학술논문

Purification and cryoelectron microscopy structure determination of human V-ATPase
Document Type
article
Source
STAR Protocols, Vol 2, Iss 1, Pp 100350- (2021)
Subject
Biophysics
Cryo-EM
Molecular biology
Protein biochemistry
Protein expression and purification
Structural biology
Science (General)
Q1-390
Language
English
ISSN
2666-1667
Abstract
Summary: Vesicular- or vacuolar-type adenosine triphosphatases (V-ATPases) are multi-component, ATP-driven proton pumps, which play important roles in many physiological processes by acidifying intracellular vesicles, organelles, and the extracellular milieu. Long-standing challenges in purifying mammalian V-ATPases have limited the biochemical and structural study of mammalian V-ATPase. Here, we provide a protocol for purifying milligrams of human V-ATPase and detail procedures for the reconstruction of its structure by cryo-EM. Our method can be applied to any biochemical and biophysical study of human V-ATPase.For complete details on the use and execution of this protocol, please refer to Wang et al. (2020).