Mechanism of ATP hydrolysis by beef heart mitochondrial ATPase. Rate constants for elementary steps in catalysis at a single site.

Grubmeyer, Charles; Cross, Richard L.; Penefsky, Harvey S.

doi:10.1016/s0021-9258(18)33683-4

Cited by 401 publications

(89 citation statements)

References 51 publications

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“…ATP hydrolysis in the F 1 can be measured under single site (unisite catalysis) or steady-state (multisite catalysis) conditions: the steady-state ATPase rate is 10 5 -10 6 fold faster than that of the ATP hydrolysis at a single site assayed with an ATP : F 1 ratio of less than 1 : 3 (Cross et al 1982). Further kinetic studies indicated that the equilibrium constant at the catalytic site is near unity, supporting that ATP is synthesized with no energy change (Grubmeyer et al 1982). Three asymmetric catalytic sites were suggested by kinetic studies on wild-type and mutant enzymes, nucleotide binding being detected with an intrinsic tryptophan probe, affinity labelling with ATP analogues or chemical modification with inhibitors (Boyer 1997;Senior et al 2002;Futai et al 2003).…”

Section: Catalysis Transport and Energy Coupling By F-atpasesmentioning

confidence: 94%

Stochastic rotational catalysis of proton pumping F-ATPase

Nakanishi‐Matsui

Futai

2008

Phil. Trans. R. Soc. B

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F-ATPases synthesize ATP from ADP and phosphate coupled with an electrochemical proton gradient in bacterial or mitochondrial membranes and can hydrolyse ATP to form the gradient. F-ATPases consist of a catalytic F 1 and proton channel F 0 formed from the a 3 b 3 gd3 and ab 2 c 10 subunit complexes, respectively. The rotation of g3c 10 couples catalyses and proton transport. Consistent with the threefold symmetry of the a 3 b 3 catalytic hexamer, 1208 stepped revolution has been observed, each step being divided into two substeps. The ATP-dependent revolution exhibited stochastic fluctuation and was driven by conformation transmission of the b subunit (phosphatebinding P-loop/a-helix B/loop/b-sheet4). Recent results regarding mechanically driven ATP synthesis finally proved the role of rotation in energy coupling.

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Section: Catalysis Transport and Energy Coupling By F-atpasesmentioning

confidence: 94%

Stochastic rotational catalysis of proton pumping F-ATPase

Nakanishi‐Matsui

Futai

2008

Phil. Trans. R. Soc. B

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“…5 does not specify the mechanism by which H+-ATP synthases favour the forward reaction over the backward reaction. In this regard, Feldman & Sigman [95] and Grubmeyer et al [96], working with the isolated F1 components of the chloroplast and mitochondrial enzymes respectively, have shown that ATP binds at a catalytic site much more tightly than do ADP and Pi, and that in consequence the equilibrium constant of the ATP synthase reaction on the enzyme is close to unity. Similar findings have been made with de-energized chloroplast lamellae [97] and submitochondrial particles [98].…”

Section: Simulation Of Relationships Between Rate Of Atp Synthesis and Aph+mentioning

confidence: 99%

An investigation of the relationships between rate and driving force in simple uncatalysed and enzyme-catalysed reactions with applications of the findings to chemiosmotic reactions

