Köcher Thomas, Savitski Mikhail M, Nielsen Michael L, Zubarev Roman A
Laboratory for Biological and Medical Mass Spectrometry, Uppsala University, Uppsala, SE-75123, Sweden.
J Proteome Res. 2006 Mar;5(3):659-68. doi: 10.1021/pr0503836.
A database independent search algorithm for the detection of phosphopeptides is described. The program interrogates the tandem mass spectra of LC-MS/MS data sets regarding the presence of phosphorylation specific signatures. To achieve maximum informational content, the complementary fragmentation techniques electron capture dissociation (ECD) and collisionally activated dissociation (CAD) are used independently for peptide fragmentation. Several criteria characteristic for peptides phosphorylated on either serine or threonine residues were evaluated. The final algorithm searches for product ions generated by either the neutral loss of phosphoric acid or the combined neutral loss of phosphoric acid and water. Various peptide mixtures were used to evaluate the program. False positive results were not observed because the program utilizes the parts-per-million mass accuracy of Fourier transform ion cyclotron resonance mass spectrometry. Additionally, false negative results were not generated owing to the high sensitivity of the chosen criteria. The limitations of database dependent data interpretation tools are discussed and the potential of the novel algorithm to overcome these limitations is illustrated.
本文描述了一种用于检测磷酸化肽段的独立于数据库的搜索算法。该程序会针对磷酸化特异性特征的存在情况，查询液相色谱-串联质谱（LC-MS/MS）数据集的串联质谱图。为了获得最大信息量，互补碎裂技术电子捕获解离（ECD）和碰撞激活解离（CAD）被分别用于肽段碎裂。评估了丝氨酸或苏氨酸残基磷酸化肽段的几个特征标准。最终算法搜索由磷酸的中性丢失或磷酸和水的联合中性丢失产生的产物离子。使用了各种肽混合物来评估该程序。由于该程序利用了傅里叶变换离子回旋共振质谱的百万分之一质量精度，未观察到假阳性结果。此外，由于所选标准的高灵敏度，也未产生假阴性结果。讨论了依赖于数据库的数据解释工具的局限性，并说明了新算法克服这些局限性的潜力。
