A New Type of Proton Coordination in an F1Fo-ATP Synthase Rotor Ring (original) (raw)
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F1000 - Post-publication peer review of the biomedical literature, 2010
We solved the crystal structure of a novel type of c-ring isolated from Bacillus pseudofirmus OF4 at 2.5 Å , revealing a cylinder with a tridecameric stoichiometry, a central pore, and an overall shape that is distinct from those reported thus far. Within the groove of two neighboring c-subunits, the conserved glutamate of the outer helix shares the proton with a bound water molecule which itself is coordinated by three other amino acids of outer helices. Although none of the inner helices contributes to ion binding and the glutamate has no other hydrogen bonding partner than the water oxygen, the site remains in a stable, ion-locked conformation that represents the functional state present at the c-ring/membrane interface during rotation. This structure reveals a new, third type of ion coordination in ATP synthases. It appears in the ion binding site of an alkaliphile in which it represents a finely tuned adaptation of the proton affinity during the reaction cycle.
High-resolution structure of the rotor ring of a proton-dependent ATP synthase
Nature Structural & Molecular Biology, 2009
The crystal structure of the c-ring from the proton-coupled F 1 F o ATP synthase from Spirulina platensis is shown at 2.1-Å resolution. The ring includes 15 membrane-embedded c subunits forming an hourglass-shaped assembly. The structure demonstrates that proton translocation across the membrane entails protonation of a conserved glutamate located near the membrane center in the c subunit outer helix. The proton is locked in this site by a precise hydrogen bond network reminiscent of that in Na + -dependent ATP synthases. However, the structure suggests that the different coordination chemistry of the bound proton and the smaller curvature of the outer helix drastically enhance the selectivity of the H + site against other cations, including H 3 O + . We propose a model for proton translocation whereby the c subunits remain in this proton-locked state when facing the membrane lipid. Proton exchange would occur in a more hydrophilic and electrostatically distinct environment upon contact with the a subunit interface.
Complete Ion-Coordination Structure in the Rotor Ring of Na+-Dependent F-ATP Synthases
Journal of Molecular Biology, 2009
The membrane-embedded rotors of Na + -dependent F-ATP synthases comprise 11 c-subunits that form a ring, with 11 Na + binding sites in between adjacent subunits. Following an updated crystallographic analysis of the c-ring from Ilyobacter tartaricus, we report the complete ioncoordination structure of the Na + sites. In addition to the four residues previously identified, there exists a fifth ligand, namely, a buried structural water molecule. This water is itself coordinated by Thr67, which, sequence analysis reveals, is the only residue involved in binding that distinguishes Na + synthases from H + -ATP synthases known to date. Molecular dynamics simulations and free-energy calculations of the c-ring in a lipid membrane lend clear support to the notion that this fifth ligand is a water molecule, and illustrate its influence on the selectivity of the binding sites. Given the evolutionary ascendancy of sodium over proton bioenergetics, this structure uncovers an ancient strategy for selective ion coupling in ATP synthases.
On the Structure of the Proton-Binding Site in the Fo Rotor of Chloroplast ATP Synthases
Journal of Molecular Biology, 2010
The recently reported crystal structures of the membrane-embedded proton-dependent c-ring rotors of a cyanobacterial F 1 F o ATP synthase and a chloroplast F 1 F o ATP synthase have provided new insights into the mechanism of this essential enzyme. While the overall features of these crings are similar, a discrepancy in the structure and hydrogen-bonding interaction network of the H + sites suggests two distinct binding modes, potentially reflecting a mechanistic differentiation. Importantly, the conformation of the key glutamate side chain to which the proton binds is also altered. To investigate the nature of these differences, we use molecular dynamics simulations of both c-rings embedded in a phospholipid membrane. We observe that the structure of the c 15 ring from Spirulina platensis is unequivocally stable within the simulation time. By contrast, the proposed structure of the H + site in the chloroplast c 14 ring changes rapidly and consistently into that reported for the c 15 ring, indicating that the latter represents a common binding mode. To assess this hypothesis, we have remodeled the c 14 ring by molecular replacement using the published structure factors. The resulting structure provides clear evidence in support of a common binding site conformation and is also considerably improved statistically. These findings, taken together with a sequence analysis of csubunits in the ATP synthase family, indicate that the so-called proton-locked conformation observed in the c 15 ring may be a common characteristic not only of light-driven systems such as chloroplasts and cyanobacteria but also of a selection of other bacterial species.
Febs Letters, 2004
The conformation of the ATP synthase c-subunit and the pKa of its essential E54 residue were characterized in alkaliphilic Bacillus pseudofirmus OF4. The c-subunit folds as a helix–loop–helix, with inter-helical contacts demonstrated by paramagnetic relaxation effects. The E54 pKa of 7.7 is significantly higher than in non-alkaliphiles, which likely prevents proton loss from the c-rotor at high pH. The E54 pKa was unchanged in a mutant, cP51A, that has a severe ATP synthesis defect at high pH only. cP51 must have some structural role that accounts for the mutant defect, such as different subunit-subunit interactions at high pH.
