Protein Probability Model for High-Throughput Protein Identification by Mass Spectrometry-Based Proteomics.

Shotgun proteomics is the method of choice for high-throughput protein identification; however, robust statistical methods are essential to automatize this task while minimizing the number of false identifications. The standard method for estimating the false discovery rate (FDR) of individual identifications and keeping it below a threshold (typically 1%) is the target-decoy approach. However, numerous works have shown that FDR at the protein level may become much larger than FDR at the peptide level. The development of an appropriate scoring model to identify proteins from their peptides using high-throughput shotgun proteomics is highly needed. In this study, we present a novel protein-level scoring algorithm that uses the scores of the identified peptides and maintains all of the properties expected for a true protein probability. We also present a refinement of the method to calculate FDR at the protein level. These algorithms can be used together as a robust identification workflow suitable for large-scale proteomics, and we show that the identification performance of this workflow is superior to that of other widely used methods in several samples and using different search engines. Our protein probability model offers the scientific community an algorithm that is easy to integrate into protein identification workflows for the automated analysis of shotgun proteomics data.