Calibrating E-values for MS2 database search methods.
BACKGROUND
The key to mass-spectrometry-based proteomics is peptide identification, which relies on software analysis of tandem mass spectra. Although each search engine has its strength, combining the strengths of various search engines is not yet realizable largely due to the lack of a unified statistical framework that is applicable to any method.


RESULTS
We have developed a universal scheme for statistical calibration of peptide identifications. The protocol can be used for both de novo approaches as well as database search methods. We demonstrate the protocol using only the database search methods. Among seven methods -SEQUEST (v27 rev12), ProbID (v1.0), InsPecT (v20060505), Mascot (v2.1), X!Tandem (v1.0), OMSSA (v2.0) and RAId_DbS - calibrated, except for X!Tandem and RAId_DbS most methods require a rescaling according to the database size searched. We demonstrate that our calibration protocol indeed produces unified statistics both in terms of average number of false positives and in terms of the probability for a peptide hit to be a true positive. Although both the protocols for calibration and the statistics thus calibrated are universal, the calibration formulas obtained from one laboratory with data collected using either centroid or profile format may not be directly usable by the other laboratories. Thus each laboratory is encouraged to calibrate the search methods it intends to use. We also address the importance of using spectrum-specific statistics and possible improvement on the current calibration protocol. The spectra used for statistical (E-value) calibration are freely available upon request.
Alves
Gelio
G
National Center for Biotechnology Information, National Library of Medicine, NIH, Bethesda, MD 20894, USA. alves@ncbi.nlm.nih.gov
Ogurtsov
Aleksey Y
AY
Wu
Wells W
WW
Wang
Guanghui
G
Shen
Rong-Fong
RF
Yu
Yi-Kuo
YK
D000595:Amino Acid Sequence; D002138:Calibration; D030562:Databases, Protein / Q000706:statistics & numerical data*; D007256:Information Systems; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010449:Peptide Mapping / Q000379:methods; D040901:Proteomics / Q000379:methods; D020539:Sequence Analysis, Protein / Q000379:methods; D012984:Software