The Mechanism of ATP Synthesis

The x-ray crystal structure of the F ATPase has recently been determined at 2.8 A resolution. The three a and three /3 chains are arranged alternately in a circle around an a-helical coiled coil of the y subunit that protrudes into the stalk region. The Fj subunit is fundamentally asymmetrical because there is only a single y subunit to three a and three /3 subunits in addition, each of the three a chains and each of the three /3 chains adopts a slightly different conformation. One /3 subunit binds ATP, one binds ADP, while the third appears to have an empty nucleotide binding site. 

The concept of a high-energy phosphorylated intermediate at coupling sites was a feature of all earlier concepts of the mechanism of ADP phosphorylation. The possibility of such a precursor, engendered by the electrochemical gradient on Fi, has however been eliminated by studies involving the use of, sO-labeled components of the reaction  

Covalent bond formation during ATP synthesis is thus restricted to the one linking the /3 and y phosphates of ATP itself. 

The energy of the electrochemical A/rH+ appears not to be utilized directly in the making of this covalent linkage but is used, rather, in the binding of ADP and Pj to Fi and in the subsequent release of ATP. 

Models based on the above concept have been proposed, such as the one shown in Fig. 14-8. Evidence for this model is based on studies of the hydrolysis of ATP labeled in the y phosphate with 1kO. The label is lost from the released P, to water (H18OH) because of the rapid reversibility of reactions 2 and -2. Uncoupling agents do not influence this loss, indicating is not involved in reactions 2 or -2, nor in reactions -3 or -1. 

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The mechanism of ATP synthesis discussed here assumes that protons extruded during electron transport are in the bulk phase surrounding the inner mitochondrial membrane (intermembrane and extramitochondrial spaces). An alternative view is that there are local proton circuits within or close to the respiratory chain and complex V, and that these protons may not be in free equilibrium with the bulk phase (Williams, 1978), although this has not been supported experimentally (for references see Nicholls and Ferguson, 1992). The chemiosmotic mechanism is both elegant and simple and explains all the known facts about ATP synthesis and its dependence on the structural integrity of the mitochondria, although the details may appear complex. This mechanism will now be discussed in more detail. 

The individual steps of the multistep chemical reduction of COj with the aid of NADPHj require an energy supply. This supply is secured by participation of ATP molecules in these steps. The chloroplasts of plants contain few mitochondria. Hence, the ATP molecules are formed in plants not by oxidative phosphorylation of ADP but by a phosphorylation reaction coupled with the individual steps of the photosynthesis reaction, particularly with the steps in the transition from PSII to PSI. The mechanism of ATP synthesis evidently is similar to the electrochemical mechanism involved in their formation by oxidative phosphorylation owing to concentration gradients of the hydrogen ions between the two sides of internal chloroplast membranes, a certain membrane potential develops on account of which the ATP can be synthesized from ADP. Three molecules of ATP are involved in the reaction per molecule of COj. 

Although the mechanism of ATP synthesis is not fully understood, there is no doubt that the ATP is synthesized in the matrix of the mitochondrion. Mitochondrial ATP is then exported to the cytoplasm. A specific carrier (translocase Mr = 40,000) is involved in the simultaneous transport of... 

The mechanism of ATP synthesis coupled to electron transport in thylakoids is discussed in Chapter 7 of this volume, and the reader is referred there. Some general aspects of photophosphorylation will be dealt with here in relation to the structure of thylakoids, their supramolecular organization and the overall efficiency of the process. 

The reduction of methanol with H2 to methane represents the simplest form of methane fermentation, and turned out to be extremely useful for studying the mechanism of ATP synthesis in cell suspensions [41] (see below). Growth at the expense of CH3OH reduction with H2 has been demonstrated so far only with Methanosphaera stadtmanae [42] and... 

N Good (1960) Activation of the Hiii reaction by amines. Biochim Biophys Acta 40 502-517 RE McCarty (1980) Deiineation of the mechanism of ATP synthesis in chioropiasts Use of uncoupiers, energy transfer inhibitors, and modifiers of coupiing factor 1. Methods in Enzymology 69 (Photosynthesis and Nitrogen Fixation, Part C (A San Pietro, ed) pp. 719-728... 

The above described difference in restoration of coupled ATP-linked activities could be due, either to the very different isolation procedures of the native Rr/ as compared to the foreign Ec/ and CFifi or to their inherent different interactions with all other RrFoFi-subunits in the reconstituted complex. Close interactions between the Fi-p subunit and other F Fx-subunits are a basic parameter in all recent models for the mechanism of ATP synthesis by the H+-translocating FoFi-ATP synthase (7-9). 

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