ATP synthesis

The protein complex that carries out this reaction is called coupling factor or, more accurately, ATP synthase. ATP synthase is a complex of several proteins, shaped like a mushroom, with the cap on the stromal side of the thylakoid disc and the stalk going through the thylakoid 

This pathway is sometimes called the Calvin-Benson cycle, after the biochemists who elucidated it. The 5-carbon, doubly phosphorylated carbohydrate, ribulose bisphosphate is the acceptor for CO2 the enzyme is called ribulose-bisphosphate carboxylase/oxygenase (called Rubisco). 

The initial product of the reaction is unstable and quickly falls apart to yield two molecules of 3-phosphoglycerate. 

The rest of the Calvin cycle is involved in interconversion of carbohydrates to make glucose (or starch) and the regeneration of the ribulose-bisphosphate acceptor. The reactions are also found in the pathways for gluconeogenesis and the pentose phosphate shunt (see Volume 1, Chapters 10 and 12). The first step is the phosphorylation of 3-phosphoglycerate by the same reactions involved in gluconeogenesis. 

The glyceraldehyde-3-phosphate can be converted into the 6-carbon sugar phosphate called fructose-6-phosphate by the reactions of triose phosphate isomerase, aldolase, and fructose bisphosphase. 

In the respiratory chain (see p. 140), electrons are transferred from NADH or ubiquinol (QH2) to O2. The energy obtained in this process is used to establish a proton gradient across the inner mitochondrial membrane. ATP synthesis is ultimately coupled to the return of protons from the intermembrane space into the matrix. 

The illustration shows the important redox systems involved in mitochondrial electron transport and their approximate redox potentials. These potentials determine the path followed by the electrons, as the members of a redox series have to be arranged in order of increasing redox potential if transport is to occur spontaneously (see p. 32). 

In complex 1, the electrons are passed from NADH+H first to FMN (see p. 104) and then on to several iron-sulfur (Fe/S) clusters. These redox systems are only stable in the interior of proteins. Depending on the type, Fe/S clusters may contain two to six iron ions, which 

The ATP synthase (EC3.6.1.34, complex V) that transports H is a complex molecular machine. The enzyme consists of two parts—a proton channel (Fq, for oligomycin-sensitive ) that is integrated into the membrane and a catalytic unit (Fi) that protrudes into the matrix. The Fo part consists of 12 membrane-spanning c-peptides and one a-subunit. The head of the Fi part is composed of three a and three p subunits, between which there are three active centers. The stem between Fo and Fi consists of one y and one e subunit. Two more polypeptides, b and 8, form a kind of stator, fixing the a and p subunits relative to the Fo part. 

The catalytic cycle can be divided into three phases, through each of which the three active sites pass in sequence. First, ADP and Pj are bound (1), then the anhydride bond forms (2), and finally the product is released (3). Each time protons pass through the Fo channel protein into the matrix, all three active sites change from their current state to the next. It has been shown that the energy for proton transport is initially converted into a rotation of the y subunit, which in turn cyclically alters the conformation of the a and p subunits, which are stationary relative to the Fo part, and thereby drives ATP synthesis. 

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Those herbicides that block mitotic entry decrease or prevent the formation of mitotic figures in meristems. Amino acid, protein, RNA, DNA, and ATP synthesis and/or utilization can all attest cell growth (163,166). Although not registered as herbicides, cycloheximide [66-81-9] inhibits mitotic entry by inhibiting protein synthesis (167) hydroxyurea/727-(97-/7 inhibits DNA synthesis (168) and actinomycin D [50-76-0] nh2oix.s RNA synthesis (167). 

Glycolysis and the citric acid cycle (to be discussed in Chapter 20) are coupled via phosphofructokinase, because citrate, an intermediate in the citric acid cycle, is an allosteric inhibitor of phosphofructokinase. When the citric acid cycle reaches saturation, glycolysis (which feeds the citric acid cycle under aerobic conditions) slows down. The citric acid cycle directs electrons into the electron transport chain (for the purpose of ATP synthesis in oxidative phosphorylation) and also provides precursor molecules for biosynthetic pathways. Inhibition of glycolysis by citrate ensures that glucose will not be committed to these activities if the citric acid cycle is already saturated. 

