High-resolution structure and mechanism of an F/V-hybrid rotor ring in a Na+-coupled ATP synthase

Matthies, Doreen; Zhou, Wenchang; Klyszejko, Adriana L.; Anselmi, Claudio; Yıldız, Özkan; Brandt, Karsten; Müller, Volker; Faraldo‐Gómez, José D.; Meier, Thomas

doi:10.1038/ncomms6286

Cited by 71 publications

(85 citation statements)

References 66 publications

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“…The functional ion-binding sites in the c-subunit rings of all rotary ATPases are located along the outer circumference of the structure, approximately halfway across the transmembrane region (21,23,(31)(32)(33)(34)(35). The amino acid composition of these binding sites varies across species, but invariably they feature at least one carboxylic side chain (most commonly glutamate).…”

Section: Resultsmentioning

confidence: 99%

“…The amino acid composition of these binding sites varies across species, but invariably they feature at least one carboxylic side chain (most commonly glutamate). In rotary ATPases that are physiologically coupled to the H + gradient, this conserved side chain enables H + binding to the c-ring through protonation (36,37); in enzymes coupled to Na + , as in I. tartaricus, the carboxylate group coordinates the bound Na + directly (21,33,35). Carboxylic amino acids are, however, often found elsewhere in the c-subunits, usually at either side of the transmembrane region.…”

Section: Resultsmentioning

confidence: 99%

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On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

Leone

Pogoryelov

Meier

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Numerous membrane transporters and enzymes couple their mechanisms to the permeation of Na + or H + , thereby harnessing the energy stored in the form of transmembrane electrochemical potential gradients to sustain their activities. The molecular and environmental factors that control and modulate the ion specificity of most of these systems are, however, poorly understood. Here, we use isothermal titration calorimetry to determine the Na + /H + selectivity of the ion-driven membrane rotor of an F-type ATP synthase. Consistent with earlier theoretical predictions, we find that this rotor is significantly H + selective, although not sufficiently to be functionally coupled to H + , owing to the large excess of Na + in physiological settings. The functional Na + specificity of this ATP synthase thus results from two opposing factors, namely its inherent chemical selectivity and the relative availability of the coupling ion. Further theoretical studies of this membrane rotor, and of two others with a much stronger and a slightly weaker H + selectivity, indicate that, although the inherent selectivity of their ion-binding sites is largely set by the balance of polar and hydrophobic groups flanking a conserved carboxylic side chain, subtle variations in their structure and conformational dynamics, for a similar chemical makeup, can also have a significant contribution. We propose that the principle of ion selectivity outlined here may provide a rationale for the differentiation of Na + -and H + -coupled systems in other families of membrane transporters and enzymes. molecular motor | ion-coupled transport | binding thermodynamics | energy transduction | membrane bioenergetics G radients in the electrochemical potential of H + or Na + across biological membranes sustain a wide range of essential cellular process. The resulting proton or sodium motive forces (pmf, smf) are the predominant energy source for secondary-active membrane transporters, which mediate the uptake of many substances required by the cell (1-3), and also enable pathogenic bacteria to protect themselves from human-made antibiotics and other toxic compounds (4-6). Downhill membrane permeation of Na + and H + across the membrane also powers the ATP synthase (7), which produces most of cellular ATP, and energizes the rotation of bacterial flagella (8). Thus, the importance of this mode of energy transduction in cells cannot be overstated. Nevertheless, little is known about the factors that control and modulate the specificity for Na + or H + in most of these processes.It seems clear, although, that there is no correlation between function type and ion specificity; that is, the same process in different species can be coupled to either Na + or H + (2, 5-7, 9-11). Organism-specific environmental factors, such as temperature or pH, also do not provide a consistent rationale; for example, ATP synthases from thermoalkaliphilic bacteria use a H + gradient despite the scarcity of H + and the potentially greater degree of H + leakage across the membrane at hig...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

Leone

Pogoryelov

Meier

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Previous modeling studies of the F-ATPase have addressed structural and mechanistic questions about the rotary mechanism (e.g., refs. [11][12][13][14][15][16][17][18][19][20], but not the metaissue of the mechanism itself compared with alternatives.…”

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confidence: 99%

Biophysical comparison of ATP synthesis mechanisms shows a kinetic advantage for the rotary process

Anandakrishnan

Zhang

Donovan-Maiye

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The ATP synthase (F-ATPase) is a highly complex rotary machine that synthesizes ATP, powered by a proton electrochemical gradient. Why did evolution select such an elaborate mechanism over arguably simpler alternating-access processes that can be reversed to perform ATP synthesis? We studied a systematic enumeration of alternative mechanisms, using numerical and theoretical means. When the alternative models are optimized subject to fundamental thermodynamic constraints, they fail to match the kinetic ability of the rotary mechanism over a wide range of conditions, particularly under low-energy conditions. We used a physically interpretable, closed-form solution for the steady-state rate for an arbitrary chemical cycle, which clarifies kinetic effects of complex free-energy landscapes. Our analysis also yields insights into the debated "kinetic equivalence" of ATP synthesis driven by transmembrane pH and potential difference. Overall, our study suggests that the complexity of the F-ATPase may have resulted from positive selection for its kinetic advantage.ATP synthase | kinetic mechanism | free-energy landscape | nonequilibrium steady state | evolution T he F-ATPase performs molecular-level free-energy (FE) transduction to phosphorylate ADP and yield ATP, the primary energy carrier that drives a vast range of cellular processes. It spurred Mitchell's chemiosmotic hypothesis (1) and Boyer's now-validated proposal for the binding-change mechanism (2) in which a proton electrochemical gradient is transduced to rotationbased mechanical energy and then back to chemical FE as ADP is phosphorylated. The F-ATPase's two-domain uniaxial rotary structure is conserved across all three domains of life (3-6), and details of its function have been the subject of a multitude of studies (e.g., refs. 7-30).Here, we address a relatively narrow question with potentially significant evolutionary implications: Why is ATP synthesized by a rotary mechanism instead of a potentially much simpler alternating-access mechanism (Fig. 1)? Using thermodynamic and kinetic constraints, we address the question of whether evolution tends to arrive at optimal molecular processes (31), building on established concepts of optimality-derived evolutionary convergence (32, 33). Presumably, performance advantages in the central task of ATP synthesis would be under significant evolutionary pressure. Previous modeling studies of the F-ATPase have addressed structural and mechanistic questions about the rotary mechanism (e.g., refs. 11-20), but not the metaissue of the mechanism itself compared with alternatives.To assess whether the rotary mechanism possesses any intrinsic performance advantage, we constructed a series of kinetic models abstracted from known mechanisms (Fig. 1). Beyond the rotarybased model, we considered a series of alternating-access analogs (Fig. 2), building on the demonstrated capacity for ATP-hydrolyzing transporters to be driven in reverse to synthesize ATP (34, 35). The discrete-state models do not include structural details, bu...

