scholarly journals Rotary substates of mitochondrial ATP synthase reveal the basis of flexible F1-Focoupling

Science ◽  
2019 ◽  
Vol 364 (6446) ◽  
pp. eaaw9128 ◽  
Author(s):  
Bonnie J. Murphy ◽  
Niklas Klusch ◽  
Julian Langer ◽  
Deryck J. Mills ◽  
Özkan Yildiz ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Metal Ion ◽  
Three Dimensional ◽  
Atp Synthesis ◽  
Access Channel ◽  
Dimensional Classification ◽  
Ring Rotation ◽  
Mitochondrial Atp Synthase ◽  
Central Stalk ◽  
Energy Consuming

F1Fo–adenosine triphosphate (ATP) synthases make the energy of the proton-motive force available for energy-consuming processes in the cell. We determined the single-particle cryo–electron microscopy structure of active dimeric ATP synthase from mitochondria ofPolytomellasp. at a resolution of 2.7 to 2.8 angstroms. Separation of 13 well-defined rotary substates by three-dimensional classification provides a detailed picture of the molecular motions that accompanyc-ring rotation and result in ATP synthesis. Crucially, the F1head rotates along with the central stalk andc-ring rotor for the first ~30° of each 120° primary rotary step to facilitate flexible coupling of the stoichiometrically mismatched F1and Fosubcomplexes. Flexibility is mediated primarily by the interdomain hinge of the conserved OSCP subunit. A conserved metal ion in the proton access channel may synchronizec-ring protonation with rotation.

scholarly journals Rotary substates of mitochondrial ATP synthase reveal the basis of flexible F1-Fo coupling

2019 ◽  
Author(s):  
Bonnie J. Murphy ◽  
Niklas Klusch ◽  
Julian D. Langer ◽  
Deryck J. Mills ◽  
Özkan Yildiz ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Metal Ion ◽  
De Novo ◽  
Atp Synthesis ◽  
Joint Movement ◽  
Access Channel ◽  
Subunit Interface ◽  
Ring Rotation ◽  
Mitochondrial Atp Synthase ◽  
Ordered Water

F1Fo-ATP synthases play a central role in cellular metabolism, making the energy of the proton-motive force across a membrane available for a large number of energy-consuming processes. We determined the single-particle cryo-EM structure of active dimeric ATP synthase from mitochondria of Polytomella sp. at 2.7- 2.8 Å resolution. Separation of 13 well-defined rotary substates by 3D classification provides a detailed picture of the molecular motions that accompany c-ring rotation and result in ATP synthesis. Crucially, the F1 head rotates along with the central stalk and c-ring rotor for the first ~30° of each 120° primary rotary step. The joint movement facilitates flexible coupling of the stoichiometrically mismatched F1 and Fo subcomplexes. Flexibility is mediated primarily by the interdomain hinge of the conserved OSCP subunit, a well-established target of physiologically important inhibitors. Our maps provide atomic detail of the c-ring/a-subunit interface in the membrane, where protonation and deprotonation of c-ring cGlu111 drives rotary catalysis. An essential histidine residue in the lumenal proton access channel binds a strong non-peptide density assigned to a metal ion that may facilitate c-ring protonation, as its coordination geometry changes with c-ring rotation. We resolve ordered water molecules in the proton access and release channels and at the gating aArg239 that is critical in all rotary ATPases. We identify the previously unknown ASA10 subunit and present complete de novo atomic models of subunits ASA1-10, which make up the two interlinked peripheral stalks that stabilize the Polytomella ATP synthase dimer.

scholarly journals A critical appraisal of evidence for localized energy coupling. Kinetic studies on liposomes containing bacteriorhodopsin and ATP synthase

1985 ◽  
Vol 230 (2) ◽  
pp. 543-549 ◽  
Author(s):  
R L Van der Bend ◽  
J Petersen ◽  
J A Berden ◽  
K Van Dam ◽  
H V Westerhoff
Keyword(s):  
Electron Transfer ◽  
Atp Synthase ◽  
Critical Appraisal ◽  
Atp Synthesis ◽  
Kinetic Studies ◽  
Energy Coupling ◽  
Proton Pumps ◽  
Specific Interactions ◽  
Mitochondrial Atp Synthase ◽  
Different Levels

In intact systems (chloroplasts, mitochondria and bacteria) many experiments have been reported which are indicative of localized coupling between ATP synthase and electron transfer complexes. We have carried out similar experiments with a system in which we may assume that specific interactions between the proton pumps are absent: reconstituted vesicles containing bacteriorhodopsin and yeast mitochondrial ATP synthase. The only experiment that gives results which differ from those previously published for intact systems concerns the effect of uncouplers on the rate of ATP synthesis at different levels of inhibition of the ATP synthase. We propose that this type of experiment may discriminate between localized and delocalized coupling.

