Nakanishi Atsuko, Kishikawa Jun-Ichi, Mitsuoka Kaoru, Yokoyama Ken
Department of Molecular Biosciences, Kyoto Sangyo University, Kyoto 603-8555, Japan.
Research Center for Ultra-High Voltage Electron Microscopy, Osaka University, Ibaraki, Osaka 567-0047 Japan.
Biophys Physicobiol. 2019 Sep 3;16:140-146. doi: 10.2142/biophysico.16.0_140. eCollection 2019.
Proton-translocating rotary ATPases couple proton influx across the membrane domain and ATP hydrolysis/synthesis in the soluble domain through rotation of the central rotor axis against the surrounding peripheral stator apparatus. It is a significant challenge to determine the structure of rotary ATPases due to their intrinsic conformational heterogeneity and instability. Recent progress of single particle analysis of protein complexes using cryogenic electron microscopy (cryo-EM) has enabled the determination of whole rotary ATPase structures and made it possible to classify different rotational states of the enzymes at a near atomic resolution. Three cryo-EM maps corresponding to different rotational states of the V/A type H-rotary ATPase from a bacterium provide insights into the rotation of the whole complex, which allow us to determine the movement of each subunit during rotation. In addition, this review describes methodological developments to determine higher resolution cryo-EM structures, such as specimen preparation, to improve the image contrast of membrane proteins.
质子转运旋转ATP合酶通过中央转子轴相对于周围的外周定子装置旋转，将跨膜结构域的质子内流与可溶性结构域中的ATP水解/合成偶联起来。由于其固有的构象异质性和不稳定性，确定旋转ATP合酶的结构是一项重大挑战。最近使用低温电子显微镜（cryo-EM）对蛋白质复合物进行单颗粒分析的进展，使得能够确定整个旋转ATP合酶的结构，并有可能以接近原子分辨率对酶的不同旋转状态进行分类。来自一种细菌的V/A型H-旋转ATP合酶的三种对应不同旋转状态的低温电子显微镜图谱，为整个复合物的旋转提供了见解，这使我们能够确定每个亚基在旋转过程中的运动。此外，本综述描述了确定更高分辨率低温电子显微镜结构的方法学进展，如样品制备，以提高膜蛋白的图像对比度。
