Protein and Lipid Interactions Driving Molecular Mechanisms of in meso Crystallization
Protein and Lipid Interactions Driving Molecular Mechanisms of in meso Crystallization
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DOI：
10.1021/ja4129839
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发表时间：
2014-02-26
影响因子：
15
通讯作者：
Khelashvili, George
中科院分区：
文献类型：
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作者：
Johner, Niklaus;Mondal, Sayan;Khelashvili, George
近年来，用于表征G蛋白偶联受体(GPCRs)结构的介观结晶技术的最新进展证实了脂立方相(LCPs)在膜蛋白结晶学领域的应用。令人惊讶的是，尽管这种方法取得了成功，但介观方法的分子机制仍然没有被很好地理解。因此，该方法必须依赖于对适当的蛋白质结构、宿主和添加剂脂类以及适当的沉淀剂和温度的广泛筛选。为了阐明In介观结晶机制，我们使用了广泛的粗粒度分子动力学模拟，详细地研究了不同条件下(与晶体形成相关的组成和温度)的LCP及其与不同类型的GPCR结构的相互作用。结果表明，LCP晶格常数的调节(在介观分析过程中添加沉淀剂触发)或宿主脂类的调节如何破坏LCP双层中单体蛋白质的稳定，从而推动它们聚集到堆叠的片层中，在那里蛋白质和膜之间残留的疏水失配可以驱动侧向接触的形成，从而导致成核和晶体生长。此外，我们还演示了特定的蛋白质设计(例如包含大极区的跨膜蛋白质)如何在第三个平面外维度促进蛋白质堆积相互作用。对立方相内蛋白质相互作用的特定分子机制的新方面所提供的见解应该有助于指导未来在成功结果的介观试验中的合理设计。
The recent advances in the in meso crystallization technique for the structural characterization of G-protein coupled receptor (GPCR) proteins have established the usefulness of the lipidic-cubic phases (LCPs) in the field of crystallography of membrane proteins. It is surprising that despite the success of the approach, the molecular mechanisms of the in meso method are still not well understood. Therefore, the approach must rely on extensive screening for a suitable protein construct, for host and additive lipids, and for the appropriate precipitants and temperature. To shed light on the in meso crystallization mechanisms, we used extensive coarse-grained molecular dynamics simulations to study, in molecular detail, LCPs under different conditions (compositions and temperatures relevant to crystallogenesis) and their interactions with different types of GPCR constructs. The results presented show how the modulation of the lattice constant of the LCP (triggered by the addition of precipitant during the in meso assay), or of the host lipid type, can destabilize monomeric proteins in the bilayer of the LCP and thus drive their aggregation into the stacked lamellae, where the residual hydrophobic mismatch between the protein and the membrane can drive the formation of lateral contacts leading to nucleation and crystal growth. Moreover, we demonstrate how particular protein designs (such as transmembrane proteins engineered to contain large polar regions) can promote protein stacking interactions in the third, out-of-plane, dimension. The insights provided by the new aspects of the specific molecular mechanisms responsible for protein protein interactions inside the cubic phase presented here should be helpful in guiding the rational design of future in meso trials with successful outcomes.