Modification of the mitochondrial F1-ATPase ∊ subunit, enhancement of the ATPase activity of the IF1–F1 complex and IF1-binding dependence of the conformation of the ∊ subunit

Treatment of bovine heart submitochondrial particles with a low concentration of 2-hydroxy-5-nitrobenzyl bromide (HNB), a selective reagent for the Trp residue of the ε subunit [Baracca, Barogi, Lenaz and Solaini (1993) Int. J. Biochem. 25, 1269-1275], enhances the ATP hydrolytic activity of the particles exclusively when the natural inhibitor protein IF1 is present. Similarly, isolated F1 [the catalytic sector of the mitochondrial H+-ATPase complex (ATP synthase)] treated with the reagent has the ATPase activity enhanced exclusively if IF1 is bound to it. These experiments suggest that the modification of the ε subunit decreases the inhibitory activity of IF1, eliciting the search for a relationship between the ε subunit and the inhibitory protein. Certainly, a reverse relationship exists because HNB binds covalently to the isolated F1 exclusively when the inhibitory protein is present. This finding is consistent with the existence of the ε subunit in different conformational states depending on whether IF1 is bound to F1 or not. Support for this assertion is obtained by measurements of the intrinsic phosphorescence decay rate of F1, a probe of the Trp ε subunit conformation in situ [Solaini, Baracca, Parenti-Castelli and Strambini (1993) Eur. J. Biochem. 214, 729-734]. A significant difference in phosphorescence decay rate is detected when IF1 is added to preparations of F1 previously devoid of the inhibitory protein. These studies indicate that IF1 and the ε subunit of the mitochondrial F1-ATPase complex are related, suggesting a possible role of the ε subunit in the mechanism of regulation of the mitochondrial ATP synthase.

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Deletion of the natural inhibitory protein Inh1 in Ustilago maydis has no effect on the dimeric state of the F1FO-ATP synthase but increases the ATPase activity and reduces the stability

Biochimica et Biophysica Acta (BBA) - Bioenergetics ◽

10.1016/j.bbabio.2021.148429 ◽

2021 ◽

Vol 1862 (7) ◽

pp. 148429

Author(s):

Romero-Aguilar Lucero ◽

Esparza-Perusquía Mercedes ◽

Langner Thorsten ◽

García-Cruz Giovanni ◽

Feldbrügge Michael ◽

...

Keyword(s):

Atpase Activity ◽

Atp Synthase ◽

Ustilago Maydis ◽

Inhibitory Protein ◽

F1fo Atp Synthase ◽

The Stability

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Expression of Bovine F1-ATPase with Functional Complementation in Yeast Saccharomyces cerevisiae

Journal of Biological Chemistry ◽

10.1074/jbc.m411113200 ◽

2005 ◽

Vol 280 (23) ◽

pp. 22418-22424 ◽

Cited By ~ 8

Author(s):

Neeti Puri ◽

Jie Lai-Zhang ◽

Scott Meier ◽

David M. Mueller

Keyword(s):

Saccharomyces Cerevisiae ◽

Molecular Mass ◽

Atp Synthase ◽

Specific Activity ◽

Functional Complementation ◽

Yeast Saccharomyces Cerevisiae ◽

F1 Atpase ◽

Mitochondrial Atp Synthase ◽

Genes Encoding ◽

Molecular Machinery

The mitochondrial F1F0-ATP synthase is a multimeric enzyme complex composed of at least 16 unique peptides with an overall molecular mass of ∼600 kDa. F1-ATPase is composed of α3β3γδϵ with an overall molecular mass of 370 kDa. The genes encoding bovine F1-ATPase have been expressed in a quintuple yeast Saccharomyces cerevisiae deletion mutant (ΔαΔβΔγΔδΔϵ). This strain expressing bovine F1 is unable to grow on medium containing a non-fermentable carbon source (YPG), indicating that the enzyme is non-functional. However, daughter strains were easily selected for growth on YPG medium and these were evolved for improved growth on YPG medium. The evolution of the strains was presumably due to mutations, but mutations in the genes encoding the subunits of the bovine F1-ATPase were not required for the ability of the cell to grow on YPG medium. The bovine enzyme expressed in yeast was partially purified to a specific activity of about half of that of the enzyme purified from bovine heart mitochondria. These results indicate that the molecular machinery required for the assembly of the mitochondrial ATP synthase is conserved from bovine and yeast and suggest that yeast may be useful for the expression, mutagenesis, and analysis of the mammalian F1- or F1F0-ATP synthase.