Phosphate exchange and ATP synthesis by DMSO-pretreated purified bovine mitochondrial ATP synthase

2001 ◽  
Vol 353 (2) ◽  
pp. 215-222
Author(s):  
Seelochan BEHARRY ◽  
Philip D. BRAGG
Keyword(s):  
Exchange Reaction ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Catalytic Site ◽  
Phosphate Removal ◽  
Subsequent Removal ◽  
F1fo Atp Synthase ◽  
Mitochondrial Atp Synthase ◽  
Phosphate Exchange

Purified soluble bovine mitochondrial F1Fo-ATP synthase contained 2mol of ATP, 2mol of ADP and 6mol of Pi/mol. Incubation of this enzyme with 1mM [32P]Pi caused the exchange of 2mol of Pi/mol of F1Fo-ATP synthase. The labelled phosphates were not displaced by ATP. Transfer of F1Fo-ATP synthase to a buffer containing 30% (v/v) DMSO and 1mM [32P]Pi resulted in the loss of bound nucleotides with the retention of 1mol of ATP/mol of F1Fo-ATP synthase. Six molecules of [32P]Pi were incorporated by exchange with the existing bound phosphate. Removal of the DMSO by passage of the enzyme through a centrifuged column of Sephadex G-50 resulted in the exchange of one molecule of bound [32P]Pi into the bound ATP. Azide did not prevent this [32P]Pi ↔ ATP exchange reaction. The bound labelled ATP could be displaced from the enzyme by exogenous ATP. Addition of ADP to the DMSO-pretreated F1Fo-ATP synthase in the original DMSO-free buffer resulted in the formation of an additional molecule of bound ATP. It was concluded that following pretreatment with and subsequent removal of DMSO the F1Fo-ATP synthase contained one molecule of ATP at a catalytic site which was competent to carry out a phosphateŐATP exchange reaction using enzyme-bound inorganic radiolabelled phosphate. In the presence of ADP an additional molecule of labelled ATP was formed from enzyme-bound Pi at a second catalytic site. The bound phosphateŐATP exchange reaction is not readily accommodated by current mechanisms for the ATP synthase.

scholarly journals Re-Understanding the Mechanisms of Action of the Anti-Mycobacterial Drug Bedaquiline

Antibiotics ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 261 ◽  
Author(s):  
Jickky Palmae Sarathy ◽  
Gerhard Gruber ◽  
Thomas Dick
Keyword(s):  
Electron Transport ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Mechanisms Of Action ◽  
Direct Mechanism ◽  
Transport Chain ◽  
Ring Rotation ◽  
New Findings ◽  
Atp Generation ◽  
Resistance Mutants

Bedaquiline (BDQ) inhibits ATP generation in Mycobacterium tuberculosis by interfering with the F-ATP synthase activity. Two mechanisms of action of BDQ are broadly accepted. A direct mechanism involves BDQ binding to the enzyme’s c-ring to block its rotation, thus inhibiting ATP synthesis in the enzyme’s catalytic α3β3-headpiece. An indirect mechanism involves BDQ uncoupling electron transport in the electron transport chain from ATP synthesis at the F-ATP synthase. In a recently uncovered second direct mechanism, BDQ binds to the enzyme’s ε-subunit to disrupt its ability to link c-ring rotation to ATP synthesis at the α3β3-headpiece. However, this mechanism is controversial as the drug’s binding affinity for the isolated ε-subunit protein is moderate and spontaneous resistance mutants in the ε-subunit cannot be isolated. Recently, the new, structurally distinct BDQ analogue TBAJ-876 was utilized as a chemical probe to revisit BDQ’s mechanisms of action. In this review, we first summarize discoveries on BDQ’s mechanisms of action and then describe the new insights derived from the studies of TBAJ-876. The TBAJ-876 investigations confirm the c-ring as a target, while also supporting a functional role for targeting the ε-subunit. Surprisingly, the new findings suggest that the uncoupler mechanism does not play a key role in BDQ’s anti-mycobacterial activity.

scholarly journals Structure and conformational states of the bovine mitochondrial ATP synthase by cryo-EM

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Anna Zhou ◽  
Alexis Rohou ◽  
Daniel G Schep ◽  
John V Bason ◽  
Martin G Montgomery ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Catalytic Mechanism ◽  
Proton Translocation ◽  
Atp Synthesis ◽  
Bioinformatic Analysis ◽  
Mitochondrial Atp Synthase ◽  
A Subunit ◽  
Classification Of Images ◽  
Atp Synthases

