different depending on the false discovery rate selected.