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The structure of the membrane extrinsic region of bovine ATP synthase.

ABSTRACT: The structure of the complex between bovine mitochondrial F(1)-ATPase and a stator subcomplex has been determined at a resolution of 3.2 A. The resolved region of the stator contains residues 122-207 of subunit b; residues 5-25 and 35-57 of F(6); 3 segments of subunit d from residues 30-40, 65-74, and 85-91; and residues 1-146 and 169-189 of the oligomycin sensitivity conferral protein (OSCP). The stator subcomplex represents its membrane distal part, and its structure has been augmented with an earlier structure of a subcomplex containing residues 79-183, 3-123, and 5-70 of subunits b, d, and F(6), respectively, which extends to the surface of the inner membrane of the mitochondrion. The N-terminal domain of the OSCP links the stator with F(1)-ATPase via alpha-helical interactions with the N-terminal region of subunit alpha(E). Its C-terminal domain makes extensive helix-helix interactions with the C-terminal alpha-helix of subunit b from residues 190-207. Subunit b extends as a continuous 160-A long alpha-helix from residue 188 back to residue 79 near to the surface of the inner mitochondrial membrane. This helix appears to be stiffened by other alpha-helices in subunits d and F(6), but the structure can bend inward toward the F(1) domain around residue 146 of subunit b. The linker region between the 2 domains of the OSCP also appears to be flexible, enabling the stator to adjust its shape as it passes over the changing profile of the F(1) domain during a catalytic cycle. The structure of the membrane extrinsic part of bovine ATP synthase is now complete.

SUBMITTER: Rees DM

PROVIDER: S-EPMC2789756 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Two subunits within the transmembrane domain of the ATP synthase-the c-ring and subunit a-energize the production of 90% of cellular ATP by transducing an electrochemical gradient of H+ or Na+ into rotational motion. The nature of this turbine-like energy conversion mechanism has been elusive for decades, owing to the lack of definitive structural information on subunit a or its c-ring interface. In a recent breakthrough, several structures of this complex were resolved by cryo-electron microscopy (cryo-EM), but the modest resolution of the data has led to divergent interpretations. Moreover, the unexpected architecture of the complex has cast doubts on a wealth of earlier biochemical analyses conducted to probe this structure. Here, we use quantitative molecular-modeling methods to derive a structure of the a-c complex that is not only objectively consistent with the cryo-EM data, but also with correlated mutation analyses of both subunits and with prior cross-linking and cysteine accessibility measurements. This systematic, integrative approach reveals unambiguously the topology of subunit a and its relationship with the c-ring. Mapping of known Cd2+ block sites and conserved protonatable residues onto the structure delineates two noncontiguous pathways across the complex, connecting two adjacent proton-binding sites in the c-ring to the space on either side of the membrane. The location of these binding sites and of a strictly conserved arginine on subunit a, which serves to prevent protons from hopping between them, explains the directionality of the rotary mechanism and its strict coupling to the proton-motive force. Additionally, mapping of mutations conferring resistance to oligomycin unexpectedly reveals that this prototypical inhibitor may bind to two distinct sites at the a-c interface, explaining its ability to block the mechanism of the enzyme irrespective of the direction of rotation of the c-ring. In summary, this study is a stepping stone toward establishing the mechanism of the ATP synthase at the atomic level.

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The structure of the membrane extrinsic region of bovine ATP synthase.

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