Prediction of Hopeless Peptides Unlikely to be Selected for Targeted Proteome Analysis

Abstract

In targeted proteomics using liquid chromatography-tandem triple quadrupole mass spectrometry (LC/MS/MS) in the selected reaction monitoring (SRM) mode, selecting the best observable or visible peptides is a key step in the development of SRM assay methods of target proteins. A direct comparison of signal intensities among all candidate peptides by brute-force LC/MS/MS analysis is a concrete approach for peptide selection. However, the analysis requires an SRM method with hundreds of transitions. This study reports on the development of a method for predicting and identifying hopeless peptides to reduce the number of candidate peptides needed for brute-force experiments. Hopeless peptides are proteotypic peptides that are unlikely to be selected for targets in SRM analysis owing to their poor ionization characteristics. Targeted proteomics data from Escherichia coli demonstrated that the relative ionization efficiency between two peptides could be predicted from sequences of two peptides, when a multivariate regression model is used. Validation of the method showed that >20% of the candidate peptides could be successfully eliminated as hopeless peptides with a false positive rate of less than 2%.