N-terminal sequence tagging using reliably determined b2 ions: A useful approach to deconvolute tandem mass spectra of co-fragmented peptides in proteomics
peer-reviewed · Journal of Proteomics · 2014
| Date | 2014-05-01 |
| Type | peer-reviewed |
| Venue | Journal of Proteomics |
| Publisher | Elsevier BV |
| Contribution | adjacent |
| DOI | 10.1016/j.jprot.2014.03.039 |
| Citations (OpenAlex) | 3 |
| Venue 2-year citedness | 3.02 |
Abstract
With the recent introduction of higher-energy collisional dissociation (HCD) in Orbitrap mass spectrometry, the popularity of that technique has grown tremendously in the proteomics society. HCD spectra, however, are characterized by a limited distribution of bn-type ions, which permit the generation of reliable sequence tags based on complementary b,y pairs both for de novo sequencing and sequence tagging strategies. Instead, most peptide HCD spectra (~95%) are dominated with b2 ions. In this work, we analyzed positive predictive values of b2 ions in HCD, and found that b2 ions can be determined with >97% certainty in the presence of a2 and its complementary yn-2 ions. Analytically, b2 ions provide information on the composition of the first two N-terminal amino acids in peptides. Their utilization in N-terminal sequence tagging leads to a significant decrease in false discovery rate by filtering out false positives while retaining true positive identifications. As a consequence, the number of peptide spectrum matches (PSMs) increased by 4.8% at fixed FDR (1%). This approach allows for deconvolution of mixture spectra and increased the number of PSM to 9.2% in a complex human sample and to 24% in a complex sample of synthetic peptides at 1% FDR.
Methods and tools
- b2-ion N-terminal sequence tagging: Shows that b2 ions in HCD spectra can be assigned with over 97% certainty when a2 and the complementary y ion are present, and uses the resulting two-residue N-terminal tags to filter PSMs and deconvolute co-fragmented spectra.