1068 VOLUME 16 NUMBER 10 OCTOBER 2009 NATURE STRUCTURAL & MOLECULAR BIOLOGY ARTICLES ATP is the universal energy currency in living cells, which constantly powers many and diverse biochemical processes. Across all life forms, ATP is mainly produced by the F 1 F o ATP synthase. The cyto- plasmic F 1 catalytic domain (subunits α 3 β 3 γδε) converts ADP and P i to ATP. A central and a peripheral stalk formed by subunits γε and b 2 δ, respectively, links F 1 to the membrane-embedded F o domain, which includes a reversible molecular rotor known as the c-ring. The transmembrane proton- or sodium-motive force (pmf, smf) drives ions to pass through the F o complex during ATP synthesis, inducing the rotation of the c-ring. The central stalk enables the transmission of this mechanical energy from the F o to the F 1 domain, by rotating alongside the c-ring. The inherently asymmetric γ subunit causes sequential conformational changes within each of the three catalytic β subunits in the F 1 head. The three different conformations of the β subunit endow the binding sites with three different nucleotide affinities at any time point 1 . A complete revolution of the rotor thus results in the synthesis of three ATP molecules. The structure of the F 1 complex has been determined 2 and a wealth of biochemical and biophysical data, including video microscopy 3 , support the rotational ATP synthesis mechanism. The F o motor consists in its simplest form of the oligomeric assem- bly of c subunits that constitute the c-ring, and of two neighboring subunits, namely a and b 2 . Each of the c subunits can reversibly bind one H + (or Na + ). Together with the γε complex, the c-ring rotor of the enzyme spins against the stator subunits ab 2 α 3 β 3 δ (ref. 4). The number of c subunits per ring has been found to vary between 10 (ref. 5) and 15 (ref. 6). The crystal structure of the rotor ring from the Ilyobacter tartaricus Na + -dependent ATP synthase 7 revealed that its c-ring comprises 11 subunits. Between each pair of adjacent sub- units a Na + -binding site can be clearly identified, within a compact coordination shell involving four amino acids and a buried struc- tural water molecule 8 . It follows also from this observation that ion exchange during enzyme activity is likely to be accompanied by con- formational changes in the binding site, presumably in the context of the a subunit interface 7 . A positively charged arginine in the stator a subunit is believed to be crucial in this regard 9–11 . To date, only a handful of species, including I. tartaricus, are known to use Na + as a coupling ion for ATP synthesis 12–15 . The vast majority of all living organisms use the pmf rather than the smf for generating ATP. Yet our knowledge of the structure of complete proton-translocating c subunit rotors has been limited to either low-resolution X-ray data 5,16 or to models derived from solution NMR measurements of monomeric c subunits in a chloroform-methanol- water mixture 17 . Current models of H + translocation are based on the notion that the c-ring forms a complex with the a subunit that creates two access channels for protons from either side of the membrane 4 . High-resolution structures to support or refute this model would be of great interest. Therefore, we have determined the structure of the F o rotor ring from the proton-translocating F 1 F o ATP synthase of alkaliphilic cyanobacterium Spirulina platensis 18 strain C1. RESULTS The c 15 ring of S. platensis C1 The c-ring of S. platensis C1 is the largest known in the F-ATP syn- thase family, and so far the only one that has been visually confirmed to be a pentadecamer 6 . Bipyramidal crystals with a yellowish-golden color were obtained. Spectroscopic analysis of isolated single crystals revealed that they contained chlorophyll a (Chl) and carotenoids (CR, mainly β-CR) in substoichiometric amounts (0.17 ± 0.03 moles of Chl+CR per mole of c 15 ). It is likely that traces of these photosynthetic 1 Department of Structural Biology, 2 Theoretical Molecular Biophysics Group and 3 Cluster of Excellence Macromolecular Complexes, Max-Planck Institute of Biophysics, Frankfurt am Main, Germany. Correspondence should be addressed to T.M. (thomas.meier@biophys.mpg.de). Received 28 May; accepted 21 August; published online 27 September 2009; doi:10.1038/nsmb.1678 High-resolution structure of the rotor ring of a proton-dependent ATP synthase Denys Pogoryelov 1 , Özkan Yildiz 1 , José D Faraldo-Gómez 2,3 & Thomas Meier 1,3 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. © 2009 Nature America, Inc. All rights reserved.