Crystal structure of the ϵ subunit of the proton-translocating ATP synthase from Escherichia coli
Structure, 1997
Background: Proton-translocating ATP synthases convert the energy generated from photosynthesis or respiration into ATP. These enzymes, termed F 0 F 1-ATPases, are structurally highly conserved. In Escherichia coli, F 0 F 1-ATPase consists of a membrane portion, F 0 , made up of three different polypeptides (a, b and c) and an F 1 portion comprising five different polypeptides in the stoichiometry α 3 β 3 γδε. The minor subunits γ, δ and ε are required for the coupling of proton translocation with ATP synthesis; the ε subunit is in close contact with the α, β, γ and c subunits. The structure of the ε subunit provides clues to its essential role in this complex enzyme. Results: The structure of the E. coli F 0 F 1-ATPase ε subunit has been solved at 2.3 Å resolution by multiple isomorphous replacement. The structure, comprising residues 2-136 of the polypeptide chain and 14 water molecules, refined to an R value of 0.214 (R free = 0.288). The molecule has a novel fold with two domains. The N-terminal domain is a β sandwich with two fivestranded sheets. The C-terminal domain is formed from two α helices arranged in an antiparallel coiled-coil. A series of alanine residues from each helix form the central contacting residues in the helical domain and can be described as an 'alanine zipper'. There is an extensive hydrophobic contact region between the two domains providing a stable interface. The individual domains of the crystal structure closely resemble the structures determined in solution by NMR spectroscopy. Conclusions: Sequence alignments of a number of ε subunits from diverse sources suggest that the C-terminal domain, which is absent in some species, is not essential for function. In the crystal the N-terminal domains of two ε subunits make a close hydrophobic interaction across a crystallographic twofold axis. This region has previously been proposed as the contact surface between the ε and γ subunits in the complete F 1-ATPase complex. In the crystal structure, we observe what is apparently a stable interface between the two domains of the ε subunit, consistent with the fact that the crystal and solution structures are quite similar despite close crystal packing. This suggests that a gross conformational change in the ε subunit, to transmit the effect of proton translocation to the catalytic domain, is unlikely, but cannot be ruled out.
Molecular Architecture of the Undecameric Rotor of a Bacterial Na+-ATP Synthase
Journal of Molecular Biology, 2002
The sodium ion-translocating F 1 F 0 ATP synthase from the bacterium Ilyobacter tartaricus contains a remarkably stable rotor ring composed of 11 c subunits. The rotor ring was isolated, crystallised in two dimensions and analysed by electron cryo-microscopy. Here, we present an a-carbon model of the c-subunit ring. Each monomeric c subunit of 89 amino acid residues folds into a helical hairpin consisting of two membrane-spanning helices and a cytoplasmic loop. The 11 N-terminal helices are closely spaced within an inner ring surrounding a cavity of , 17 Å (1.7 nm). The tight helix packing leaves no space for side-chains and is accounted for by a highly conserved motif of four glycine residues in the inner, N-terminal helix. Each inner helix is connected by a clearly visible loop to an outer C-terminal helix. The outer helix has a kink near the position of the ion-binding site residue Glu65 in the centre of the membrane and another kink near the C terminus. Two helices from the outer ring and one from the inner ring form the ion-binding site in the middle of the membrane and a potential access channel from the binding site to the cytoplasmic surface. Three possible inter-subunit ion-bridges are likely to account for the remarkable temperature stability of I. tartaricus c-rings compared to those of other organisms.
Unusual features of the c-ring of F1FO ATP synthases
Scientific Reports, 2019
Membrane integral ATP synthases produce adenosine triphosphate, the universal “energy currency” of most organisms. However, important details of proton driven energy conversion are still unknown. We present the first high-resolution structure (2.3 Å) of the in meso crystallized c-ring of 14 subunits from spinach chloroplasts. The structure reveals molecular mechanisms of intersubunit contacts in the c14-ring, and it shows additional electron densities inside the c-ring which form circles parallel to the membrane plane. Similar densities were found in all known high-resolution structures of c-rings of F1FO ATP synthases from archaea and bacteria to eukaryotes. The densities might originate from isoprenoid quinones (such as coenzyme Q in mitochondria and plastoquinone in chloroplasts) that is consistent with differential UV-Vis spectroscopy of the c-ring samples, unusually large distance between polar/apolar interfaces inside the c-ring and universality among different species. Althou...
A New Type of Na+-Driven ATP Synthase Membrane Rotor with a Two-Carboxylate Ion-Coupling Motif
PLoS Biology, 2013
The anaerobic bacterium Fusobacterium nucleatum uses glutamate decarboxylation to generate a transmembrane gradient of Na + . Here, we demonstrate that this ion-motive force is directly coupled to ATP synthesis, via an F 1 F o -ATP synthase with a novel Na + recognition motif, shared by other human pathogens. Molecular modeling and free-energy simulations of the rotary element of the enzyme, the c-ring, indicate Na + specificity in physiological settings. Consistently, activity measurements showed Na + stimulation of the enzyme, either membrane-embedded or isolated, and ATP synthesis was sensitive to the Na + ionophore monensin. Furthermore, Na + has a protective effect against inhibitors targeting the ionbinding sites, both in the complete ATP synthase and the isolated c-ring. Definitive evidence of Na + coupling is provided by two identical crystal structures of the c 11 ring, solved by X-ray crystallography at 2.2 and 2.6 Å resolution, at pH 5.3 and 8.7, respectively. Na + ions occupy all binding sites, each coordinated by four amino acids and a water molecule. Intriguingly, two carboxylates instead of one mediate ion binding. Simulations and experiments demonstrate that this motif implies that a proton is concurrently bound to all sites, although Na + alone drives the rotary mechanism. The structure thus reveals a new mode of ion coupling in ATP synthases and provides a basis for drug-design efforts against this opportunistic pathogen.