Mechanical inhibition of isolated Vo from V/A-ATPase for proton conductance

Abstract
Data availability
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Abstract

V-ATPase is an energy converting enzyme, coupling ATP hydrolysis/synthesis in the hydrophilic V₁ domain, with proton flow through the V_o membrane domain, via rotation of the central rotor complex relative to the surrounding stator apparatus. Upon dissociation from the V₁ domain, the V_o domain of the eukaryotic V-ATPase can adopt a physiologically relevant auto-inhibited form in which proton conductance through the V_o domain is prevented, however the molecular mechanism of this inhibition is not fully understood. Using cryo-electron microscopy, we determined the structure of both the holo V/A-ATPase and isolated V_o at near-atomic resolution, respectively. These structures clarify how the isolated V_o domain adopts the auto-inhibited form and how the holo complex prevents formation of the inhibited V_o form.

Data availability

The density maps and the built models for Tth VoV1, Tth V1 (focused refined), and Tth Vo were deposited in EMDB (EMDB ID; 30013, 30014, and 30015) and PDB (PDB ID; 6LY8 for V1 and 6LY9 for isolated Vo), respectively. All data is available in the main text or the supplementary materials.

The following data sets were generated

1. Kishikawa J
2. Nakanishi A
3. Furuta A
4. Kato T
5. Namba K
6. Tamakoshi M
7. Mitsuoka K
8. Yokoyama K
(2020) V/A-ATPase from Thermus thermophilus
30013.

https://www.ebi.ac.uk/pdbe/emdb/
1. Kishikawa J
2. Nakanishi A
3. Furuta A
4. Kato T
5. Namba K
6. Tamakoshi M
7. Mitsuoka K
8. Yokoyama K
(2020) V1-ATPase built from cryo-EM map of V/A-ATPase from Thermus thermophilus.
30014.

https://www.ebi.ac.uk/pdbe/emdb/
1. Kishikawa J
2. Nakanishi A
3. Furuta A
4. Kato T
5. Namba K
6. Tamakoshi M
7. Mitsuoka K
8. Yokoyama K
(2020) The membrane-embedded Vo domain of V/A-ATPase from Thermus thermophilus
30015.

https://www.ebi.ac.uk/pdbe/emdb/
1. Kishikawa J
2. Nakanishi A
3. Furuta A
4. Kato T
5. Namba K
6. Tamakoshi M
7. Mitsuoka K
8. Yokoyama K
(2020) V/A-ATPase from Thermus thermophilus, the soluble domain, including V1, d, two EG stalks, and N-terminal domain of a-subunit.
6LY8.

https://www.rcsb.org/structure/unreleased/6LY8
1. Kishikawa J
2. Nakanishi A
3. Furuta A
4. Kato T
5. Namba K
6. Tamakoshi M
7. Mitsuoka K
8. Yokoyama K
(2020) The membrane-embedded Vo domain of V/A-ATPase from Thermus thermophilus
6LY9.

https://www.rcsb.org/structure/unreleased/6LY9

Article and author information

Author details

Jun-ichi Kishikawa

Department of Molecular Biosciences, Kyoto Sangyo University, Kyoto, Japan

Competing interests
The authors declare that no competing interests exist.
Atsuko Nakanishi

Department of Molecular Biosciences, Kyoto Sangyo University, Kyoto, Japan

Competing interests
The authors declare that no competing interests exist.
Aya Furuta

Department of Molecular Biosciences, Kyoto Sangyo University, Kyoto, Japan

Competing interests
The authors declare that no competing interests exist.
Takayuki Kato

Institute for Protein Research, Osaka University, Osaka, Japan

Competing interests
The authors declare that no competing interests exist.
Keiichi Namba

Institute for Protein Research, Osaka University, Osaka, Japan

Competing interests
The authors declare that no competing interests exist.
Masatada Tamakoshi

Department of Molecular Biology, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan

Competing interests
The authors declare that no competing interests exist.
Kaoru Mitsuoka

Research Center for Ultra-High Voltage Electron Microscopy, Osaka University, Osaka, Japan

Competing interests
The authors declare that no competing interests exist.
Ken Yokoyama

Department of Molecular Biosciences, Kyoto Sangyo University, Kyoto, Japan

For correspondence
yokoken@cc.kyoto-su.ac.jp

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-6813-1096

Funding

Japan Society for the Promotion of Science (17H03648)

Ken Yokoyama

Japan Agency for Medical Research and Development (JP17am0101001)

Kaoru Mitsuoka

Ministry of Education, Culture, Sports, Science, and Technology (12024046)

Kaoru Mitsuoka

Takeda Science Foundation

Ken Yokoyama

Japan Science and Technology Agency (JPMJCR1865)

Kaoru Mitsuoka

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.