Fujiyoshi Yoshinori
Structural Physiology, Department of Biophysics, Graduate School of Science, Kyoto University, Oiwake, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan.
J Electron Microsc (Tokyo). 2011;60 Suppl 1:S149-59. doi: 10.1093/jmicro/dfr033.
Membrane proteins are important research targets for basic biological sciences and drug design, but studies of their structure and function are considered difficult to perform. Studies of membrane structures have been greatly facilitated by technological and instrumental advancements in electron microscopy together with methodological advancements in biology. Electron crystallography is especially useful in studying the structure and function of membrane proteins. Electron crystallography is now an established method of analyzing the structures of membrane proteins in lipid bilayers, which resembles their natural biological environment. To better understand the neural system function from a structural point of view, we developed the cryo-electron microscope with a helium-cooled specimen stage, which allows for analysis of the structures of membrane proteins at a resolution higher than 3 Å. This review introduces recent instrumental advances in cryo-electron microscopy and presents some examples of structure analyses of membrane proteins, such as bacteriorhodopsin, water channels and gap junction channels. This review has two objectives: first, to provide a personal historical background to describe how we came to develop the cryo-electron microscope and second, to discuss some of the technology required for the structural analysis of membrane proteins based on cryo-electron microscopy.
膜蛋白是基础生物学和药物设计的重要研究靶点，但对其结构和功能的研究被认为难以开展。电子显微镜技术和仪器的进步以及生物学方法的进步极大地推动了膜结构的研究。电子晶体学在研究膜蛋白的结构和功能方面特别有用。电子晶体学现在是分析脂质双分子层中膜蛋白结构的一种成熟方法，脂质双分子层类似于膜蛋白的天然生物环境。为了从结构角度更好地理解神经系统功能，我们开发了带有氦冷却样品台的冷冻电子显微镜，它能够以高于3 Å的分辨率分析膜蛋白的结构。本综述介绍了冷冻电子显微镜最近的仪器进展，并展示了一些膜蛋白结构分析的实例，如细菌视紫红质、水通道和间隙连接通道。本综述有两个目的：第一，提供个人历史背景，描述我们是如何开发冷冻电子显微镜的；第二，讨论基于冷冻电子显微镜的膜蛋白结构分析所需的一些技术。
