Global proteomic profiling of phosphopeptides using electron transfer dissociation tandem mass spectrometry
Global proteomic profiling of phosphopeptides using electron transfer dissociation tandem mass spectrometry
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DOI：
10.1073/pnas.0611217104
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发表时间：
2007-02-13
影响因子：
11.1
通讯作者：
Pandey, Akhilesh
中科院分区：
文献类型：
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作者：
Molina, Henrik;Horn, David M.;Pandey, Akhilesh
电子转移解离（ETD）是最近引入的质谱技术，提供了更全面的肽序列和翻译后修饰的覆盖。在这里，我们评估了使用ETD的全球磷酸化蛋白质组分析。总之，我们从人胚肾293 T细胞中鉴定了总共1，435个磷酸化位点，其中1，141个（约80%）以前没有描述过。ETD和碰撞诱导解离（CID）模式的详细比较表明，ETD识别的磷酸肽比CID多60%，平均多40%的碎片离子，有利于磷酸化位点的定位。虽然我们的数据表明，ETD是上级的磷酸化分析CID，这两种方法可以有效地结合在交替ETD和CID模式，更全面的分析。结合ETD和CID，从这项研究中，我们能够在分析的> 11，000个磷酸化肽中鉴定出80%的已知磷酸化位点。对已识别的磷酸化位点进行分层聚类，使我们发现了15个以前未报道过的磷酸化基序。总的来说，ETD是定位磷酸化位点的一种很好的方法，应该是任何综合磷酸化分析策略的一个组成部分。
Electron transfer dissociation (ETD) is a recently introduced mass spectrometric technique that provides a more comprehensive coverage of peptide sequences and posttranslational modifications. Here, we evaluated the use of ETD for a global phosphoproteome analysis. In all, we identified a total of 1,435 phosphorylation sites from human embryonic kidney 293T cells, of which 1,141 (approximate to 80%) were not previously described. A detailed comparison of ETD and collision-induced dissociation (CID) modes showed that ETD identified 60% more phosphopeptides than CID, with an average of 40% more fragment ions that facilitated localization of phosphorylation sites. Although our data indicate that ETD is superior to CID for phosphorylation analysis, the two methods can be effectively combined in alternating ETD and CID modes for a more comprehensive analysis. Combining ETD and CID, from this single study, we were able to identify 80% of the known phosphorylation sites in > 11,000 phosphorylated peptides analyzed. A hierarchical clustering of the identified phosphorylation sites allowed us to discover 15 phosphorylation motifs that have not been reported previously. Overall, ETD is an excellent method for localization of phosphorylation sites and should be an integral component of any strategy for comprehensive phosphorylation analysis.