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Enhanced identification of peptides lacking basic residues by LC‐ESI‐MS/MS analysis of singly charged peptides
Author(s) -
Biniossek Martin L.,
Schilling Oliver
Publication year - 2012
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201100569
Subject(s) - chemistry , electrospray ionization , chromatography , tandem mass spectrometry , proteome , peptide , mass spectrometry , trypsin , histidine , isobaric labeling , lysine , tandem mass tag , bottom up proteomics , proteomics , electrospray , liquid chromatography–mass spectrometry , protein mass spectrometry , biochemistry , amino acid , quantitative proteomics , enzyme , gene
Peptide sequences lacking basic residues (arginine, lysine, or histidine, referred to as “base‐less”) are of particular importance in proteomic experiments targeting protein C ‐termini or employing nontryptic proteases such as G lu C or chymotrypsin. We demonstrate enhanced identification of base‐less peptides by focused analysis of singly charged precursors in liquid chromatography ( LC ) electrospray ionization (ESI) tandem mass spectrometry ( MS/MS ). Singly charged precursors are often excluded from fragmentation and sequence analysis in LC ‐ MS/MS . We generated different pools of base‐less and base‐containing peptides by tryptic and nontryptic digestion of bacterial proteomes. Focused LC ‐ MS/MS analysis of singly charged precursor ions yielded predominantly base‐less peptide identifications. Similar numbers of base‐less peptides were identified by LC‐MS/MS analysis targeting multiply charged precursors. There was little redundancy between the base‐less sequences derived by both MS/MS schemes. In the present experimental outcome, additional LC‐MS/MS analysis of singly charged precursors substantially increased the identification rate of base‐less sequences derived from multiply charged precursors. In conclusion, LC‐MS/MS based identification of base‐less peptides is substantially enhanced by additional focused analysis of singly charged precursors.