Abstract
BackgroundConcurrent peptide fragmentation (i.e. shotgun CID, parallel CID or MSE) has emerged as an alternative to data-dependent acquisition in generating peptide fragmentation data in LC-MS/MS proteomics experiments. Concurrent peptide fragmentation data acquisition has been shown to be advantageous over data-dependent acquisition by providing greater detection dynamic range and providing more accurate quantitative information. Nevertheless, concurrent peptide fragmentation data acquisition remains to be widely adopted due to the lack of published algorithms designed specifically to process or interpret such data acquired on any mass spectrometer.ResultsAn algorithm called Elution Time Ion Sequencing (ETISEQ), has been developed to enable automated conversion of concurrent peptide fragmentation data acquisition data to LC-MS/MS data. ETISEQ generates MS/MS-like spectra based on the correlation of precursor and product ion elution profiles. The performance of ETISEQ is demonstrated using concurrent peptide fragmentation data from tryptic digests of standard proteins and whole influenza virus. It is shown that the number of unique peptides identified from the digests is broadly comparable between ETISEQ processed concurrent peptide fragmentation data and the data-dependent acquired LC-MS/MS data.ConclusionThe ETISEQ algorithm has been designed for easy integration with existing MS/MS analysis platforms. It is anticipated that it will popularize concurrent peptide fragmentation data acquisition in proteomics laboratories.
Highlights
Concurrent peptide fragmentation has emerged as an alternative to data-dependent acquisition in generating peptide fragmentation data in Liquid chromatography (LC)-MS/MS proteomics experiments
For the purpose for this algorithm, concurrent peptide fragmentation data acquisition (CDA) data is defined as an LCMS experiment where, throughout the duration of the experiment, survey MS scans are alternated with MS/MSlike scans
For the whole influenza virus digest, there were a number of identified peptides that were unique to CDA data despite the fact that less unique peptides were identified for the reconstructed LC-MS/MS data in comparison to dependent acquisition (DDA) data (Table 2)
Summary
Concurrent peptide fragmentation (i.e. shotgun CID, parallel CID or MSE) has emerged as an alternative to data-dependent acquisition in generating peptide fragmentation data in LC-MS/MS proteomics experiments. While technological improvements are continually being made in chromatography [4], mass spectrometry [5,6] and mass spectra interpretation algorithms [7], the detection of lower abundance proteins or proteolytic peptides in complex mixtures remains an obstacle in most proteomics experiments [8,9]. These (page number not for citation purposes).
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