Correlation of multiple peptide mass spectra for phosphoprotein identification.

When collision induced dissociation is used to fragment phosphorylated peptides during tandem mass spectrometry (MS(2)), an ion exhibiting the neutral loss of phosphoric acid can be the major product. The neutral loss ion can then be fragmented during MS(3) for additional resolution of the peptide sequence. Together, MS(2) and MS(3) spectral pairs can offer supporting identification of phosphorylated peptides and proteins. Here, the software program PANORAMICS has been modified to make use of Mascot results for MS(2) and MS(3) spectral data sets. For pairs, the algorithm considers the number of shared m/z peaks used for peptide assignment and then adjusts the score evaluating that a peptide was correctly matched to these spectra using a mathematical model. The algorithm then calculates peptide probabilities for paired and unpaired spectra, and deduces a probability that a protein was identified given the set of matched peptides. The output provides information useful for determining whether peptides and proteins are phosphorylated. The program can process large result files often generated by multidimensional protein identification technology (MudPIT).