Lookup peaks: A hybrid of de novo sequencing and database search for protein identification by tandem mass spectrometry
Lookup peaks: A hybrid of de novo sequencing and database search for protein identification by tandem mass spectrometry
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DOI：
10.1021/ac0617013
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发表时间：
2007-02-15
影响因子：
7.4
通讯作者：
Goldberg, David
中科院分区：
文献类型：
--
作者：
Bern, Marshall;Cai, Yuhan;Goldberg, David
肽和蛋白质鉴定的一种强大技术是串联质谱法，然后使用SEQUEST或Mascot等程序进行数据库搜索。然而，当允许非特异性切割或肽修饰时，这些程序变得缓慢并失去灵敏度。从头测序和杂交方法，如序列标记，为更广泛的搜索提供了速度和鲁棒性，但这些方法需要更好的光谱，更完整和连续的片段，因此，对低丰度肽不太敏感。在这里，我们描述了一种新的混合方法，保留了纯数据库搜索的灵敏度。该方法使用少量的从头分析来识别可能的B-和y-离子峰“查找峰“，其然后可以用于从数据库中提取候选肽，其中候选肽的数量可调以适合计算预算。我们描述了一个名为ByOnic的程序，它实现了这种方法，我们在几个数据集上对ByOnic进行了基准测试，其中包括一个掺入低浓度重组人蛋白的小鼠血浆。我们证明了ByOnic比序列标记更敏感，事实上，比三个最流行的纯数据库搜索工具SEQUEST，Mascot和X更敏感！Tandemon的肽和蛋白质水平。在小鼠血浆样本中，ByOnic始终发现其他工具遗漏的加标蛋白质。
A powerful technique for peptide and protein identification is tandem mass spectrometry followed by database search using a program such as SEQUEST or Mascot. These programs, however, become slow and lose sensitivity when allowing nonspecific cleavages or peptide modifications. De novo sequencing and hybrid methods such as sequence tagging offer speed and robustness for wider searches, yet these approaches require better spectra with more complete and consecutive fragmentation and, hence, are less sensitive to low-abundance peptides. Here we describe a new hybrid method that retains the sensitivity of pure database search. The method uses a small amount of de novo analysis to identify likely b- and y-ion peaks"lookup peaks"that can then be used to extract candidate peptides from the database, with the number of candidates tunable to fit a computing budget. We describe a program called ByOnic that implements this method, and we benchmark ByOnic on several data sets, including one of mouse blood plasma spiked with low concentrations of recombinant human proteins. We demonstrate that ByOnic is more sensitive than sequence tagging and, indeed, more sensitive than the three most popular pure database search toolsSEQUEST, Mascot, and X!Tandemon both the peptide and protein levels. On the mouse plasma samples, ByOnic consistently found spiked proteins missed by the other tools.