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The electron transport chain in the mitochondrion is the site of oxidative phosphorylation in eukaryotes. The NADH and succinate generated in the citric acid cycle is oxidized, releasing energy to power the ATP synthase.
Oxidative phosphorylation is a metabolic pathway that uses energy released by the oxidation of nutrients to produce adenosine triphosphate (ATP). Although the many forms of life on earth use a range of different nutrients, almost all carry out oxidative phosphorylation to produce ATP, the molecule that supplies energy to metabolism. This pathway is probably so pervasive because it is a highly-efficient way of storing energy, compared to alternative fermentation processes such as glycolysis.
During oxidative phosphorylation, electrons are transferred from electron donors to electron acceptors such as oxygen, in a redox reaction. These redox reactions release energy, which is used to form ATP. In eukaryotes, these redox reactions are carried out by a series of protein complexes within mitochondria, whereas, in prokaryotes, these proteins are located in the cells\' inner membranes. These linked sets of enzymes are called electron transport chains. In eukaryotes, five main protein complexes are involved, whereas in prokaryotes many different enzymes are present, using a variety of electron donors and acceptors.
The energy released as electrons flow through this electron transport chain is used to transport protons across the inner mitochondrial membrane, in a process called chemiosmosis. This generates potential energy in the form of a pH gradient and an electrical potential across this membrane. This store of energy is tapped by allowing protons to flow back across the membrane and down this gradient, through a large enzyme called ATP synthase. This enzyme uses this energy to generate ATP from adenosine diphosphate (ADP), in a phosphorylation reaction. Unusually, the ATP synthase is driven by the proton flow which forces the rotation of a part of the enzyme—it is a rotary mechanical motor.
Although oxidative phosphorylation is a vital part of metabolism, it produces reactive oxygen species such as superoxide and hydrogen peroxide that lead to propagation of free-radicals, damaging cells and contributing to aging and disease. The enzymes carrying out this metabolic pathway are also the target of many drugs and poisons that inhibit their activities.
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Oxidative phosphorylation works by using energy-releasing chemical reactions to drive energy-requiring reactions: The two sets of reactions are said to be coupled. This means one cannot occur without the other. The flow of electrons through the electron transport chain, from electron donors such as NADH to electron acceptors such as oxygen, is an exergonic process – it releases energy, whereas the synthesis of ATP is an endergonic process that requires an input of energy.
Both the electron transport chain and the ATP synthase are embedded in a membrane, and energy is transferred from electron transport chain to the ATP synthase by movements of protons across this membrane, in a process called chemiosmosis.Mitchell P, Moyle J (1967). "Chemiosmotic hypothesis of oxidative phosphorylation". Nature 213 (5072): 137–9. PMID 4291593. In practice, this is like a simple electric circuit, with a current of protons being driven from the negative N-side of the membrane to the positive P-side by the proton-pumping enzymes of the electron transport chain. These enzymes are like a battery, as they perform work to drive current through the circuit. The movement of protons creates an electrochemical gradient across the membrane, which is often called the proton-motive force. This gradient has two components: a difference in proton concentration (a pH gradient) and a difference in electric potential, with the N-side having a negative charge. The energy is stored largely as the difference of electric potentials in mitochondria, but as a pH gradient in chloroplasts.Dimroth P, Kaim G, Matthey U (2000). "Crucial role of the membrane potential for ATP synthesis by F(1)F(o) ATP synthases". J. Exp. Biol. 203 (Pt 1): 51–9. PMID 10600673.
ATP synthase releases this stored energy by completing the circuit and allowing protons to flow down the electrochemical gradient, back to the N-side of the membrane.Schultz B, Chan S (2001). "Structures and proton-pumping strategies of mitochondrial respiratory enzymes". Annu Rev Biophys Biomol Struct 30: 23–65. PMID 11340051. This enzyme is like an electric motor as it uses the proton-motive force to drive the rotation of part of its structure and couples this motion to the synthesis of ATP.
The amount of energy released by oxidative phosphorylation is high, compared with the amount produced by anaerobic fermentation. Glycolysis produces only 2 ATP molecules, but 26 ATPs are produced by the oxidative phosphorylation of the 10 NADH and 2 succinate molecules made by converting one molecule of glucose to carbon dioxide and water.Rich PR (2003). "The molecular machinery of Keilin\'s respiratory chain". Biochem. Soc. Trans. 31 (Pt 6): 1095–105. PMID 14641005. This ATP yield is the theoretical maximum value; in practice, some protons leak across the membrane, lowering the yield of ATP.Porter RK, Brand MD (1995). "Mitochondrial proton conductance and H+/O ratio are independent of electron transport rate in isolated hepatocytes". Biochem. J. 310 ( Pt 2): 379–82. PMID 7654171.
