Verification of automated peptide identifications from proteomic tandem mass spectra.
Shotgun proteomics yields tandem mass spectra of peptides that can be identified by database search algorithms. When only a few observed peptides suggest the presence of a protein, establishing the accuracy of the peptide identifications is necessary for accepting or rejecting the protein identification. In this protocol, we describe the properties of peptide identifications that can differentiate legitimately identified peptides from spurious ones. The chemistry of fragmentation, as embodied in the 'mobile proton' and 'pathways in competition' models, informs the process of confirming or rejecting each spectral match. Examples of ion-trap and tandem time-of-flight (TOF/TOF) mass spectra illustrate these principles of fragmentation.
Tabb
David L
DL
Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennessee 37232-8340, USA. david.l.tabb@vanderbilt.edu
Friedman
David B
DB
Ham
Amy-Joan L
AJ
D000818:Animals; D001426:Bacterial Proteins / Q000032:analysis; D016480:Helicobacter pylori / Q000737:chemistry; D051379:Mice; D008871:Microvilli / Q000737:chemistry; D010455:Peptides / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D001426:Bacterial Proteins; D010455:Peptides