Reporter Ion Data Analysis Reduction (R.I.D.A.R) for isobaric proteomics quantification studies

Isobaric labeling-based relative quantification techniques such as iTRAQ and TMT were introduced 15 years ago and are now practically ubiquitous in shotgun proteomics labs around the world. The methods of data processing for these experiments has changed little since inception, with peptide database searching of all MS/MS spectra occurring concurrent or asynchronous to the quantification of the reporter fragment regions. In this study we present an alternative method for data processing whereby the reporter ion region of all MS/MS spectra are first examined and spectra that are not quantitatively interesting to the end user are discarded. The remaining MS/MS spectra that are retained can then be more rapidly searched for computationally expensive database alterations such as post-translational modifications and single amino acid variations in more practical time. We have termed this method Reporter Ion Data Analysis Reduction (RIDAR). To demonstrate the application of RIDAR, we reprocess a recent CPTAC 2 study containing approximately 7.8 million MS/MS spectra. Post RIDAR processing we can search this public dataset versus a human canonical FASTA database and a compiled proteogenomic database of over 875,000 known cancer mutations in a single day on a standard desktop computer, a time reduction of 85% compared to the conventional workflow. With the rapidly increasing size and density of shotgun proteomics data files, RIDAR facilitates rapid analysis of large proteomics datasets for researchers without access to high performance computational resources.