SEC-TMT facilitates quantitative differential analysis of protein interaction networks.
SEC-TMT facilitates quantitative differential analysis of protein interaction networks.
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SEC-TMT 有助于蛋白质相互作用网络的定量差异分析。
DOI：
10.1101/2023.01.12.523793
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Jovanovic,Marko
中科院分区：
文献类型：
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作者：
Doron-Mandel,Ella;Bokor,BenjaminJ;Ma,Yanzhe;Street,LenaA;Tang,LaurenC;Abdou,AhmedA;Shah,NeelH;Rosenberger,GeorgeA;Jovanovic,Marko
蛋白质分子相互作用和翻译后修饰（PTMs），如磷酸化，可以相互依赖和相互调节。尽管这种相互作用是许多生物过程的核心，但同时研究来自同一样品的组装状态和ptm的系统方法严重缺乏。本文介绍了SEC-MX（粒径排除色谱法），这是一种将粒径排除色谱法和ptm富集相结合的全球性定量方法，可同时表征PTMs和组装态。SEC-MX提高了吞吐量，允许磷酸肽富集，并促进了生物条件之间的定量差异比较。将SEC-MX应用于HEK293和HCT116细胞，我们生成了一个概念验证数据集，绘制了数千个磷酸肽及其组装状态。我们的分析揭示了磷酸化事件和组装状态之间的复杂关系，并为后续研究产生了可测试的假设。总之，我们建立了SEC-MX作为探索蛋白质功能和丰度变化之外的调控的有价值的工具。
Protein molecular interactions and post-translational modifications (PTMs), such as phosphorylation, can be co-dependent and reciprocally co-regulate each other. Although this interplay is central for many biological processes, a systematic method to simultaneously study assembly-states and PTMs from the same sample is critically missing. Here, we introduce SEC-MX (Size Exclusion Chromatography fractions MultipleXed), a global quantitative method combining Size Exclusion Chromatography and PTM-enrichment for simultaneous characterization of PTMs and assembly-states. SEC-MX enhances throughput, allows phosphopeptide enrichment, and facilitates quantitative differential comparisons between biological conditions. Applying SEC-MX to HEK293 and HCT116 cells, we generated a proof-of-concept dataset mapping thousands of phosphopeptides and their assembly-states. Our analysis revealed intricate relationships between phosphorylation events and assembly-states and generated testable hypotheses for follow-up studies. Overall, we establish SEC-MX as a valuable tool for exploring protein functions and regulation beyond abundance changes.