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Cited by 51ATP Synthase and ATPases ResearchAdvanced Electron Microscopy Techniques and ApplicationsEnzyme Structure and Function

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Structure of ATP synthase under strain during catalysis

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Guo, Hui, Rubinstein, John L.·2022·Nature Communications
DOI 10.1038/s41467-022-29893-2PMC PMC9038767

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Abstract

ATP synthases are macromolecular machines consisting of an ATP-hydrolysis-driven F 1 motor and a proton-translocation-driven F O motor. The F 1 and F O motors oppose each other’s action on a shared rotor subcomplex and are held stationary relative to each other by a peripheral stalk. Structures of resting mitochondrial ATP synthases revealed a left-handed curvature of the peripheral stalk even though rotation of the rotor, driven by either ATP hydrolysis in F 1 or proton translocation through F O , would apply a right-handed bending force to the stalk. We used cryoEM to image yeast mitochondrial ATP synthase under strain during ATP-hydrolysis-driven rotary catalysis, revealing a large deformation of the peripheral stalk. The structures show how the peripheral stalk opposes the bending force and suggests that during ATP synthesis proton translocation causes accumulation of strain in the stalk, which relaxes by driving the relative rotation of the rotor through six sub-steps within F 1 , leading to catalysis.

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Fig. 1 Rotation in ATP synthase. a ATP synthase (left) consists of an F1 and an FO region with a shared rotor subcomplex (outlined in black) and a peripheral stalk (green). Rotation driven by proton translocation through the FO region, or the opposite rotation driven by ATP hydrolysis in the F1 region, are predicted to induce a right-handed bend of the peripheral stalk (right). b ATP hydrolysis or synthesis in the F1 region requires three catalytic (black, yellow, and red) and three bind dwell (light blue, blue, and green) conformations. c High-resolution structure of the yeast mitochondrial ATP synthase. d In the absence of free ATP, the peripheral stalk exhibits limited flexibility with a left-handed curvature. e During ATP hydrolysis, ATP synthase can adopt conformations that show a right-handed curvature of the peripheral stalk. f Histograms of the distribution of conformations in the absence of ATP (top, blue) and during ATP hydrolysis (bottom, purple).
Fig. 1 Rotation in ATP synthase. a ATP synthase (left) consists of an F1 and an FO region with a shared rotor subcomplex (outlined in black) and a peripheral stalk (green). Rotation driven by proton translocation through the FO region, or the opposite rotation driven by ATP hydrolysis in the F1 region, are predicted to induce a right-handed bend of the peripheral stalk (right). b ATP hydrolysis or synthesis in the F1 region requires three catalytic (black, yellow, and red) and three bind dwell (light blue, blue, and green) conformations. c High-resolution structure of the yeast mitochondrial ATP synthase. d In the absence of free ATP, the peripheral stalk exhibits limited flexibility with a left-handed curvature. e During ATP hydrolysis, ATP synthase can adopt conformations that show a right-handed curvature of the peripheral stalk. f Histograms of the distribution of conformations in the absence of ATP (top, blue) and during ATP hydrolysis (bottom, purple).CC BY · PubMed Central
Fig. 2 Atomic models of ATP synthase under strain during ATP hydrolysis. a Atomic models of the enzyme were generated by flexible fitting of the atomic models of ATP synthase without ATP into maps of the strained ATP synthase conformations. b Overlay of fitted models from catalytic dwell (left) and binding dwell (right) structures show limited conformational changes in the α3β3γδεc10 subcomplex. c Overlay of models from catalytic dwell conformations show pronounced bending of the peripheral stalk while the enzyme is under strain during ATP hydrolysis.
Fig. 2 Atomic models of ATP synthase under strain during ATP hydrolysis. a Atomic models of the enzyme were generated by flexible fitting of the atomic models of ATP synthase without ATP into maps of the strained ATP synthase conformations. b Overlay of fitted models from catalytic dwell (left) and binding dwell (right) structures show limited conformational changes in the α3β3γδεc10 subcomplex. c Overlay of models from catalytic dwell conformations show pronounced bending of the peripheral stalk while the enzyme is under strain during ATP hydrolysis.CC BY · PubMed Central