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Inhibitors of the catalytic domain of mitochondrial ATP synthase

Biochemical Society Transactions ◽

10.1042/bst0340989 ◽

2006 ◽

Vol 34 (5) ◽

pp. 989-992 ◽

Cited By ~ 17

Author(s):

J.R. Gledhill ◽

J.E. Walker

Keyword(s):

Atp Synthase ◽

Binding Sites ◽

Catalytic Domain ◽

Structural Studies ◽

Inhibitor Protein ◽

X Ray ◽

F1 Atpase ◽

X Ray Crystallography ◽

Mitochondrial Atp Synthase ◽

Natural Inhibitor

An understanding of the mechanism of ATP synthase requires an explanation of how inhibitors act. The catalytic F1-ATPase domain of the enzyme has been studied extensively by X-ray crystallography in a variety of inhibited states. Four independent inhibitory sites have been identified by high-resolution structural studies. They are the catalytic site, and the binding sites for the antibiotics aurovertin and efrapeptin and for the natural inhibitor protein, IF1.

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Inverse regulation of F1-ATPase activity by a mutation at the regulatory region on the γ subunit of chloroplast ATP synthase

Biochemical Journal ◽

10.1042/0264-6021:3520783 ◽

2000 ◽

Vol 352 (3) ◽

pp. 783 ◽

Cited By ~ 11

Author(s):

Hiroki KONNO ◽

Masahide YODOGAWA ◽

Michael T. STUMPP ◽

Peter KROTH ◽

Heinrich STROTMANN ◽

...

Keyword(s):

Atpase Activity ◽

Atp Synthase ◽

Regulatory Region ◽

F1 Atpase ◽

Chloroplast Atp Synthase ◽

Γ Subunit

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Structure of F1-ATPase from the obligate anaerobeFusobacterium nucleatum

Open Biology ◽

10.1098/rsob.190066 ◽

2019 ◽

Vol 9 (6) ◽

pp. 190066 ◽

Cited By ~ 1

Author(s):

Jessica Petri ◽

Yoshio Nakatani ◽

Martin G. Montgomery ◽

Scott A. Ferguson ◽

David Aragão ◽

...

Keyword(s):

Crystal Structure ◽

Adenosine Triphosphate ◽

Atp Synthase ◽

Mycobacterium Smegmatis ◽

Anaerobic Bacterium ◽

Catalytic Domain ◽

Atp Hydrolysis ◽

Fusobacterium Nucleatum ◽

Hydrolytic Activity ◽

F1 Atpase

The crystal structure of the F1-catalytic domain of the adenosine triphosphate (ATP) synthase has been determined from the pathogenic anaerobic bacteriumFusobacterium nucleatum. The enzyme can hydrolyse ATP but is partially inhibited. The structure is similar to those of the F1-ATPases fromCaldalkalibacillus thermarum, which is more strongly inhibited in ATP hydrolysis, and inMycobacterium smegmatis, which has a very low ATP hydrolytic activity. The βE-subunits in all three enzymes are in the conventional ‘open’ state, and in the case ofC. thermarumandM. smegmatis, they are occupied by an ADP and phosphate (or sulfate), but inF. nucleatum, the occupancy by ADP appears to be partial. It is likely that the hydrolytic activity of theF. nucleatumenzyme is regulated by the concentration of ADP, as in mitochondria.

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Neuroprotective Effect of a Novel ATP-Synthase Inhibitor Bedaquiline in Cerebral Ischemia-Reperfusion Injury

International Journal of Molecular Sciences ◽

10.3390/ijms22189717 ◽

2021 ◽

Vol 22 (18) ◽

pp. 9717

Author(s):

Danielius Umbrasas ◽

Odeta Arandarcikaite ◽

Ramune Grigaleviciute ◽

Rimantas Stakauskas ◽

Vilmante Borutaite

Keyword(s):

Atpase Activity ◽

Atp Synthase ◽

Mitochondrial Respiration ◽

Ischemia Reperfusion Injury ◽

Neuroprotective Effect ◽

Ischemia Reperfusion ◽

Atp Depletion ◽

Neurological Dysfunction ◽

Mitochondrial Atp Synthase ◽

Mitochondrial Functions

Mitochondrial dysfunction during ischemic stroke ultimately manifests as ATP depletion. Mitochondrial ATP synthase upon loss of mitochondrial membrane potential during ischemia rapidly hydrolyses ATP and thus contributes to ATP depletion. Increasing evidence suggests that inhibition of ATP synthase limits ATP depletion and is protective against ischemic tissue damage. Bedaquiline (BDQ) is an anti-microbial agent, approved for clinical use, that inhibits ATP synthase of Mycobacteria; however recently it has been shown to act on mitochondrial ATP synthase, inhibiting both ATP synthesis and hydrolysis in low micromolar concentrations. In this study, we investigated whether preconditioning with BDQ can alleviate ischemia/reperfusion-induced brain injury in Wistar rats after middle cerebral artery occlusion-reperfusion and whether it affects mitochondrial functions. We found that BDQ was effective in limiting necrosis and neurological dysfunction during ischemia-reperfusion. BDQ also caused inhibition of ATPase activity, mild uncoupling of respiration, and stimulated mitochondrial respiration both in healthy and ischemic mitochondria. Mitochondrial calcium retention capacity was unaffected by BDQ preconditioning. We concluded that BDQ has neuroprotective properties associated with its action on mitochondrial respiration and ATPase activity.

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