Surface Accessibility and Dynamics of Macromolecular Assemblies Probed by Covalent Labeling Mass Spectrometry and Integrative Modeling.
Surface Accessibility and Dynamics of Macromolecular Assemblies Probed by Covalent Labeling Mass Spectrometry and Integrative Modeling.
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DOI：
10.1021/acs.analchem.6b02875
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发表时间：
2017-02-07
影响因子：
7.4
通讯作者：
Politis A
中科院分区：
文献类型：
--
作者：
Schmidt C;Macpherson JA;Lau AM;Tan KW;Fraternali F;Politis A
质谱已成为研究大分子组装体结构和动力学的重要工具。在这里，我们表明，共价标记的溶剂可访问的残基，然后由他们的MS为基础的识别产量建模的限制，允许映射的位置和方向的亚基内的蛋白质组装。连同来自交联和本地MS的互补限制，我们建立了四个具有已知亚基结构的杂合物的类似本地的模型，并将其与可用的X射线晶体结构进行比较。结果表明，共价标记后，MS显着增加的预测能力的综合建模策略，使更准确的蛋白质组装模型。我们将这一策略应用于菠菜叶绿体的F型ATP合酶（cATPase），为其作为nanoparticles的功能提供了结构基础。通过我们的限制为基础的策略产生的模型进行分子动力学（MD）模拟，我们揭示了周边柄的构象状态和分配灵活的区域中的酶。我们的策略可以很容易地将互补的化学标记策略，我们预计，它将适用于许多其他系统提供新的见解的蛋白质复合物的结构和功能。
Mass spectrometry (MS) has become an indispensable tool for investigating the architectures and dynamics of macromolecular assemblies. Here we show that covalent labeling of solvent accessible residues followed by their MS-based identification yields modeling restraints that allow mapping the location and orientation of subunits within protein assemblies. Together with complementary restraints derived from cross-linking and native MS, we built native-like models of four heterocomplexes with known subunit structures and compared them with available X-ray crystal structures. The results demonstrated that covalent labeling followed by MS markedly increased the predictive power of the integrative modeling strategy enabling more accurate protein assembly models. We applied this strategy to the F-type ATP synthase from spinach chloroplasts (cATPase) providing a structural basis for its function as a nanomotor. By subjecting the models generated by our restraint-based strategy to molecular dynamics (MD) simulations, we revealed the conformational states of the peripheral stalk and assigned flexible regions in the enzyme. Our strategy can readily incorporate complementary chemical labeling strategies and we anticipate that it will be applicable to many other systems providing new insights into the structure and function of protein complexes.