Reduction of coenzyme Q from its ubiquinone form (Q) to the reduced ubiquinol form (QH2).
The electron transport chain carries both protons and electrons, passing electrons from donors to acceptors, and transporting protons across a membrane. These processes use both soluble and protein-bound transfer molecules. In mitochondria, electrons are transferred within the intermembrane space by the water-soluble electron transfer protein cytochrome c.Mathews FS (1985). "The structure, function and evolution of cytochromes". Prog. Biophys. Mol. Biol. 45 (1): 1–56. PMID 3881803. This carries only electrons, and these are transferred by the reduction and oxidation of an iron atom that the protein holds within a heme group in its structure. Cytochrome c is also found in some bacteria, where it is located within the periplasmic space.Wood PM (1983). "Why do c-type cytochromes exist?". FEBS Lett. 164 (2): 223–6. PMID 6317447.
Within the inner mitochondrial membrane, the lipid-soluble electron carrier coenzyme Q10 (Q) carries both electrons and protons by a redox cycle.Crane FL (2001). "Biochemical functions of coenzyme Q10". Journal of the American College of Nutrition 20 (6): 591–8. PMID 11771674. This small benzoquinone molecule is very hydrophobic, so it diffuses freely within the membrane. When Q accepts two electrons and two protons, it becomes reduced to the ubiquinol form (QH2); when QH2 releases two electrons and two protons, it becomes oxidized back to the ubiquinone (Q) form. As a result, if two enzymes are arranged so that Q is reduced on one side of the membrane and QH2 oxidized on the other, ubiquinone will couple these reactions and shuttle protons across the membrane.Mitchell P (1979). "Keilin\'s respiratory chain concept and its chemiosmotic consequences". Science 206 (4423): 1148–59. PMID 388618. Some bacterial electron transport chains use different quinones, such as menaquinone, in addition to ubiquinone.Søballe B, Poole RK (1999). "Microbial ubiquinones: multiple roles in respiration, gene regulation and oxidative stress management". Microbiology (Reading, Engl.) 145 ( Pt 8): 1817–30. PMID 10463148.
Within proteins, electrons are transferred between flavin cofactors Johnson D, Dean D, Smith A, Johnson M (2005). "Structure, function, and formation of biological iron-sulfur clusters". Annu Rev Biochem 74: 247–81. PMID 15952888. , iron–sulfur clusters, and cytochromes. There are several types of iron–sulfur cluster. The simplest kind found in the electron transfer chain consists of two iron atoms joined by two atoms of inorganic sulfur; these are called [2Fe–2S] clusters. The second kind, called [4Fe–4S], contains a cube of four iron atoms and four sulfur atoms. Each iron atom in these clusters is coordinated by an additional amino acid, usually by the sulfur atom of cysteine. Metal ion cofactors undergo redox reactions without binding or releasing protons, so in the electron transport chain they serve solely to transport electrons through proteins. Electrons move through quite long distances in proteins by hopping along between chains of these cofactors.Page CC, Moser CC, Chen X, Dutton PL (1999). "Natural engineering principles of electron tunnelling in biological oxidation-reduction". Nature 402 (6757): 47–52. PMID 10573417. This occurs by quantum tunnelling, which is rapid over distances of less than 14 Å.Leys D, Scrutton NS (2004). "Electrical circuitry in biology: emerging principles from protein structure". Curr. Opin. Struct. Biol. 14 (6): 642–7. PMID 15582386.
Many catabolic biochemical processes, such as glycolysis, the citric acid cycle and beta oxidation, produce the reduced coenzyme NADH. This coenzyme contains electrons that have a high transfer potential; in other words, they will release a large amount of energy upon oxidation. However, the cell does not release this energy all at once, as this would be an uncontrollable reaction. Instead, the electrons are removed from NADH and passed to oxygen through a series of enzymes that each release a small amount of the energy. This set of enzymes, consisting of complexes I through IV, is called the electron transport chain and is found in the inner membrane of the mitochondrion. Succinate is also oxidized by the electron transport chain, but feeds into the pathway at a different point.
