Structural insights into the membrane receptor ShuA in DDM micelles and in a model of gram-negative bacteria outer membrane as seen by SAXS and MD simulations
Structural insights into the membrane receptor ShuA in DDM micelles and in a model of gram-negative bacteria outer membrane as seen by SAXS and MD simulations
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DOI：
10.1016/j.bbamem.2020.183504
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发表时间：
2021-02-01
影响因子：
3.4
通讯作者：
Bonnete, Francoise
中科院分区：
文献类型：
--
作者：
Abel, Stephane;Marchi, Massimo;Bonnete, Francoise
膜蛋白在洗涤剂胶束中的成功结晶取决于关键因素，如胶束组装体中蛋白质的构象稳定性、蛋白质-洗涤剂复合物（PDC）的单分散性以及通过洗涤剂带适当屏蔽蛋白质疏水表面而形成的有利的蛋白质晶体接触。为了研究两亲性环境对膜蛋白结构、稳定性和可结晶性的影响，我们将联合收割机分子动力学（MD）模拟与SEC-MALLS和SEC-SAXS（尺寸排阻色谱结合多角度激光散射或小角度X射线散射）实验相结合，来描述蛋白-去污剂相互作用，这有助于合理化PDC结晶。在这种情况下，我们比较了蛋白质洗涤剂的相互作用ShuA从志贺菌n-Dodecyl β-D-Maltopyranoside（DDM）与ShuA插入在一个现实的模型，革兰氏阴性菌外膜（OM）含有细菌脂多糖和磷脂的混合物。为了评估PDC模型的质量，我们从MD轨迹计算相应的SAXS曲线，并与实验结果进行比较。我们表明，计算的SAXS曲线从MD轨迹再现更好的SAXS从SEC-SAXS实验ShuA包围268 DDM分子。分子动力学结果表明DDM分子在ShuA周围形成一个封闭的带，其疏水厚度似乎略小于（类似于22埃）由蛋白质在膜中的取向（OPM）数据库建议的疏水跨膜结构域（24.6埃）。模拟还表明，ShuA跨膜结构域是非常稳定的，在所有的系统中，除了细胞外和周质环，表现出较大的运动，由于特定的分子与脂多糖（LPS）的相互作用。我们最后指出，这种去污剂的行为可能会导致闭塞的周质亲水表面和不良的晶体接触，导致难以结晶的ShuA在DDM。
Successful crystallization of membrane proteins in detergent micelles depends on key factors such as conformational stability of the protein in micellar assemblies, the protein-detergent complex (PDC) monodispersity and favorable protein crystal contacts by suitable shielding of the protein hydrophobic surface by the detergent belt. With the aim of studying the influence of amphiphilic environment on membrane protein structure, stability and crystallizability, we combine molecular dynamics (MD) simulations with SEC-MALLS and SEC-SAXS (Size Exclusion Chromatography in line with Multi Angle Laser Light Scattering or Small Angle X-ray Scattering) experiments to describe the protein-detergent interactions that could help to rationalize PDC crystallization. In this context, we compare the protein-detergent interactions of ShuA from Shigella dysenteriae in n-Dodecyl beta-D-Maltopyranoside (DDM) with ShuA inserted in a realistic model of gram-negative bacteria outer membrane (OM) containing a mixture of bacterial lipopolysaccharide and phospholipids. To evaluate the quality of the PDC models, we compute the corresponding SAXS curves from the MD trajectories and compare with the experimental ones. We show that computed SAXS curves obtained from the MD trajectories reproduce better the SAXS obtained from the SEC-SAXS experiments for ShuA surrounded by 268 DDM molecules. The MD results show that the DDM molecules form around ShuA a closed belt whose the hydrophobic thickness appears slightly smaller (similar to 22 angstrom) than the hydrophobic transmembrane domain of the protein (24.6 angstrom) suggested by Orientations of Proteins in Membranes (OPM) database. The simulations also show that ShuA transmembrane domain is remarkably stable in all the systems except for the extracellular and periplasmic loops that exhibit larger movements due to specific molecular interactions with lipopolysaccharides (LPS). We finally point out that this detergent behavior may lead to the occlusion of the periplasmic hydrophilic surface and poor crystal contacts leading to difficulties in crystallization of ShuA in DDM.