Rapid validation of protein identifications with the borderline statistical confidence via de novo sequencing and MS BLAST searches
Rapid validation of protein identifications with the borderline statistical confidence via de novo sequencing and MS BLAST searches
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DOI：
10.1021/pr060200v
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发表时间：
2006-09-01
影响因子：
4.4
通讯作者：
Shevchenko, Andrej
中科院分区：
文献类型：
--
作者：
Wielsch, Natalie;Thomas, Henrik;Shevchenko, Andrej
具有临界统计置信度的蛋白质鉴定通常通过将一些边际质量MS/MS谱与数据库肽序列相匹配来产生，并且代表了蛋白质组的可靠和可再现表征中的显著瓶颈。在这里，我们提出了一种快速验证边界命中的方法，该方法避免了对原始MS/MS光谱进行手动检查的需要。该方法利用PepNovo de novo测序软件对相应MS/MS光谱的独立解释，然后利用所有部分不准确、简并和冗余的候选肽序列进行质谱驱动的BLAST（MS BLAST）序列相似性数据库搜索。在一项涉及在线性离子阱LTQ质谱仪上通过nanoLC-MS/MS分析鉴定超过180种秀丽隐杆线虫蛋白质的案例研究中，该方法能够快速分配（确认或拒绝）超过70%的Mascot命中临界统计置信度。
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.