Supplementary Figure 1: Comparison of methods on the Pyrococcus furiosus example | Nature Methods

Supplementary Figure 1: Comparison of methods on the Pyrococcus furiosus example

From: Mass spectrometrists should search for all peptides, but assess only the ones they care about

Supplementary Figure 1

Histograms of MS-GF+ scores (grey) with estimated number of correct PSMs (#target - #decoys, red) and incorrect PSMs (#decoys, blue), 1% FDR cutoff (dashed line) for the search-all-assess-all (all-all, panel a), the search-subset-assess-subset (sub-sub, panels b, d) and the search-all-assess-subset strategy (all- sub, panel c). The GO term “ATP binding” was used to generate the subset of interest. In panel (a) and (c), the spectra are searched against the complete Pyrococcus database and in panel (b) and (d) against an “ATP binding proteins” only database. Panel (a) shows PSM scores for all PSMs, panels (b) - (d) for the “ATP binding proteins” subset, only. The fraction of incorrect PSMs (π0, first mode in the target distribution) is lower in the complete Pyrococcus set (all-all, panel a) than in the “ATP binding proteins” subset (all-sub, panel c) indicating that the all-all FDR is too liberal. The 1% FDR cutoff in the subsub strategy (panel b) shifted to higher values and this leads to a decrease in the number of subset PSMs found compared to the all-all strategy and the all-sub strategy. It also shows that sub-sub forces many PSMs on incorrect subset PSMs (orange bars in panel d). Indeed 6110 (8617-2507) spectra matching to other Pyrococcus targets/decoys in the complete Pyrococcus search switch to an “ATPbinding” sequence in the subset-search. 1.4% of the sub-sub PSMs above the 1% FDR cutoff have switched peptides sequences (panel d orange) as compared to the complete search (all-all and all-sub strategies). They have much lower scores than in a complete search and are questionable at best (black and orange boxplot below histogram in panel d).

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