scholarly journals Kinetic Analysis Of ATP Synthesis Catalyzed By E. coli FoF1 ATP Synthase Reconstituted Into Egg Yolk Liposomes: Evidence For Bi-site Activation

2009 ◽  
Vol 96 (3) ◽  
pp. 142a
Author(s):  
Mikhail Galkin ◽  
Robert K. Nakamoto
Keyword(s):  
Kinetic Analysis ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Egg Yolk ◽  
E Coli ◽  
Fof1 Atp Synthase

Assembly of the Escherichia coli FoF1 ATP synthase involves distinct subcomplex formation

2013 ◽  
Vol 41 (5) ◽  
pp. 1288-1293 ◽  
Author(s):  
Gabriele Deckers-Hebestreit
Keyword(s):  
Escherichia Coli ◽  
Atp Synthase ◽  
Cytoplasmic Membrane ◽  
Atp Synthesis ◽  
E Coli ◽  
Assembly Pathway ◽  
Polypeptide Chains ◽  
Fof1 Atp Synthase

The ATP synthase (FoF1) of Escherichia coli couples the translocation of protons across the cytoplasmic membrane by Fo to ATP synthesis or hydrolysis in F1. Whereas good knowledge of the nanostructure and the rotary mechanism of the ATP synthase is at hand, the assembly pathway of the 22 polypeptide chains present in a stoichiometry of ab2c10α3β3γδϵ has so far not received sufficient attention. In our studies, mutants that synthesize different sets of FoF1 subunits allowed the characterization of individually formed stable subcomplexes. Furthermore, the development of a time-delayed in vivo assembly system enabled the subsequent synthesis of particular missing subunits to allow the formation of functional ATP synthase complexes. These observations form the basis for a model that describes the assembly pathway of the E. coli ATP synthase from pre-formed subcomplexes, thereby avoiding membrane proton permeability by a concomitant assembly of the open H+-translocating unit within a coupled FoF1 complex.

scholarly journals The structure of the catalytic domain of the ATP synthase fromMycobacterium smegmatisis a target for developing antitubercular drugs

2019 ◽  
Vol 116 (10) ◽  
pp. 4206-4211 ◽  
Author(s):  
Alice Tianbu Zhang ◽  
Martin G. Montgomery ◽  
Andrew G. W. Leslie ◽  
Gregory M. Cook ◽  
John E. Walker
Keyword(s):  
Atp Synthase ◽  
Mycobacterium Smegmatis ◽  
Catalytic Domain ◽  
Atp Hydrolysis ◽  
Atp Synthesis ◽  
Geobacillus Stearothermophilus ◽  
Antitubercular Drugs ◽  
Atp Binding Site ◽  
E Coli ◽  
Catalytic Cycles

The crystal structure of the F1-catalytic domain of the adenosine triphosphate (ATP) synthase has been determined fromMycobacterium smegmatiswhich hydrolyzes ATP very poorly. The structure of the α3β3-component of the catalytic domain is similar to those in active F1-ATPases inEscherichia coliandGeobacillus stearothermophilus. However, its ε-subunit differs from those in these two active bacterial F1-ATPases as an ATP molecule is not bound to the two α-helices forming its C-terminal domain, probably because they are shorter than those in active enzymes and they lack an amino acid that contributes to the ATP binding site in active enzymes. InE. coliandG. stearothermophilus, the α-helices adopt an “up” state where the α-helices enter the α3β3-domain and prevent the rotor from turning. The mycobacterial F1-ATPase is most similar to the F1-ATPase fromCaldalkalibacillus thermarum, which also hydrolyzes ATP poorly. The βE-subunits in both enzymes are in the usual “open” conformation but appear to be occupied uniquely by the combination of an adenosine 5′-diphosphate molecule with no magnesium ion plus phosphate. This occupation is consistent with the finding that their rotors have been arrested at the same point in their rotary catalytic cycles. These bound hydrolytic products are probably the basis of the inhibition of ATP hydrolysis. It can be envisaged that specific as yet unidentified small molecules might bind to the F1domain inMycobacterium tuberculosis, prevent ATP synthesis, and inhibit the growth of the pathogen.

scholarly journals Direct observation of stepped proteolipid ring rotation in E. coli FoF1-ATP synthase

2010 ◽  
Vol 29 (23) ◽  
pp. 3911-3923 ◽  
Author(s):  
Robert Ishmukhametov ◽  
Tassilo Hornung ◽  
David Spetzler ◽  
Wayne D Frasch
Keyword(s):  
Atp Synthase ◽  
Direct Observation ◽  
E Coli ◽  
Ring Rotation ◽  
Fof1 Atp Synthase

scholarly journals Dye-ligand chromatographic purification of intact multisubunit membrane protein complexes: application to the chloroplast H+-F0F1-ATP synthase