In eukaryotes, the enzymes in this electron transport system use the energy released from the oxidation of NADH to pump protons across the inner membrane of the mitochondrion. This causes protons to build up in the intermembrane space, and generates an electrochemical gradient across the membrane. The energy stored in this potential is then used by ATP synthase to produce ATP. Oxidative phosphorylation in the eukaryotic mitochondrion is the best-understood example of this process. The mitochondrion is present in almost all eukaryotes, with the exception of anaerobic protozoa such as Trichomonas vaginalis that instead reduce protons to hydrogen in a remnant mitochondrion called a hydrogenosome.Boxma B, de Graaf RM, van der Staay GW, et al (2005). "An anaerobic mitochondrion that produces hydrogen". Nature 434 (7029): 74–9. PMID 15744302.
Complex I or NADH-Q oxidoreductase. The abbreviations are discussed in the text. In all diagrams of respiratory complexes, the matrix is at the bottom, with the intermembrane space above.
NADH-coenzyme Q oxidoreductase, also known as NADH dehydrogenase or complex I, is the first protein in the electron transport chain.Hirst J (2005). "Energy transduction by respiratory complex I--an evaluation of current knowledge". Biochem. Soc. Trans. 33 (Pt 3): 525–9. PMID 15916556. Complex I is a giant enzyme with the mammalian complex I having 46 subunits and a molecular mass of about 1,000 kilodaltons (kDa).Lenaz G, Fato R, Genova M, Bergamini C, Bianchi C, Biondi A (2006). "Mitochondrial Complex I: structural and functional aspects". Biochim Biophys Acta 1757 (9–10): 1406–20. PMID 16828051. The structure is known in detail only from a bacterium Sazanov L.A., Hinchliffe P. (2006) Structure of the hydrophilic domain of respiratory complex I from Thermus thermophilus. Science 311, 1430-1436 ; in most organisms the complex resembles a boot with a large “ball” poking out from the membrane into the mitochondrion.Baranova EA, Holt PJ, Sazanov LA (2007). "Projection structure of the membrane domain of Escherichia coli respiratory complex I at 8 A resolution". J. Mol. Biol. 366 (1): 140–54. PMID 17157874. Friedrich T, Böttcher B (2004). "The gross structure of the respiratory complex I: a Lego System". Biochim. Biophys. Acta 1608 (1): 1–9. PMID 14741580. The genes that encode the individual proteins are contained in both the cell nucleus and the mitochondrial genome, as is the case for many enzymes present in the mitochondrion.
The reaction which is catalyzed by this enzyme is the two electron reduction by NADH of coenzyme Q10 or ubiquinone (represented as Q in the equation below), a lipid-soluble quinone that is found in the mitochondrion membrane:
The start of the reaction, and indeed of the entire electron chain, is the binding of a NADH molecule to complex I and the donation of two electrons. The electrons enter complex I via a prosthetic group attached to the complex, flavin mononucleotide (FMN). The addition of electrons to FMN converts it to its reduced form, FMNH2. The electrons are then transferred through a series of iron–sulfur clusters: the second kind of prosthetic group present in the complex.Sazanov L.A., Hinchliffe P. (2006) Structure of the hydrophilic domain of respiratory complex I from Thermus thermophilus. Science 311, 1430-1436 There are both [2Fe–2S] and [4Fe–4S] iron–sulfur clusters in complex I.
As the electrons pass through this complex, four protons are pumped from the matrix into the intermembrane space. Exactly how this occurs is unclear, but it seems to involve conformational changes in complex I that cause the protein to bind protons on the N-side of the membrane and then move them onto the P-side of the membrane.Brandt U, Kerscher S, Dröse S, Zwicker K, Zickermann V (2003). "Proton pumping by NADH:ubiquinone oxidoreductase. A redox driven conformational change mechanism?". FEBS Lett. 545 (1): 9–17. PMID 12788486. Finally, the electrons are transferred from the chain of iron–sulfur clusters to a ubiquinone molecule in the membrane. Reduction of ubiquinone also contributes to the generation of a proton gradient, as two protons are taken up from the matrix as it is reduced to ubiquinol (QH2).
Complex II: Succinate-Q oxidoreductase.
