Blasie J Kent
University of Pennsylvania, Philadelphia, PA, United States; University of California Irvine, Irvine, CA, United States.
Methods Enzymol. 2018;603:67-90. doi: 10.1016/bs.mie.2018.01.019. Epub 2018 Mar 15.
The mechanism of electromechanical coupling for voltage-gated ion channels (VGICs) involved in neurological signal transmission, primarily Nav- and Kv-channels, remains unresolved. Anesthetics have been shown to directly impact this mechanism, at least for Kv-channels. Molecular dynamics computer simulations can now predict the structures of VGICs embedded within a hydrated phospholipid bilayer membrane as a function of the applied transmembrane voltage, but significant assumptions are still necessary. Nevertheless, these simulations are providing new insights into the mechanism of electromechanical coupling at the atomic level in 3-D. We show that time-resolved neutron interferometry can be used to investigate directly the profile structure of a VGIC, vectorially oriented within a single hydrated phospholipid bilayer membrane at the solid-liquid interface, as a function of the applied transmembrane voltage in the absence of any assumptions or potentially perturbing modifications of the VGIC protein and/or the host membrane. The profile structure is a projection of the membrane's 3-D structure onto the membrane normal and, in the absence of site-directed deuterium labeling, is provided at substantially lower spatial resolution than the atomic level. Nevertheless, this novel approach can be used to directly test the validity of the predictions from molecular dynamics simulations. We describe the key elements of our novel experimental approach, including why each is necessary and important to providing the essential information required for this critical comparison of "simulation" vs "experiment." In principle, the approach could be extended to higher spatial resolution and to include the effects of anesthetics on the electromechanical coupling mechanism in VGICs.
参与神经信号传递的电压门控离子通道（VGICs），主要是Nav通道和Kv通道，其机电耦合机制仍未得到解决。麻醉剂已被证明会直接影响这一机制，至少对Kv通道是这样。分子动力学计算机模拟现在可以预测嵌入水合磷脂双分子层膜中的VGICs结构，作为施加跨膜电压的函数，但仍需要重大假设。尽管如此，这些模拟正在为三维原子水平的机电耦合机制提供新的见解。我们表明，时间分辨中子干涉测量法可用于直接研究在固液界面单个水合磷脂双分子层膜中矢量取向的VGIC的轮廓结构，作为在没有任何假设或对VGIC蛋白和/或宿主膜进行潜在干扰性修饰的情况下施加跨膜电压的函数。轮廓结构是膜的三维结构在膜法线方向上的投影，并且在没有定点氘标记的情况下，其空间分辨率远低于原子水平。尽管如此，这种新方法可用于直接检验分子动力学模拟预测的有效性。我们描述了我们新实验方法的关键要素，包括为什么每个要素对于提供“模拟”与“实验”这一关键比较所需的基本信息都是必要且重要的。原则上，该方法可以扩展到更高的空间分辨率，并包括麻醉剂对VGICs机电耦合机制的影响。
