Oxidative phosphorylation electrochemical proton gradient

Figure 12-8. Principles of the chemiosmotic theory of oxidative phosphorylation. The main proton circuit is created by the coupling of oxidation in the respiratory chain to proton translocation from the inside to the outside of the membrane, driven by the respiratory chain complexes I, III, and IV, each of which acts as a protonpump. Q, ubiquinone C, cytochrome c F Fq, protein subunits which utilize energy from the proton gradient to promote phosphorylation. Uncoupling agents such as dinitrophenol allow leakage of H" across the membrane, thus collapsing the electrochemical proton gradient. Oligomycin specifically blocks conduction of H" through Fq.

Phosphorylation of ADP to ATP by mitochondria is driven by an electrochemical proton gradient established across the inner mitochondrial membrane as a consequence of vectoral transport of protons from NADH and succinate during oxidation by the respiratory chain (see Chapter 17). Hence, lipophilic weak acids or bases (such as 2,4-dinitrophenol) that can shuttle protons across membranes will dissipate the proton gradient and uncouple oxidation from ADP phosphorylation. Intrami-tochondrial ADP can be rate-limiting as demonstrated by inhibition of the mitochondrial adenosine nucleotide carrier by atractyloside. Inhibition of ATP synthesis... 

The thylakoid membrane is asymmetrically organized, or sided, like the mitochondrial membrane. It also shares the property of being a barrier to the passive diffusion of H ions. Photosynthetic electron transport thus establishes an electrochemical gradient, or proton-motive force, across the thylakoid membrane with the interior, or lumen, side accumulating H ions relative to the stroma of the chloroplast. Like oxidative phosphorylation, the mechanism of photophosphorylation is chemiosmotic. 

FIGURE 11-40 Reversibility of F-type ATPases. An ATP-driven proton transporter also can catalyze ATP synthesis (red arrows) as protons flow down their electrochemical gradient. This is the central reaction in the processes of oxidative phosphorylation and photophosphorylation, both described in detail in Chapter 19. 

Oxidative phosphorylation is ATP synthesis linked to the oxidation of NADH and FADH2 by electron transport through the respiratory chain. This occurs via a mechanism originally proposed as the chemiosmotic hypothesis. Energy liberated by electron transport is used to pump H+ ions out of the mitochondrion to create an electrochemical proton (H+) gradient. The protons flow back into the mitochondrion through the ATP synthase located in the inner mitochondrial membrane, and this drives ATP synthesis. Approximately three ATP molecules are synthesized per NADH oxidized and approximately two ATPs are synthesized per FADH2 oxidized. 

In the chemiosmotic theory for oxidative phosphorylation (Chap. 14), electron flow in the electron-transport chain is coupled to the generation of a proton concentration gradient across the inner mitochondrial membrane. Derive an expression for the difference in electrochemical potential for a proton across the membrane. 

The idea that oxidative phosphorylation and photophosphorylation systems are coupled with the transfer of a proton through the membrane was introduced by Mitchell (1966) and is now widely accepted. H+-ATPase (ATP synthase, F,Fo-ATPase) catalyzes ATP synthesis coupled to an electrochemical gradient and ATP hydrolysis driven by proton translocation in mitochondrial or bacterial membranes. (Boyer, 2001 Babcock and Wikstroem, 1992 Abraham et al., 1994 Allison, 1998 Ogilvie et al. 1997 Musser and Theg, 2000 Backgren et al., 2000 Arechada and Jones, 2001 Gibbsons et. al., 2000 and references therein). The enzyme from Escherichia coli consists of two parts, a water-... 

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. 

If the inner mitochondrial membrane is damaged such that it no longer provides a barrier to the movement of protons, then the proton electrochemical gradient is destroyed and oxidative phosphorylation will not occur. Uncoupling agents, such as 2,4-dinitrophenol, which permeabilize the inner mitochondrial membrane to protons, also inhibits oxidative phosphorylation for the same reason. 

In phase 2 of cellular respiration, the energy derived from fuel oxidation is converted to the high-energy phosphate bonds of ATP by the process of oxidative phosphorylation (see Fig. 2). Electrons are transferred from NADH and FAD(2H) to O2 by the electron transport chain, a series of electron transfer proteins that are located in the inner mitochondrial membrane. Oxidation of NADH and FAD(2H) by O2 generates an electrochemical potential across the inner mitochondrial membrane in the form of a transmembrane proton gradient (Ap). This electrochemical potential drives the synthesis of ATP form ADP and Pi by a transmembrane enzyme called ATP synthase (or FoFjATPase). 

Fig. 19.8. Overview of energy transformations in oxidative phosphorylation. The electrochemical potential gradient across the mitochondrial membrane is represented by ApH, the proton gradient, and A F, the membrane potential. The role of the electrochemical potential in oxidative phosphorylation is discussed in more depth in Chapter 21.

Our understanding of oxidative phosphorylation is based on the chemiosmotic hypothesis, which proposes that the energy for ATP synthesis is provided by an electrochemical gradient across the inner mitochondrial membrane. This electrochemical gradient is generated by the components of the electron transport chain, which pump protons across the inner mitochondrial membrane as they sequentially accept and donate electrons (see Fig. 21.1). The final acceptor is O2, which is reduced to H2O. 

In the cell, electron flow in the electron transport chain must be sequential from NADH or a flavoprotein all the way to O2 to generate ATP (see Fig. 21.5). In the absence of O2, there is no ATP generated from oxidative phosphorylation because electrons back up in the chain. Even complex I cannot pump protons to generate the electrochemical gradient, because every molecule of CoQ already has electrons that it cannot pass down the chain without an O2 to accept them at the end. The action of the respiratory chain inhibitor cyanide, which binds to cytochrome oxidase, is similar to that of anoxia it prevents proton pumping by all three complexes. Complete inhibition of the b-c complex prevents pumping at cytochrome... 

The electrochemical gradient couples the rate of the electron transport chain to the rate of ATP synthesis. Because electron flow requires proton pumping, electron flow cannot occur faster than protons are used for ATP synthesis (coupled oxidative phosphorylation) or returned to the matrix by a mechanism that short circuits the ATP synthase pore (uncoupling). 

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