Quantifying the stabilizing effects of protein-ligand interactions in the gas phase.
Quantifying the stabilizing effects of protein-ligand interactions in the gas phase.
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DOI：
10.1038/ncomms9551
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发表时间：
2015-10-06
影响因子：
16.6
通讯作者：
Robinson CV
中科院分区：
文献类型：
--
作者：
Allison TM;Reading E;Liko I;Baldwin AJ;Laganowsky A;Robinson CV
蛋白质-配体相互作用对蛋白质稳定性的影响通常通过许多已建立的溶液相测定来监测。很少有蛋白质容易翻译成膜蛋白。我们已经开发了一种离子迁移率质谱法，识别配体结合的可溶性和膜蛋白直接通过质量和离子迁移率的变化，并评估这些相互作用对蛋白质稳定性的影响，通过测量阻力展开。蛋白质解折叠是通过碰撞激活诱导的，这导致蛋白质结构的变化，从而导致气相流动性。这使得能够以前所未有的灵敏度详细表征配体对蛋白质的结合作用。在这里，我们描述的方法和软件需要从离子迁移率数据中提取的参数，使单个结合事件的定量分析。这种方法具有很大的希望，为调查膜蛋白和药物和脂质之间的生物学意义的相互作用，这是通过其他手段的表征。相对较少的技术可以定量测量配体对膜蛋白稳定性的影响。在这里，作者演示了使用离子迁移率质谱法来准确测量和量化配体诱导的蛋白质稳定在气相中。
The effects of protein–ligand interactions on protein stability are typically monitored by a number of established solution-phase assays. Few translate readily to membrane proteins. We have developed an ion-mobility mass spectrometry approach, which discerns ligand binding to both soluble and membrane proteins directly via both changes in mass and ion mobility, and assesses the effects of these interactions on protein stability through measuring resistance to unfolding. Protein unfolding is induced through collisional activation, which causes changes in protein structure and consequently gas-phase mobility. This enables detailed characterization of the ligand-binding effects on the protein with unprecedented sensitivity. Here we describe the method and software required to extract from ion mobility data the parameters that enable a quantitative analysis of individual binding events. This methodology holds great promise for investigating biologically significant interactions between membrane proteins and both drugs and lipids that are recalcitrant to characterization by other means. Relatively few techniques can quantitatively measure the effect of ligands on membrane protein stability. Here the authors demonstrate the use of ion-mobility mass spectrometry to accurately measure and quantify ligand-induced protein stabilization in the gas phase.