Protein Identification from Tandem Mass Spectra by Database Searching

Protein identification from tandem mass spectra is one of the most versatile and widely used proteomics workflows, able to identify proteins, characterize post-translational modifications, and provide semi-quantitative measurements of relative protein abundance. This manuscript describes the concepts, prerequisites, and methods required to analyze a tandem mass spectrometry dataset in order to identify its proteins, by using a tandem mass spectrometry search engine to search protein sequence databases. The discussion includes instructions for extraction, preparation, and formatting of spectral datafiles; selection of appropriate search parameter settings; and basic interpretation of the results.