METHOD TO CORRELATE TANDEM MASS-SPECTRA OF MODIFIED PEPTIDES TO AMINO-ACID-SEQUENCES IN THE PROTEIN DATABASE

A method to correlate uninterpreted tandem mass spectra of modified pe ptides, produced under low-energy (10-50 eV) collision conditions, wit h amino acid sequences in a protein database has been developed, The f ragmentation patterns observed in the tandem mass-spectra of peptides containing covalent modifications is used to directly search and fit l inear amino acid sequences in the database, Specific information relev ant to sites of modification is not contained in the character-based s equence information of the databases, The search method considers each putative modification site as both modified and unmodified in one pas s through the database and simultaneously considers up to three differ ent sites of modification, The search method will identify the correct sequence if the tandem mass spectrum did not represent a modified pep tide. This approach is demonstrated with peptides containing modificat ions such as S-carboxymethylated cysteine, oxidized methionine, phosph oserine, phosphothreonine, or phosphotyrosine. In addition, a scanning approach is used in which neutral loss scans are used to initiate the acquisition of product ion MS/MS spectra of doubly charged phosphoryl ated peptides during a single chromatographic ran for data analysis wi th the database-searching algorithm, The approach described in this pa per provides a convenient method to match the nascent tandem mass spec tra of modified peptides to sequences in a protein database and thereb y identify previously unknown sites of modification.