Single Particle Analysis of V-type ATPase Using Cryo-EM

Abstract

V-ATPase is a rotary molecular motor which couples ATP synthesis/hydrolysis in the soluble domain with proton flow in the membrane domain by a rotation of the central rotor complex. Here, we present three rotational states of V-ATPase from the bacterium Thermus thermophilus by single particle analysis using cryo-EM. By using a combination of masked classification/refinement and focused classification techniques, the resolutions of these EM maps were improved. These EM maps provide the first detailed insight into the contact surface between each subunit, and the assignment of the movement of each subunit during rotation. In this article, I would like to report the results of structural analysis of V-ATPase and introduce the actual situations of sample preparation and analytical methods for single particle analysis.