Wall Piece IV (1985), a kinetic sculpture by George Rhoads. This complex meehanieal art form can be viewed as a metaphor for the molecular apparatus underlying electron transport and ATP synthesis by oxidative phosphorylation. (1985 ty George Rhoaeh)... 

Whereas ATP made in glycolysis and the TCA cycle is the result of substrate-level phosphorylation, NADH-dependent ATP synthesis is the result of oxidative phosphorylation. Electrons stored in the form of the reduced coenzymes, NADH or [FADHa], are passed through an elaborate and highly orga-... 

This is a crucial point because (as we will see) proton transport is coupled with ATP synthesis. Oxidation of one FADHg in the electron transport chain results in synthesis of approximately two molecules of ATP, compared with the approximately three ATPs produced by the oxidation of one NADH. Other enzymes can also supply electrons to UQ, including mitochondrial 5w-glyc-erophosphate dehydrogenase, an inner membrane-bound shuttle enzyme, and the fatty acyl-CoA dehydrogenases, three soluble matrix enzymes involved in fatty acid oxidation (Figure 21.7 also see Chapter 24). The path of electrons from succinate to UQ is shown in Figure 21.8. 

Peter Mitchell s chemiosmotic hypothesis revolutionized our thinking about the energy coupling that drives ATP synthesis by means of an electrochemical gradient. How much energy is stored in this electrochemical gradient For the transmembrane flow of protons across the inner membrane (from inside [matrix] to outside), we could write... 

Engelhardt s experiments in 1930 led to the notion that ATP is synthesized as the result of electron transport, and, by 1940, Severo Ochoa had carried out a measurement of the P/O ratio, the number of molecules of ATP generated per atom of oxygen consumed in the electron transport chain. Because two electrons are transferred down the chain per oxygen atom reduced, the P/O ratio also reflects the ratio of ATPs synthesized per pair of electrons consumed. After many tedious and careful measurements, scientists decided that the P/O ratio was 3 for NADH oxidation and 2 for succinate (that is, [FADHg]) oxidation. Electron flow and ATP synthesis are very tightly coupled in the sense that, in normal mitochondria, neither occurs without the other. 

Many models were proposed to account for the coupling of electron transport and ATP synthesis. A persuasive model, advanced by E. C. Slater in 1953, proposed that energy derived from electron transport was stored in a high-energy intermediate (symbolized as X P). This chemical species—in essence an activated form of phosphate—functioned according to certain relations according to Equations (21.22)-(21.25) (see below) to drive ATP synthesis. 

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... 

How might such cycling occur Important clues have emerged from several experiments that show that the y subunit rotates with respect to the af3 complex. How such rotation might be linked to transmembrane proton flow and ATP synthesis is shown in Figure 21.25. In this model, the c subunits of Fq... 

What is the cost of ATP-ADP exchange relative to the energy cost of ATP synthesis itselD We already noted that moving 1 ATP out and 1 ADP in is the equivalent of one proton moving from the cytosol to the matrix. Synthesis of an... 

Under these conditions, what is the maximum ratio of [ATP]/[ADP] attainable by oxidative phosphorylation when [PJ = 1 mM (Assume AG° for ATP synthesis = +30.5 kj/mol.)... 

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 ... 

What molecular architecture couples the absorption of light energy to rapid electron-transfer events, in turn coupling these e transfers to proton translocations so that ATP synthesis is possible Part of the answer to this question lies in the membrane-associated nature of the photosystems. Membrane proteins have been difficult to study due to their insolubility in the usual aqueous solvents employed in protein biochemistry. A major breakthrough occurred in 1984 when Johann Deisenhofer, Hartmut Michel, and Robert Huber reported the first X-ray crystallographic analysis of a membrane protein. To the great benefit of photosynthesis research, this protein was the reaction center from the photosynthetic purple bacterium Rhodopseudomonas viridis. This research earned these three scientists the 1984 Nobel Prize in chemistry. 

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.)... 

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