ATP-synthase

Electron micrographs of mitochondria show globular structures protruding from the matrix side of the inner mitochondrial membrane. These globular units can be detached from the membranes by relatively mild treatment such as low ionic strength or urea. The solubilized particles, known as the F[ component of ATP synthase, can hydrolyze ATP, but cannot synthesize it, and hence has been termed an ATPase. 

ATP synthase activity can be restored by adding back the F] complex to the depleted membranes. The F[ complexes bind to membrane channels known as the F complex, which are also composed of multiple subunits. The polypeptides of the F0 component are very hydrophobic and form a proton transport channel through the membrane, which links the proton gradient to ATP synthesis. This channel appears to be lined with hydrophilic residues such as seryl, threonyl and carboxyl groups. The stalk that connects the F, to the F complex comprises one copy each of the polypeptide known as the oligomycin-sensitivity-conferring protein (OSCP) and another protein known as F6. 

The major subunit of the FH complex is a small (Mr = 5,400) polypeptide referred to as proteolipid because of the high proportion of phospholipid bound to it. This proteolipid, now sequenced from a number of sources, forms a hairpin loop that traverses the inner mitochondrial membrane and has unique hydrophobic amino acid sequences flanking a central, short, highly charged segment that interacts with the OSCP or with F, component 5. 

The compound dicyclohexylcarbodiimide (DCCD) inhibits proton translocation through F0 by reacting with the carboxyl group of a single glutamate residue in the channel-forming loop of the proteolipid. 

Oligomycin is an antibiotic that inhibits respiration in intact mitochondria. Respiration is not inhibited in uncoupled mitochondria, i.e., those mitochondria in which 02 consumption occurs but in which no ATP is synthesized. Thus, oligomycin does not block respiratory carriers, in contrast to inhibitors such as rotenone and cyanide. Instead, oligomycin blocks proton translocation through the F component to the F( component, through a specific interaction with a subunit of the membrane-associated F . The subscript o in the term F was originally used to indicate the oligomycin-sensitive complex. 

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Figure C3.2.17. Diagram of a liposome-based artificial photosynthetic membrane showing the photocycle that pumps protons into the interior of the liposome and the CFqF j-ATP synthase enzyme. From [55],...

Steinberg-Yfrach G, Rigaud G-L, Durantini E N, Moore A L, Gust D and Moore T A 1998 Light driven production of ATP catalysed by F0F1-ATP synthase in an artificial photosynthetic membrane Nature 392 479-82... 

Complex IV Cytochrome c Oxidase The Thermodynamic View of Chemiosmotic Coupling ATP Synthase... 

Inhibitors of Oxidative Phosphorylatioi Unconplers Disrupt die Coupling of Electron Transport and ATP Synthase ATP Exits die Mitochondria via an ATP-ADP Transloca.se... 

FIGURE 21.23 Electron micrograph of sub-mitochondrial particles showing the 8.5-nm projections or particles on the inner membrane, eventnally shown to be Fj-ATP synthase. (Parsons, D. E, 1963. Science 140 985)... 

The mitochondrial complex that carries out ATP synthesis is called ATP synthase or sometimes FjFo-ATPase (for the reverse reaction it catalyzes). ATP synthase was observed in early electron micrographs of submitochondrial particles (prepared by sonication of inner membrane preparations) as round, 8.5-nm-diameter projections or particles on the inner membrane (Figure 21.23). In micrographs of native mitochondria, the projections appear on the matrixfacing surface of the inner membrane. Mild agitation removes the particles from isolated membrane preparations, and the isolated spherical particles catalyze ATP hydrolysis, the reverse reaction of the ATP synthase. Stripped of these particles, the membranes can still carry out electron transfer but cannot synthesize ATP. In one of the first reconstitution experiments with membrane proteins, Efraim Racker showed that adding the particles back to stripped membranes restored electron transfer-dependent ATP synthesis. 

