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Wang L, Chen Z, Wu H, et al. Purification and cryoelectron microscopy structure determination of human V-ATPase. STAR Protoc. 2021;2(1):100350doi: 10.1016/j.xpro.2021.100350.
Wang, L., Chen, Z., Wu, H., & Fu, T. M. (2021). Purification and cryoelectron microscopy structure determination of human V-ATPase. STAR protocols, 2(1), 100350. https://doi.org/10.1016/j.xpro.2021.100350
Wang, Longfei, et al. "Purification and cryoelectron microscopy structure determination of human V-ATPase." STAR protocols vol. 2,1 (2021): 100350. doi: https://doi.org/10.1016/j.xpro.2021.100350
Wang L, Chen Z, Wu H, Fu TM. Purification and cryoelectron microscopy structure determination of human V-ATPase. STAR Protoc. 2021 Feb 16;2(1):100350. doi: 10.1016/j.xpro.2021.100350. eCollection 2021 Mar 19. PMID: 33665630; PMCID: PMC7902551.
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STAR Protoc. 2021 Feb 16;2(1):100350. doi: 10.1016/j.xpro.2021.100350. eCollection 2021 Mar 19.

Purification and cryoelectron microscopy structure determination of human V-ATPase.

STAR protocols

Longfei Wang, Zhenhang Chen, Hao Wu, Tian-Min Fu

Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
  2. Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, MA 02115, USA.
  3. Department of Biological Chemistry and Pharmacology, The Ohio State University, Columbus, OH 43210, USA.
  4. The Ohio State University Comprehensive Cancer Center, Columbus, OH 43210, USA.

PMID: 33665630 PMCID: PMC7902551 DOI: 10.1016/j.xpro.2021.100350

Abstract

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).

© 2021 The Author(s).

Keywords: Biophysics; Cryo-EM; Molecular biology; Protein biochemistry; Protein expression and purification; Structural biology

Conflict of interest statement

H.W. is a member of scientific advisory board of Cell. The other authors declare no competing interests.

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