Global Ion Suppression Limits the Potential of Mass Spectrometry Based Phosphoproteomics,Journal of Proteome Research

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Global Ion Suppression Limits the Potential of Mass Spectrometry Based Phosphoproteomics
Journal of Proteome Research ( IF 3.8 ) Pub Date : 2018-11-12 , DOI: 10.1021/acs.jproteome.8b00812
Roland Felix Dreier ₁ , Erik Ahrné ₁ , Petr Broz ₁ , Alexander Schmidt ₁

Affiliation

Mass spectrometry based proteomics has become the method of choice for pinpointing and monitoring thousands of post-translational modifications, predominately phosphorylation sites, in cellular signaling studies. Critical for achieving this analytical depth is the enrichment of phosphorylated peptides prior to liquid chromatography–mass spectrometry (MS) analysis. Despite the high prevalence of this modification, the numbers of identified phosphopeptides lag behind those achieved for unmodified peptides, and the cause for this still remains controversial. Here, we use an effective phosphatase protocol that considerably improves global ionization efficiency and, therefore, the overall sensitivity and coverage of standard phosphoproteomics studies. We demonstrate the power of our method on the model system of Salmonella-infected macrophages by extending the current quantitative picture of immune signaling pathways involved in infection. In combination with sensitive, label-free targeted MS for phosphorylation site validation, our approach is ideally suited to exploring cellular phosphorylation based signaling networks in high detail.

中文翻译：

全球离子抑制限制了基于质谱的磷酸化蛋白质组学的潜力

基于质谱的蛋白质组学已成为精确定位和监测细胞信号研究中成千上万个翻译后修饰（主要是磷酸化位点）的选择方法。达到此分析深度的关键是在液相色谱-质谱（MS）分析之前先富集磷酸化的肽。尽管这种修饰的流行率很高，但是鉴定出的磷酸肽的数量仍落后于未修饰的肽，而其原因仍然是有争议的。在这里，我们使用有效的磷酸酶方案，可显着提高整体电离效率，因此可提高标准磷酸化蛋白质组学研究的总体敏感性和覆盖范围。我们证明了我们的方法在沙门氏菌模型系统中的作用感染巨噬细胞，方法是扩大目前涉及感染的免疫信号通路的定量图像。与敏感的，无标记的靶向MS结合用于磷酸化位点验证，我们的方法非常适合于详细研究基于细胞磷酸化的信号网络。

更新日期：2018-11-12

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