Adenosine triphosphate (ATP), the chemical energy currency of biology, is synthesized in eukaryotic cells primarily by the mitochondrial ATP synthase. ATP synthases operate by a rotary catalytic mechanism where proton translocation through the membrane-inserted FO region is coupled to ATP synthesis in the catalytic F1 region via rotation of a central rotor subcomplex. We report here single particle electron cryomicroscopy (cryo-EM) analysis of the bovine mitochondrial ATP synthase. Combining cryo-EM data with bioinformatic analysis allowed us to determine the fold of the a subunit, suggesting a proton translocation path through the FO region that involves both the a and b subunits. 3D classification of images revealed seven distinct states of the enzyme that show different modes of bending and twisting in the intact ATP synthase. Rotational fluctuations of the c8-ring within the FO region support a Brownian ratchet mechanism for proton-translocation-driven rotation in ATP synthases.

scholarly journals IF1, a natural inhibitor of mitochondrial ATP synthase, is not essential for the normal growth and breeding of mice

2013 ◽  
Vol 33 (5) ◽  
Author(s):  
Junji Nakamura ◽  
Makoto Fujikawa ◽  
Masasuke Yoshida
Keyword(s):  
Atp Synthase ◽  
Mouse Strain ◽  
Reactive Oxygen Species Generation ◽  
Atp Synthesis ◽  
Normal Growth ◽  
Test Results ◽  
Mitochondrial Atp Synthase ◽  
Evolutionarily Conserved ◽  
Natural Inhibitor ◽  
Synthesis Activity

IF1 is an endogenous inhibitor protein of mitochondrial ATP synthase. It is evolutionarily conserved throughout all eukaryotes and it has been proposed to play crucial roles in prevention of the wasteful reverse reaction of ATP synthase, in the metabolic shift from oxidative phosphorylation to glycolysis, in the suppression of ROS (reactive oxygen species) generation, in mitochondria morphology and in haem biosynthesis in mitochondria, which leads to anaemia. Here, we report the phenotype of a mouse strain in which IF1 gene was destroyed. Unexpectedly, individuals of this IF1-KO (knockout) mouse strain grew and bred without defect. The general behaviours, blood test results and responses to starvation of the IF1-KO mice were apparently normal. There were no abnormalities in the tissue anatomy or the autophagy. Mitochondria of the IF1-KO mice were normal in morphology, in the content of ATP synthase molecules and in ATP synthesis activity. Thus, IF1 is not an essential protein for mice despite its ubiquitous presence in eukaryotes.

scholarly journals An Algal Nucleus-encoded Subunit of Mitochondrial ATP Synthase Rescues a Defect in the Analogous Human Mitochondrial-encoded Subunit

2002 ◽  
Vol 13 (11) ◽  
pp. 3836-3844 ◽  
Author(s):  
Joseline Ojaimi ◽  
Junmin Pan ◽  
Sumana Santra ◽  
William J. Snell ◽  
Eric A. Schon
Keyword(s):  
Atp Synthase ◽  
Nuclear Gene ◽  
Mitochondrial Diseases ◽  
Atp Synthesis ◽  
Single Copy ◽  
Human Cells ◽  
Mitochondrial Targeting ◽  
Mitochondrial Atp Synthase ◽  
Targeting Signal ◽  
Atpase 6

Unlike most organisms, the mitochondrial DNA (mtDNA) ofChlamydomonas reinhardtii, a green alga, does not encode subunit 6 of F0F1-ATP synthase. We hypothesized that C. reinhardtii ATPase 6 is nucleus encoded and identified cDNAs and a single-copy nuclear gene specifying this subunit (CrATP6, with eight exons, four of which encode a mitochondrial targeting signal). Although the algal and humanATP6 genes are in different subcellular compartments and the encoded polypeptides are highly diverged, their secondary structures are remarkably similar. When CrATP6 was expressed in human cells, a significant amount of the precursor polypeptide was targeted to mitochondria, the mitochondrial targeting signal was cleaved within the organelle, and the mature polypeptide was assembled into human ATP synthase. In spite of the evolutionary distance between algae and mammals, C. reinhardtii ATPase 6 functioned in human cells, because deficiencies in both cell viability and ATP synthesis in transmitochondrial cell lines harboring a pathogenic mutation in the human mtDNA-encoded ATP6 gene were overcome by expression of CrATP6. The ability to express a nucleus-encoded version of a mammalian mtDNA-encoded protein may provide a way to import other highly hydrophobic proteins into mitochondria and could serve as the basis for a gene therapy approach to treat human mitochondrial diseases.

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