Tabb David L, Friedman David B, Ham Amy-Joan L
Department of Biochemistry, Vanderbilt University Medical Center, Nashville, Tennessee 37232-8340, USA.
Nat Protoc. 2006;1(5):2213-22. doi: 10.1038/nprot.2006.330.
Shotgun proteomics yields tandem mass spectra of peptides that can be identified by database search algorithms. When only a few observed peptides suggest the presence of a protein, establishing the accuracy of the peptide identifications is necessary for accepting or rejecting the protein identification. In this protocol, we describe the properties of peptide identifications that can differentiate legitimately identified peptides from spurious ones. The chemistry of fragmentation, as embodied in the 'mobile proton' and 'pathways in competition' models, informs the process of confirming or rejecting each spectral match. Examples of ion-trap and tandem time-of-flight (TOF/TOF) mass spectra illustrate these principles of fragmentation.
鸟枪法蛋白质组学可产生肽段的串联质谱,这些肽段可通过数据库搜索算法进行鉴定。当只有少数观察到的肽段提示某种蛋白质的存在时,确定肽段鉴定的准确性对于接受或拒绝蛋白质鉴定是必要的。在本方案中,我们描述了能够区分真实鉴定的肽段和虚假肽段的肽段鉴定属性。碎片化的化学过程,如“移动质子”和“竞争途径”模型所体现的,为确认或拒绝每个光谱匹配的过程提供了依据。离子阱和串联飞行时间(TOF/TOF)质谱的示例说明了这些碎片化原理。