Chemiosmotic hypothesis Mitchell

Mitchell s chemiosmotic hypothesis. The ratio of protons transported per pair of electrons passed through the chain—the so-called HV2 e ratio—has been an object of great interest for many years. Nevertheless, the ratio has remained extremely difficult to determine. The consensus estimate for the electron transport pathway from succinate to Og is 6 H /2 e. The ratio for Complex I by itself remains uncertain, but recent best estimates place it as high as 4 H /2 e. On the basis of this value, the stoichiometry of transport for the pathway from NADH to O2 is 10 H /2 e. Although this is the value assumed in Figure 21.21, it is important to realize that this represents a consensus drawn from many experiments. 

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... 

In 1961, Peter Mitchell proposed a novel coupling mechanism involving a proton gradient across the inner mitochondrial membrane. In Mitchell s chemiosmotic hypothesis, protons are driven across the membrane from the matrix to the intermembrane... 

When Mitchell first described his chemiosmotic hypothesis in 1961, little evidence existed to support it, and it was met with considerable skepticism by the scientific community. Eventually, however, considerable evidence accumulated to support this model. It is now clear that the electron transport chain generates a proton gradient, and careful measurements have shown that ATP is synthesized when a pH gradient is applied to mitochondria that cannot carry out electron transport. Even more relevant is a simple but crucial experiment reported in 1974 by Efraim Racker and Walther Stoeckenius, which provided specific confirmation of the Mitchell hypothesis. In this experiment, the bovine mitochondrial ATP synthasereconstituted in simple lipid vesicles with bac-teriorhodopsin, a light-driven proton pump from Halobaeterium halobium. As shown in Eigure 21.28, upon illumination, bacteriorhodopsin pumped protons... 

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

Mitchell s Nobel lecture, outlining the evolution of the chemiosmotic hypothesis. 

The chemiosmotic hypothesis (also known as the Mitchell hypothe sis) explains how the free energy generated by the transport of elec trons by the electron transport chain is used to produce ATP from ADP + Pj. 

The chemiosmotic hypothesis had the great virtue of predicting the following consequences which could be tested (1) electron-transport driven proton pumps with defined stoichiometries and (2) a separate ATP synthase, which could be driven by a pH gradient or membrane potential. Mitchell s hypothesis was initially greeted with skepticism but it encouraged many people, including Mitchell and his associate Jennifer Moyle, to test these predictions, which were soon found to be correct.178... 

Mitchell postulated the chemiosmotic hypothesis for the mechanism of oxidative phosphorylation. 

Since biochemists clearly understood that H+ ion was involved in oxidative phosphorylation, the alternative ATP formation concept occurred as a counter to chemical conjugation. This concept was called the chemiosmotic hypothesis of the oxidative phosphorylation mechanism. This hypothesis was developed by Mitchell, the famous English biochemist [20], who turned is attention to the blind sides of the chemical conjugation concept. 

Oxidative phosphorylation is the name given to the synthesis of ATP (phosphorylation) that occurs when NADH and FADH2 are oxidized (hence oxidative) by electron transport through the respiratory chain. Unlike substrate level phosphorylation (see Topics J3 and LI), it does not involve phosphorylated chemical intermediates. Rather, a very different mechanism was proposed by Peter Mitchell in 1961, the chemiosmotic hypothesis. This proposes that energy liberated by electron transport is used to create a proton gradient across the mitochondrial inner membrane and that it is this that is used to drive ATP synthesis. Thus the proton gradient couples electron transport and ATP synthesis, not a chemical intermediate. The evidence is overwhelming that this is indeed the way that oxidative phosphorylation works. The actual synthesis of ATP is carried out by an enzyme called ATP synthase located in the inner mitochondrial membrane (Fig. 3). 

In the previous chapter we indicated that the components involved with electron flow are situated in the lamellar membranes of chloroplasts such that they lead to a vectorial or unidirectional movement of electrons and protons (see Fig. 5-19). We now return to this theme and focus on the gradients in H+ (protons) thus created. In the light, the difference in the chemical potential of H+ from the inside to the outside of a thylakoid acts as the energy source to drive photophosphorylation. This was first clearly recognized in the 1960s by Peter Mitchell, who received the 1978 Nobel Prize in chemistry for his enunciation of what has become known as the chemiosmotic hypothesis for interpreting the relationship among electron flow, proton movements, and ATP formation. 

Figure 1. The major transmembrane photosynthetic reaction centers (RC) (top) and respiratory complexes (bottom) are composed of light (zigzag) activated chains (dark gray) of redox centers (open polygons) that create a transmembrane electric field and move protons (double arrows) to create a transmembrane proton gradient, fulfilling the requirements of Mitchell s chemiosmotic hypothesis. Diffusing substrates include ubiquinone (hexagon) and other sources of oxidants and reductants. PSI and PSII, photosystems I and II, respectively.

Two major ATP synthesizing reactions in living organisms are oxidative phosphorylation and photophosphorylation. Both reactions take place in H -ATPase (FqF,), which is driven by an electrochemical potential difference of protons across the biomembrane, as predicted by Mitchell [1]. In Racket s laboratory, ATPases related to oxidative phosphorylation were prepared, but their relationship to Mitchell s chemiosmotic hypothesis [1] was not described [2], Later, an insoluble ATPase (H -ATPase) was shown to translocate protons across the membrane when it was reconstituted into liposomes [3], H -ATPase was shown to be composed of a catalytic moiety called F, (coupling factor 1) [4], and a membrane moiety called Fq [5], which confers inhibitor sensitivity to F,. F was shown to be a proton channel, which translocates down an electrochemical potential gradient across the membrane when Fg is reconstituted into liposomes (Fig. 5.1) [6]. Thus, -ATPase was called FqFj or ATP synthetase. 

The answer is b. (Murray, pp 123-148. Scriver, pp 2367-2424. Sack, pp 159-115. Wilson, pp 287-3111) The chemiosmotic hypothesis of Mitchell describes the coupling of oxidative phosphorylation and electron transport. The movement of electrons along the electron transport chain allows protons to be pumped from the matrix of the mitochondria to the cytoplasmic side. The protons are pumped at three sites in the electron transport chain to produce a proton gradient. When protons flow back through proton channels of the asymmetrically oriented ATPase of the inner mitochondrial membrane, ATP is synthesized. 

A mechanism that incorporates these observations is the chemiosmotic hypothesis first suggested by Peter Mitchell in 1961. His ideas were greeted with much scepticism when they were first formulated but they have gradually come to be accepted by the majority of biochemists, so much so that Peter Mitchell has been awarded the 1978 Nobel Prize for Chemistry for this work and... 

His early observations provided some of the most important experimental evidence in support of Mitchell s chemiosmotic hypothesis. 

P. Mitchell and I. Moyle, Chemiosmotic hypothesis of oxidative phosphorylation, Nature, 213 (1967) 137-139. 

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