title: Properties of isotope patterns and their utility for peptide identification in large-scale proteomic experiments.
abstract: BACKGROUND
  In proteomic experiments, isotope patterns are routinely generated for all detected peptides. While this pattern is determined by peptide composition, it has not been evaluated as a parameter that can help in the process of peptide identification.
  
  
  METHODS
  First, we investigated how the relative isotope abundance (RIA) accuracy in proteomic data sets depends on the spectral intensity, resolution, and the number of mass spectrometry (MS) 1 scans, using an Orbitrap Velos mass spectrometer. Next, we explored the discriminatory power of isotope patterns in the context of proteome analyses of various complexities, either alone or in combination with a Mascot database search. Finally, we provide a theoretical framework for the required accuracies of both peptide mass and RIA for peptide identification.
  
  
  RESULTS
  We demonstrate that the RIA error obtained in routine proteome analyses is 4-5%, and that this is only modestly influenced by spectral intensity, resolution, and the number of MS1 scans. While RIA alone has no discriminatory power, in a Mascot search isotope patterns can distinguish top scoring hits from runner-up hits in 70-95% of cases. Our theoretical approach shows that RIA accuracy needs to be ~0.2% in order to uniquely identify peptides in full proteomes.
  
  
  CONCLUSIONS
  Our results demonstrate that isotope patterns can have discriminatory power when used in combination with a classical database search. Inclusion of this parameter in proteomic workflows may help to increase confidence in peptide identification, but in practical terms this will be limited to small proteomes.
authors.affiliation: European Molecular Biology Laboratory, Genome Biology Unit, Heidelberg, Germany.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Okawa
authors.forename: Bernd
authors.initials: B
authors.lastname: Fischer
authors.forename: Jeroen
authors.initials: J
authors.lastname: Krijgsveld
mesh_terms: D006367:HeLa Cells; D006801:Humans; D007554:Isotopes / Q000032:analysis; D010455:Peptides / Q000032:analysis*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D007554:Isotopes; D010455:Peptides; D020543:Proteome