期刊
JOURNAL OF PROTEOME RESEARCH
卷 5, 期 9, 页码 2448-2456出版社
AMER CHEMICAL SOC
DOI: 10.1021/pr060200v
关键词
de novo sequencing; database searching; borderline hits; MS/MS; MS BLAST; PepNovo
资金
- NIGMS NIH HHS [1R01GM070986-01A1] Funding Source: Medline
Protein identifications with the borderline statistical confidence are typically produced by matching a few marginal quality MS/MS spectra to database peptide sequences and represent a significant bottleneck in the reliable and reproducible characterization of proteomes. Here, we present a method for rapid validation of borderline hits that circumvents the need in, often biased, manual inspection of raw MS/MS spectra. The approach takes advantage of the independent interpretation of corresponding MS/MS spectra by PepNovo de novo sequencing software followed by mass spectrometry-driven BLAST ( MS BLAST) sequence-similarity database searches that utilize all partially inaccurate, degenerate and redundant candidate peptide sequences. In a case study involving the identification of more than 180 Caenorhabditis elegans proteins by nanoLC-MS/MS analysis on a linear ion trap LTQ mass spectrometer, the approach enabled rapid assignment ( confirmation or rejection) of more than 70% of Mascot hits of borderline statistical confidence.
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