Fig. 3 Deformation of the peripheral stalk of ATP synthase while under strain during ATP hydrolysis. a Structure of the peripheral stalk shows that subunit h bridges F1 and FO. b Overlay of the eight State 1catalytic atomic models, produced by molecular dynamics flexible fitting of the atomic model of resting ATP synthase into the eight 3D maps of State 1catalytic, shows a large deformation of the peripheral stalk. c The peripheral stalk bends at two pivot points near F1 (yellow arrow) and near FO (red arrow). d Unstructured regions in subunit h (blue asterisks) allow it to withstand the bending of subunit d.
Fig. 3 Deformation of the peripheral stalk of ATP synthase while under strain during ATP hydrolysis. a Structure of the peripheral stalk shows that subunit h bridges F1 and FO. b Overlay of the eight State 1catalytic atomic models, produced by molecular dynamics flexible fitting of the atomic model of resting ATP synthase into the eight 3D maps of State 1catalytic, shows a large deformation of the peripheral stalk. c The peripheral stalk bends at two pivot points near F1 (yellow arrow) and near FO (red arrow). d Unstructured regions in subunit h (blue asterisks) allow it to withstand the bending of subunit d.CC BY · PubMed Central
Fig. 4 Sequence of conformations in the ATP hydrolysis and ATP synthesis cycles. a The unstrained conformation of State 1catalytic shows density consistent with MgADP in the αβtight catalytic pair (left, yellow), suggesting an ADP-inhibited state, while the strained conformations show density consistent with MgATP (right, purple) suggesting an active state. b Measurement of the rotation of the rotor subunit γ within the F1 region shows 36° and 84° sub-steps between catalytic and binding dwell conformations. c Measurement of the rotation of the c-ring relative to subunit a in FO. Within each binding or catalytic dwell conformation the peripheral stalk becomes increasingly strained as the c-ring rotates in the ATP synthesis direction and increasingly relaxed as the c-ring rotates in the ATP hydrolysis direction. Transition between catalytic and binding dwell conformations would require back-stepping of the c-ring (shaded areas) if every conformation occurred during the rotary cycle. Colors in (b) and (c) are defined as in Fig. 1b.
Fig. 4 Sequence of conformations in the ATP hydrolysis and ATP synthesis cycles. a The unstrained conformation of State 1catalytic shows density consistent with MgADP in the αβtight catalytic pair (left, yellow), suggesting an ADP-inhibited state, while the strained conformations show density consistent with MgATP (right, purple) suggesting an active state. b Measurement of the rotation of the rotor subunit γ within the F1 region shows 36° and 84° sub-steps between catalytic and binding dwell conformations. c Measurement of the rotation of the c-ring relative to subunit a in FO. Within each binding or catalytic dwell conformation the peripheral stalk becomes increasingly strained as the c-ring rotates in the ATP synthesis direction and increasingly relaxed as the c-ring rotates in the ATP hydrolysis direction. Transition between catalytic and binding dwell conformations would require back-stepping of the c-ring (shaded areas) if every conformation occurred during the rotary cycle. Colors in (b) and (c) are defined as in Fig. 1b.CC BY · PubMed Central

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Citations

Cites

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Metadata

Authors
Guo, H., & Rubinstein, J. L.
Affiliations
University of Toronto (CA)
Corresponding author
Rubinstein, John L.
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2022-04-25
Volume
13
Issue
1
Pages
2232-2232
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-022-29893-2
PMC
PMC9038767
OpenAlex
W4225253156
Cited by
51
References
2
Citations by year
2026: 6 · 2025: 14 · 2024: 13 · 2023: 10 · 2022: 8
FWCI
3.67
Type
journal article
Language
en
Update notices
correction (2022-05-17)
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
Funders
  • Gouvernement du Canada | Canadian Institutes of Health Research (PJT162186)
  • Canada Research Chairs
  • Canada Foundation for Innovation
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › ATP Synthase and ATPases Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Advanced Electron Microscopy Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
Keywords
ATP synthase, peripheral stalk, mechanical strain, ATP hydrolysis, proton translocation, rotary catalysis, mitochondrial ATP synthase, yeast, cryo-electron microscopy
MeSH terms
Adenosine Triphosphate, Catalysis, Protons, Saccharomyces cerevisiae, Nitric Oxide Synthase, Mitochondrial Proton-Translocating ATPases

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