2000 ◽  
Vol 346 (1) ◽  
pp. 41-44
Author(s):  
Holger SEELERT ◽  
Ansgar POETSCH ◽  
Meino ROHLFS ◽  
Norbert A. DENCHER
Keyword(s):  
Membrane Protein ◽  
Phosphatidic Acid ◽  
Ammonium Sulphate ◽  
Atp Synthase ◽  
Protein Complex ◽  
Protein Complexes ◽  
Atp Synthesis ◽  
Chromatographic Purification ◽  
Membrane Protein Complexes ◽  
Fof1 Atp Synthase

n-Dodecyl-β-D-maltoside was used as a detergent to solubilize the ammonium sulphate precipitate of chloroplast FOF1-ATP synthase, which was purified further by dye-ligand chromatography. Upon reconstitution of the purified protein complex into phosphatidylcholine/phosphatidic acid liposomes, ATP synthesis, driven by an artificial ∆pH/∆ψ, was observed. The highest activity was achieved with ATP synthase solubilized in n-dodecyl-β-D-maltoside followed by chromatography with Red 120 dye. The optimal dye for purification with CHAPS was Green 5. All known subunits were present in the monodisperse proton-translocating ATP synthase preparation obtained from chloroplasts.

scholarly journals Perfect chemomechanical coupling of FoF1-ATP synthase

2017 ◽  
Vol 114 (19) ◽  
pp. 4960-4965 ◽  
Author(s):  
Naoki Soga ◽  
Kazuya Kimura ◽  
Kazuhiko Kinosita ◽  
Masasuke Yoshida ◽  
Toshiharu Suzuki
Keyword(s):  
Atp Synthase ◽  
Chemical Potential ◽  
Initial Rate ◽  
Coupling Efficiency ◽  
Atp Synthesis ◽  
Energy Yield ◽  
Coupling Mechanism ◽  
Chemomechanical Coupling ◽  
Base Transition ◽  
Fof1 Atp Synthase

FoF1-ATP synthase (FoF1) couples H+ flow in Fo domain and ATP synthesis/hydrolysis in F1 domain through rotation of the central rotor shaft, and the H+/ATP ratio is crucial to understand the coupling mechanism and energy yield in cells. Although H+/ATP ratio of the perfectly coupling enzyme can be predicted from the copy number of catalytic β subunits and that of H+ binding c subunits as c/β, the actual H+/ATP ratio can vary depending on coupling efficiency. Here, we report actual H+/ATP ratio of thermophilic Bacillus FoF1, whose c/β is 10/3. Proteoliposomes reconstituted with the FoF1 were energized with ΔpH and Δψ by the acid−base transition and by valinomycin-mediated diffusion potential of K+ under various [ATP]/([ADP]⋅[Pi]) conditions, and the initial rate of ATP synthesis/hydrolysis was measured. Analyses of thermodynamically equilibrated states, where net ATP synthesis/hydrolysis is zero, show linear correlation between the chemical potential of ATP synthesis/hydrolysis and the proton motive force, giving the slope of the linear function, that is, H+/ATP ratio, 3.3 ± 0.1. This value agrees well with the c/β ratio. Thus, chemomechanical coupling between Fo and F1 is perfect.

scholarly journals The phototroph-specific β-hairpin structure of the γ subunit of FoF1-ATP synthase is important for efficient ATP-synthesis of cyanobacteria

2021 ◽  
pp. 101027
Author(s):  
Kumiko Kondo ◽  
Masayuki Izumi ◽  
Kosuke Inabe ◽  
Keisuke Yoshida ◽  
Mari Imashimizu ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Atp Synthesis ◽  
Hairpin Structure ◽  
Γ Subunit ◽  
Fof1 Atp Synthase

scholarly journals Cooperation among c-subunits of FoF1-ATP synthase in rotation-coupled proton translocation

2021 ◽  
Author(s):  
Noriyo Mitome ◽  
Shintaroh Kubo ◽  
Sumie Ohta ◽  
Hikaru Takashima ◽  
Yuto Shigefuji ◽  
...  
Keyword(s):  
Atp Synthase ◽  
Proton Pump ◽  
Proton Translocation ◽  
Atp Synthesis ◽  
Wild Type ◽  
Single Chain ◽  
Proton Release ◽  
Biochemical Assays ◽  
Proton Uptake ◽  
Fof1 Atp Synthase

In FoF1-ATP synthase, proton translocation through Fo drives rotation of the c-subunit oligomeric ring relative to the a-subunit. Recent studies suggest that in each step of the rotation, key glutamic acid residues in different c-subunits contribute to proton release to and proton uptake from the a-subunit. However, no studies have demonstrated cooperativity among c-subunits toward FoF1-ATP synthase activity. Here, we addressed this using Bacillus PS3 ATP synthase harboring c-ring with various combinations of wild-type and cE56D, enabled by genetically fused single-chain c-ring. ATP synthesis and proton pump activities were significantly decreased by a single cE56D mutation and further decreased by double cE56D mutations. Moreover, activity further decreased as the two mutation sites were separated, indicating cooperation among c-subunits. Similar results were obtained for proton transfer-coupled molecular simulations. Simulations revealed that prolonged proton uptake in mutated c-subunits is shared between two c-subunits, explaining the cooperation observed in biochemical assays.

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