ARS | Publication request: The problem with peptide presumption and the downfall of target-decoy false discovery rates

Submitted to: Analytical Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 25, 2012
Publication Date: October 29, 2012
Citation: Cooper, B. 2012. The problem with peptide presumption and the downfall of target-decoy false discovery rates. Analytical Chemistry. 84:9663-9667.

Interpretive Summary: Mass spectrometry is used to identify proteins in biological samples. Given a mixture of peptides derived from the proteins, the mass spectrometer produces a spectrum which can be interpreted by software. The software assigns a score that says if the spectrum is a particular peptide versus some other peptide or some other substance like a sugar, fatty acid or oil. It is necessary to apply a scientific-based threshold to determine the lower end of acceptability for scores. However, some scientists are now accepting lower than normal scores. This study shows how the acceptance of low scores leads to questionable scientific data. This study also shows that the reasoning used to justify low scores is based on whim. The purpose of this research is to encourage the development of standards that lead to better scientific data quality. These findings are important to scientists at universities, institutes, government agencies and companies who want to assign better confidence to the peptides and proteins they discover by mass spectrometry.

Technical Abstract: Mascot Expect values and target-decoy database derived false discovery rates (FDR) are confidence indicators describing the quality of individual and sets of peptide-tandem mass spectrum matches. A user can impose a quality standard by prescribing a limit to these values, equivalent to drawing an arbitrary line that separates better from poorer quality matches. As a result of force-feeding peptide sequence candidates to search engines, a bias inherent to the method, and setting narrower parent ion mass tolerances to reflect the resolution of a mass spectrometer, target-decoy derived FDRs can diminish and Expect values can improve. For FDRs in particular, this can also drive down the lower-limit for match score acceptance. For each, data quality confidence appears to improve even while fragmentation evidence for some spectra remains weak. One negative outcome can be the presumed identification of peptides that do not exist despite confidence indicators that suggest otherwise.