Succinate-Q oxidoreductase, also known as complex II, is a second entry point to the electron transport chain.Cecchini G (2003). "Function and structure of complex II of the respiratory chain". Annu Rev Biochem 72: 77–109. PMID 14527321. It is unusual as it is the only enzyme that participates in both the citric acid cycle and the electron transport chain. Complex II consists of four protein subunits and contains a bound flavin adenine dinucleotide (FAD) cofactor, iron–sulfur clusters, and a heme group that does not participate in electron transfer to coenzyme Q, but is believed to be important in decreasing production of reactive oxygen species. Yankovskaya V., Horsefield R., Tornroth S., Luna-Chavez C., Miyoshi H., Leger C., Byrne B., Cecchini G., Iwata S. (2003) Architecture of succinate dehydrogenase and reactive oxygen species generation. Science 299, 700-704 Horsefield R, Iwata S, Byrne B (2004). "Complex II from a structural perspective". Curr. Protein Pept. Sci. 5 (2): 107–18. PMID 15078221. It oxidizes succinate to fumarate and reduces ubiquinone. As this reaction releases less energy than the oxidation of NADH, complex II does not transport protons across the membrane and does not contribute to the proton gradient.
In some eukaryotes, such as the parasitic worm Ascaris suum, an enzyme similar to complex II, fumarate reductase (menaquinol:fumarate oxidoreductase, or QFR) operates in reverse to oxidize ubiquinol and reduce fumarate. This allows the worm to survive in the anaerobic environment of the large intestine, carrying out anaerobic oxidative phosphorylation with fumarate as the electron acceptor.Kita K, Hirawake H, Miyadera H, Amino H, Takeo S (2002). "Role of complex II in anaerobic respiration of the parasite mitochondria from Ascaris suum and Plasmodium falciparum". Biochim. Biophys. Acta 1553 (1–2): 123–39. PMID 11803022. Another unconventional function of complex II is seen in the malaria parasite Plasmodium falciparum. Here, the reversed action of complex II as an oxidase is important in regenerating ubiquinol, which the parasite uses in an unusual form of pyrimidine biosynthesis.Painter HJ, Morrisey JM, Mather MW, Vaidya AB (2007). "Specific role of mitochondrial electron transport in blood-stage Plasmodium falciparum". Nature 446 (7131): 88–91. PMID 17330044.
Electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-Q oxidoreductase), also known as electron transferring-flavoprotein dehydrogenase, is a third entry point to the electron transport chain. It is an enzyme that accepts electrons from electron-transferring flavoprotein in the mitochondrial matrix, and uses these electrons to reduce ubiquinone.Ramsay RR, Steenkamp DJ, Husain M (1987). "Reactions of electron-transfer flavoprotein and electron-transfer flavoprotein: ubiquinone oxidoreductase". Biochem. J. 241 (3): 883–92. PMID 3593226. This enzyme contains a flavin and a [4Fe–4S] cluster, but unlike the other respiratory complexes, it attaches to the surface of the membrane and does not cross the lipid bilayer.Zhang J, Frerman FE, Kim JJ (2006). "Structure of electron transfer flavoprotein-ubiquinone oxidoreductase and electron transfer to the mitochondrial ubiquinone pool". Proc. Natl. Acad. Sci. U.S.A. 103 (44): 16212–7. doi:10.1073/pnas.0604567103. PMID 17050691.
In mammals, this metabolic pathway is important in beta oxidation of fatty acids and catabolism of amino acids and choline, as it accepts electrons from multiple acetyl-CoA dehydrogenases.Ikeda Y, Dabrowski C, Tanaka K (1983). "Separation and properties of five distinct acyl-CoA dehydrogenases from rat liver mitochondria. Identification of a new 2-methyl branched chain acyl-CoA dehydrogenase". J. Biol. Chem. 258 (2): 1066–76. PMID 6401712. Ruzicka FJ, Beinert H (1977). "A new iron-sulfur flavoprotein of the respiratory chain. A component of the fatty acid beta oxidation pathway". J. Biol. Chem. 252 (23): 8440–5. PMID 925004. In plants, ETF-Q oxidoreductase is also important in the metabolic responses that allow survival in extended periods of darkness.Ishizaki K, Larson TR, Schauer N, Fernie AR, Graham IA, Leaver CJ (2005). "The critical role of Arabidopsis electron-transfer flavoprotein:ubiquinone oxidoreductase during dark-induced starvation". Plant Cell 17 (9): 2587–600. PMID 16055629.
