ATP-synthase mitochondriale : de la structure de l'enzyme à l'organelle. – F1Fo

The F1Fo ATP-synthase is a key enzyme since it synthesises ATP, the fuel molecule of any living organism. The ATP-synthase is located in the inner membrane of mitochondria, in the thylakoid membrane of chloroplasts, or in the plasma membrane of bacteria. The ATP-synthase converts the electrochemical potential energy of the proton gradient that is generated across the membranes by respiration or photoreaction to the chemical energy of ATP synthesis, through cooperative conformational changes. The mitochondrial ATP-synthase is more complex than the other ATP-synthases. In yeast, this large enzyme of 600 kDa comprises 17 distinct subunits, whereas only 8 and 9 subunits compose ATPsynthases from bacteria and chloroplasts, respectively. Its membranous part (Fo) conducts protons across the inner mitochondrial membrane and its hydrophilic protruding portion (F1) catalyses ATP synthesis at three catalytic sites. The functioning of this enzyme as a rotary engine is now clearly demonstrated. Protons translocation across the Fo drives the rotation of a ring of subunits 9, which in turn induces the rotation of a central stalk (subunits ³¿µ) inside a barrel formed by three ±² pairs. This rotation generates conformational changes of ² subunits and enables catalytic sites to alternatively modulate their affinities for substrates and products. To prevent the ( ±²)3 head from rotating with the central stalk and subunit 9-ring, an elongated part (called peripheral or second stalk) composed of subunits OSCP, 4, d, h, 8, f and i, is proposed to play the role of a stator by connecting and anchoring the catalytic subunits to the membrane. This region extends from the top of F1, lies along the external surface of the ( ±²)3 domain, and then reaches the membrane part to interact with subunit 6. Although many crystallographers have worked since the seventies to get structural data on the F1Fo ATP-synthase, only a partial model of this enzyme has been obtained by 3D crystallography in Walker's laboratory. This model contains the rotor (central stalk + subunit 9-ring) and the catalytic subunits ( ±²)3. Peripheral stalk subunits and subunit 6 were lost upon crystallization. An overall envelope of the enzyme has been obtained at a 32 Å resolution by cryo-electron microscopy 3D reconstruction on isolated particles. However this envelope did not provide any information on subunit 6 or on the organization of the second stalk components. Thus, subunit 6 (which together with subunit 9-ring, constitutes the proton channel) and the peripheral stalk are the less defined regions of the enzyme, and their structural properties in the complex have not yet been exactly determined. Subunit 6 is of very particular interest since a few mutations of the mitochondrial gene encoding this subunit have been found in patients with severe mitochondrial diseases. Recently, the second stalk has been shown to be involved in ATP-synthase oligomerisation. Indeed the mitochondrial ATP-synthase was found in dimeric and oligomeric forms upon extraction with mild detergent and analysis using native PAGE. Three subunits (e, g and k) have been found to be specifically associated with dimeric species. In yeast, disruptions of subunit g or e do not dramatically affect oxidative phosphorylations, but cells display abnormal mitochondria with large onion-like structures instead of classical inner membrane invaginations called mitochondrial cristae. In strains lacking subunit e or g, dimeric forms are less stable but still exist in the inner mitochondrial membrane. Cross-linking experiments have shown that subunits of the second stalk (subunits i, 4, and h) likely mediate the interface involved in dimer formation and that subunit e and g stabilise those dimers and create another interface to generate ATP-synthase tetramers. Although mitochondrial cristae were first observed in the sixties by electron microscopy, their genesis and their role have not extensively been investigated. As mentioned