PhosphoHunter: An Efficient Software Tool for Phosphopeptide Identification

Figure 1

Summary of PhosphoHunter procedure. Block A (implemented via the create_database.pl script): a database in FASTA format is used to create a target database according to appropriate digestion rules and other parameters provided in an input file, such as the number of allowed consecutive missing cleavages. The database is then used to obtain a decoy database and a single composite database is obtained from target and decoy databases. Block B (implemented via the merge.pl script): individual dta files, corresponding to experimental spectra, are merged into a single dta file. Block C: experimental spectra are normalized and processed by discarding charges higher than 4, low-intensity peaks, and peptides not showing neutral loss. The intensity threshold of neutral loss is specified in the input file. Block D: theoretical and processed spectra are compared according to a scoring function and a list of phosphopeptides with scores is associated with each spectrum. Block E: for each spectrum, a -value is computed for each element of the list and only the peptides with a -value below a specific threshold, defined in the input file (relation not shown in the figure), are kept in the final list. Blocks C, D, and E are all implemented via the phosphopeptide_ID.pl script.