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“…Net ATP formation is achieved via electron transport phosphorylation involving the membrane-associated complexes RnfA to -G and F 1 F o ATP synthase. The exergonic reduction of NAD ϩ with reduced ferredoxin catalyzed by the RnfA to -G complex is associated with the buildup of an electrochemical sodium ion potential (1 Na ϩ translocated per electron) (19), which in turn drives the phosphorylation of ADP via the F 1 F o ATP synthase complex (3.3 Na ϩ translocated per ATP) (20). The reduced ferredoxin is regenerated via reduction of ferredoxin and NAD ϩ with 2 H 2 catalyzed by the electron-bifurcating and NAD-dependent [FeFe]-hydrogenase HydABCD (21).…”

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Energy Conservation Associated with Ethanol Formation from H ₂ and CO ₂ in Clostridium autoethanogenum Involving Electron Bifurcation

et al. 2015

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Most acetogens can reduce CO 2 with H 2 to acetic acid via the Wood-Ljungdahl pathway, in which the ATP required for formate activation is regenerated in the acetate kinase reaction. However, a few acetogens, such as Clostridium autoethanogenum, Clostridium ljungdahlii, and Clostridium ragsdalei, also form large amounts of ethanol from CO 2 and H 2 . How these anaerobes with a growth pH optimum near 5 conserve energy has remained elusive. We investigated this question by determining the specific activities and cofactor specificities of all relevant oxidoreductases in cell extracts of H 2 /CO 2 -grown C. autoethanogenum. The activity studies were backed up by transcriptional and mutational analyses. Most notably, despite the presence of six hydrogenase systems of various types encoded in the genome, the cells appear to contain only one active hydrogenase. The active [FeFe]-hydrogenase is electron bifurcating, with ferredoxin and NADP as the two electron acceptors. Consistently, most of the other active oxidoreductases rely on either reduced ferredoxin and/or NADPH as the electron donor. An exception is ethanol dehydrogenase, which was found to be NAD specific. Methylenetetrahydrofolate reductase activity could only be demonstrated with artificial electron donors. Key to the understanding of this energy metabolism is the presence of membrane-associated reduced ferredoxin:NAD ؉ oxidoreductase (Rnf), of electron-bifurcating and ferredoxin-dependent transhydrogenase (Nfn), and of acetaldehyde:ferredoxin oxidoreductase, which is present with very high specific activities in H 2 /CO 2 -grown cells. Based on these findings and on thermodynamic considerations, we propose metabolic schemes that allow, depending on the H 2 partial pressure, the chemiosmotic synthesis of 0.14 to 1.5 mol ATP per mol ethanol synthesized from CO 2 and H 2 . IMPORTANCEEthanol formation from syngas (H 2 , CO, and CO 2 ) and from H 2 and CO 2 that is catalyzed by bacteria is presently a much-discussed process for sustainable production of biofuels. Although the process is already in use, its biochemistry is only incompletely understood. The most pertinent question is how the bacteria conserve energy for growth during ethanol formation from H 2 and CO 2 , considering that acetyl coenzyme A (acetyl-CoA), is an intermediate. Can reduction of the activated acetic acid to ethanol with H 2 be coupled with the phosphorylation of ADP? Evidence is presented that this is indeed possible, via both substrate-level phosphorylation and electron transport phosphorylation. In the case of substrate-level phosphorylation, acetyl-CoA reduction to ethanol proceeds via free acetic acid involving acetaldehyde:ferredoxin oxidoreductase (carboxylate reductase).

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High-resolution structure and mechanism of an F/V-hybrid rotor ring in a Na+-coupled ATP synthase

Cited by 71 publications

References 66 publications

On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

Biophysical comparison of ATP synthesis mechanisms shows a kinetic advantage for the rotary process

Energy Conservation Associated with Ethanol Formation from H ₂ and CO ₂ in Clostridium autoethanogenum Involving Electron Bifurcation

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High-resolution structure and mechanism of an F/V-hybrid rotor ring in a Na+-coupled ATP synthase

Cited by 71 publications

References 66 publications

On the principle of ion selectivity in Na + /H + -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

On the principle of ion selectivity in Na + /H + -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

Biophysical comparison of ATP synthesis mechanisms shows a kinetic advantage for the rotary process

Energy Conservation Associated with Ethanol Formation from H 2 and CO 2 in Clostridium autoethanogenum Involving Electron Bifurcation

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On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

On the principle of ion selectivity in Na ⁺ /H ⁺ -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor

Energy Conservation Associated with Ethanol Formation from H ₂ and CO ₂ in Clostridium autoethanogenum Involving Electron Bifurcation