CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Université Paris-Saclay, 91198 Gif-sur-Yvette, France.
J Phys Chem B. 2020 Dec 31;124(52):11802-11818. doi: 10.1021/acs.jpcb.0c08902. Epub 2020 Dec 21.
Structural studies of proteins and, in particular, integral membrane proteins (IMPs) using solution NMR spectroscopy approaches are challenging due to not only their inherent structural complexities but also the fact that they need to be solubilized in biomimetic environments (such as micelles), which enhances the slow molecular reorientation. To deal with these difficulties and increase the effective rate of molecular reorientation, the encapsulation of IMPs in the aqueous core of the reverse micelle (RM) dissolved in a low-viscosity solvent has been proven to be a viable approach. However, the effect of the reverse micelle (RM) environment on the IMP structure and function is little known. To gain insight into these aspects, this article presents a series of atomistic unconstrained molecular dynamics (MD) of a model ion channel (gramicidin A, gA) with RMs formed with anionic surfactant diacyl chain bis(2-ethylhexyl) sodium succinate (AOT) in pentane at a water-to-surfactant molar ratio () of 6. The simulations were carried out with different protocols and starting conditions for a total of 2.4 μs and were compared with other MDs used with the gA channel inserted in models of the SDS micelle or the DMPC membrane. We show here that in the presence of AOT RMs the gA dimer did not look like the "dumbbell-like" model anticipated by experiments, where the C-terminal parts of the gA are capped with two RMs and the rest of the dimer is protected from the oil solvent by the AOT acyl chains. In contrast, the MD simulations reveal that the AOT, Na, and water formed two well-defined and elongated RMs attached to the C-terminal ends of the gA dimer, while the rest is in direct contact with the pentane. The initial β secondary structure of the gA is well conserved and filled with 6-9 waters, as in SDS micelles or the DMPC membrane. Finally, the water movement inside the gA is strongly affected by the presence of RMs at each extremity, and no passage of water molecules through the gA channel is observed even after a long simulation period, whereas the opposite was found for gA in SDS and DMPC.
使用溶液 NMR 光谱学方法研究蛋白质，特别是整合膜蛋白 (IMP)，具有挑战性，不仅因为它们具有固有结构复杂性，还因为它们需要在仿生环境（如胶束）中溶解，这会增加分子的缓慢重取向。为了应对这些困难并提高分子重取向的有效速率，已证明将 IMP 包封在溶解在低粘度溶剂中的反胶束 (RM) 的水核中是一种可行的方法。然而，反胶束 (RM) 环境对 IMP 结构和功能的影响知之甚少。为了深入了解这些方面，本文介绍了一系列使用阴离子表面活性剂二酰基链双(2-乙基己基)琥珀酸钠 (AOT) 在戊烷中形成的反胶束 (RM) 的模型离子通道（革兰氏菌素 A，gA）的原子非约束分子动力学 (MD) 模拟。在水与表面活性剂摩尔比（）为 6 的情况下，使用不同的方案和起始条件进行了模拟，总模拟时间为 2.4 μs，并与其他在 SDS 胶束或 DMPC 膜模型中插入 gA 通道的 MD 进行了比较。我们在这里表明，在 AOT RM 的存在下，gA 二聚体看起来不像实验预期的“哑铃状”模型，其中 gA 的 C 末端部分被两个 RM 封闭，二聚体的其余部分被 AOT 酰基链保护免受油溶剂的影响。相反，MD 模拟表明，AOT、Na 和水形成了两个定义明确且拉长的 RM，附着在 gA 二聚体的 C 末端，而其余部分则直接与戊烷接触。gA 的初始β二级结构得到很好的保留，并填充了 6-9 个水分子，就像在 SDS 胶束或 DMPC 膜中一样。最后，RM 的存在强烈影响 gA 内水的运动，即使在长时间的模拟后，也观察不到水分子通过 gA 通道，而在 SDS 和 DMPC 中则发现了相反的情况。