ATP synthase actually consists of two principal complexes. The spheres observed in electron micrographs make up the Fj unit, which catalyzes ATP synthesis. These Fj spheres are attached to an integral membrane protein aggregate called the Fq unit. Fj consists of five polypeptide chains named a, j3, y, 8, and e, with a subunit stoichiometry ajjSaySe (Table 21.3). Fq consists of three hydrophobic subunits denoted by a, b, and c, with an apparent stoichiometry of ajbgCg.ig- Fq forms the transmembrane pore or channel through which protons move to drive ATP synthesis. The a, j3, y, 8, and e subunits of Fj contain 510, 482, 272, 146, and 50 amino acids, respectively, with a total molecular mass... 

Escherichia coli FiFg ATP Synthase Subunit Organization ... 

FIGURE 21.24 Molecular graphic images (a) side view and (b) top view of the Fj-ATP synthase showing the individnal component peptides. The 7-snbnnit is the pink strnctnre visible in the center of view (b). 

FIGURE 21.25 A model of the Fj and Fg components of the ATP synthase, a rotating molecnlar motor. The a, b, a, /3, and 8 snbnnits constitute the stator of the motor, and the c, y, and e subunits form the rotor. Flow of protons through the structure turns the rotor and drives the cycle of conformational changes in a and fi that synthesize ATP. 

FIGURE 21.28 The reconstituted vesicles containing ATP synthase and bacteriorhodopsin used by Stoeckenius and Racker to confirm the Mitchell chemiosmotic hypothesis. 

Uncouplers Disrupt the Coupling of Electron Transport and ATP Synthase... 

FIGURE 21.31 Structures of several uiicouplers, molecules that dissipate the proton gradient across the inner mitochondrial membrane and thereby destroy the tight coupling between electron transport and the ATP synthase reaction. 

Assume that the free energy change, AG, associated with the movement of one mole of protons from the outside to the inside of a bacterial cell is —23 kJ/mol and 3 must cross the bacterial plasma membrane per ATP formed by the bacterial FjEo-ATP synthase. ATP synthesis thus takes place by the coupled process ... 

FIGURE 22.17 The R. viridis reaction center is coupled to the cytochrome h/Cl complex through the quinone pool (Q). Quinone molecules are photore-duced at the reaction center Qb site (2 e [2 hv] per Q reduced) and then diffuse to the cytochrome h/ci complex, where they are reoxidized. Note that e flow from cytochrome h/ci back to the reaction center occurs via the periplasmic protein cytochrome co- Note also that 3 to 4 are translocated into the periplasmic space for each Q molecule oxidized at cytochrome h/ci. The resultant proton-motive force drives ATP synthesis by the bacterial FiFo ATP synthase. (Adapted from Deisenhofer, and Michel, H., 1989. The photosynthetic reaction center from the purple bac-terinm Rhod.opseud.omoaas viridis. Science 245 1463.)... 

CFiCFo ATP Synthase Is the Chloroplast Equivalent of the Mitochondrial FiFq ATP Synthase... 

FIGURE 22.21 The mechanism of photophosphorylation. Photosynthetic electron transport establishes a proton gradient that is tapped by the CFiCFo ATP synthase to drive ATP synthesis. Critical to this mechanism is the fact that the membrane-bound components of light-induced electron transport and ATP synthesis are asymmetrical with respect to the thylakoid membrane so that vectorial discharge and uptake of ensue, generating the proton-motive force. 

If noncyclic photosynthetic electron transport leads to the translocation of 3 H /e and cyclic photosynthetic electron transport leads to the translocation of 2 H /A, what is the relative photosynthetic efficiency of ATP synthesis (expressed as the number of photons absorbed per ATP synthesized) for noncyclic versus cyclic photophosphorylation (Assume that the CFiCEq ATP synthase yields 1 ATP/3 H. )... 

Berry, S., and Rnmberg, B., 1996. H" /ATP coupling ratio at the unmodulated CFiCFq-ATP synthase determined by proton flux measurements. 

Complex V (ATP Synthase, Mitochondrial Proton-Translocating ATPase)... 

With this model, the energy-requiring step is not the formation of ATP but the conformational change that allows release of tightly bound ATP. The role of the a-subunits may be to maintain the functional conformation of the P-subunits. Another subtmit is sometimes associated with F this may regulate ATP synthase... 

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