Stoner

1992

Biochemical Journal

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Both the rate and the driving force of a reaction can be expressed in terms of the concentrations of the reactants and products. Consequently, rate and driving force can be expressed as a function of each other. This has been done for a single-reactant, single-product, uncatalysed reaction and its enzyme-catalysed equivalent using the van't Hoff reaction isotherm and Haldane's generalized Michaelis-Menten rate equation, the primary objective being explanation of the exponential and sigmoidal relationships between reaction rate and delta mu H+ commonly observed in studies on chemiosmotic reactions. Acquisition of a purely thermodynamic rate vs. driving-force relationship requires recognition of the intensive and extensive variables and maintenance of the extensive variables constant. This relationship is identical for the two reactions and is hyperbolic or sigmoidal, depending on whether the equilibrium constant is smaller or larger than unity. In the case of the catalysed reaction, acquisition of the purely thermodynamic relationship requires the assumption that the enzyme be equally effective in catalysing the forward and backward reactions. If this condition is not met, the relationship is modified by the enzyme in a manner which can be determined from the ratio of the Michaelis constants of the reactant and product. Under conditions of enzyme saturation in respect to reactant+product, the rate vs. driving-force relationship is determined exclusively by the thermodynamics of the reaction and a single kinetic parameter, the magnitude of which is determined by the relative effectiveness of the enzyme in catalysing the forward and backward reactions. In view of this finding, it is pointed out that, since the catalytic components of chemiosmotic reactions appear to be saturated with respect to the reactant-product pair that is varied in experimental rate vs. delta mu H+ determinations, and that, since many complex enzymic reactions conform to the simple Michaelis-Menten equation with respect to a single reactant-product pair when the concentrations of all other reactants and products are maintained constant, one might expect to be capable of simulating the experimental relationships simply from knowledge of the thermodynamics of the reaction and the relative effectiveness of the catalytic component in catalysing the forward and backward reactions using the simple Michaelis-Menten equation. That this expectation appears to be largely correct is demonstrated with model reactions.(ABSTRACT TRUNCATED AT 400 WORDS)

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“…We focused on two typical reaction schemes, one suggested recently for mitochondrial F 1 (as MF 1 ) [18], and the other determined for bacterial F 1 (as BF 1 ) [19]. For an individual ATPase site, formed by one α-β hetero-dimer, the uni-site reaction rates for the product ADP and Pi releases are particularly low [20,21]. To allow sufficiently fast reactions as that achieved on the tri-site F 1 ring, it is necessary that the inter-subunit couplings significantly accelerate the respective product releases, but how that happen is elusive.…”

Section: Introductionmentioning

confidence: 99%

Deciphering Intrinsic Inter-subunit Couplings that Lead to Sequential Hydrolysis of F 1 -ATPase Ring

Dai

Flechsig

Jin

2017

Biophysical Journal

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Rotary sequential hydrolysis of the metabolic machine F1-ATPase is a prominent manifestation of high coordination among multiple chemical sites in ring-shaped molecular machines, and it is also functionally essential for F1 to tightly couple chemical reactions and central γ-shaft rotation. High-speed AFM experiments have identified that sequential hydrolysis is maintained in the F1 stator ring even in the absence of the γ-rotor. To explore the origins of intrinsic sequential performance, we computationally investigated essential inter-subunit couplings on the hexameric ring of mitochondrial and bacterial F1. We first reproduced in stochastic Monte Carlo simulations the experimentally determined sequential hydrolysis schemes by kinetically imposing inter-subunit couplings and following subsequent tri-site ATP hydrolysis cycles on the F1 ring. We found that the key couplings to support the sequential hydrolysis are those that accelerate neighbor-site ADP and Pi release upon a certain ATP binding or hydrolysis reaction. The kinetically identified couplings were then examined in atomistic molecular dynamics simulations at a coarse-grained level to reveal the underlying structural mechanisms. To do that, we enforced targeted conformational changes of ATP binding or hydrolysis to one chemical site on the F1 ring and monitored the ensuing conformational responses of the neighboring sites using structure-based simulations. Notably, we found asymmetrical neighbor-site opening that facilitates ADP release upon enforced ATP binding. We also captured a complete charge-hopping process of the Pi release subsequent to enforced ATP hydrolysis in the neighbor site, confirming recent single-molecule analyses with regard to the role of ATP hydrolysis in F1. Our studies therefore elucidate both the coordinated chemical kinetics and structural dynamics mechanisms underpinning the sequential operation of the F1 ring.

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Mechanism of ATP hydrolysis by beef heart mitochondrial ATPase. Rate constants for elementary steps in catalysis at a single site.

Cited by 401 publications

References 51 publications

Stochastic rotational catalysis of proton pumping F-ATPase

Stochastic rotational catalysis of proton pumping F-ATPase

An investigation of the relationships between rate and driving force in simple uncatalysed and enzyme-catalysed reactions with applications of the findings to chemiosmotic reactions

Deciphering Intrinsic Inter-subunit Couplings that Lead to Sequential Hydrolysis of F 1 -ATPase Ring

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