Q-cytochrome c oxidoreductase is also known as cytochrome c reductase, cytochrome bc1 complex, or simply complex III.Berry E, Guergova-Kuras M, Huang L, Crofts A (2000). "Structure and function of cytochrome bc complexes". Annu Rev Biochem 69: 1005–75. PMID 10966481. Crofts AR (2004). "The cytochrome bc1 complex: function in the context of structure". Annu. Rev. Physiol. 66: 689–733. PMID 14977419. In mammals, this enzyme is a dimer, with each subunit complex containing 11 protein subunits, an [2Fe-2S] iron–sulfur cluster and three cytochromes; one cytochrome c1 and two b cytochromes.Iwata S, Lee JW, Okada K, et al (1998). "Complete structure of the 11-subunit bovine mitochondrial cytochrome bc1 complex". Science 281 (5373): 64–71. PMID 9651245. A cytochrome is a kind of electron-transferring protein that contains at least one heme group. The iron atoms inside complex III’s heme groups alternate between a reduced ferrous (+2) and oxidized ferric (+3) state as the electrons are transferred through the protein.
The reaction catalyzed by complex III is the oxidation of one molecule of ubiquinol and the reduction of two molecules of cytochrome c, a heme protein loosely associated with the mitochondrion. Unlike coenzyme Q, which carries two electrons, cytochrome c carries only one electron.
As only one of the electrons can be transferred from the QH2 donor to a cytochrome c acceptor at a time, the reaction mechanism of complex III is more elaborate than those of the other respiratory complexes, and occurs in two steps called the Q cycle.Trumpower BL (1990). "The protonmotive Q cycle. Energy transduction by coupling of proton translocation to electron transfer by the cytochrome bc1 complex". J. Biol. Chem. 265 (20): 11409–12. PMID 2164001. In the first step, the enzyme binds three substrates, first, QH2, which is then oxidized, with one electron being passed to the second substrate, cytochrome c. The two protons released from QH2 pass into the intermembrane space. The third substrate is Q, which accepts the second electron from the QH2 and is reduced to Q.-, which is the ubisemiquinone free-radical. The first two substrates are released, but this ubisemiquinone intermediate remains bound. In the second step, a second molecule of QH2 is bound and again passes its first electron to a cytochrome c acceptor. The second electron is passed to the bound ubisemiquinone, reducing it to QH2 as it gains two protons from the mitochondrial matrix. This QH2 is then released from the enzyme.Hunte C, Palsdottir H, Trumpower BL (2003). "Protonmotive pathways and mechanisms in the cytochrome bc1 complex". FEBS Lett. 545 (1): 39–46. PMID 12788490.
As coenzyme Q is reduced to ubiquinol on the inner side of the membrane and oxidized to ubiquinone on the other, this causes the net transfer of protons across the membrane, adding to the proton gradient. The rather complex two-step mechanism by which this occurs is important as it increases the efficiency of proton transfer. If instead of the Q cycle, one molecule of QH2 was used to directly reduce two molecules of cytochrome c, the efficiency would be halved, with only one proton transferred per cytochrome c reduced.
Complex IV: cytochrome c oxidase.
Cytochrome c oxidase, also known as complex IV, is the final protein complex in the electron transport chain.Calhoun M, Thomas J, Gennis R (1994). "The cytochrome oxidase superfamily of redox-driven proton pumps". Trends Biochem Sci 19 (8): 325–30. PMID 7940677. The mammalian enzyme has an extremely complex structure and contains 13 subunits, two heme groups, as well as multiple metal ion cofactors –- in all three atoms of copper, one of magnesium and one of zinc.Tsukihara T, Aoyama H, Yamashita E, Tomizaki T, Yamaguchi H, Shinzawa-Itoh K, Nakashima R, Yaono R, Yoshikawa S. (1996). "TThe whole structure of the 13-subunit oxidized cytochrome c oxidase at 2.8 A.". Science 272 (5265): 1136–44. PMID 8638158.
This enzyme mediates the final reaction in the electron transport chain and transfers electrons to oxygen, while pumping protons across the membrane.Yoshikawa S, Muramoto K, Shinzawa-Itoh K, et al (2006). "Proton pumping mechanism of bovine heart cytochrome c oxidase". Biochim. Biophys. Acta 1757 (9–10): 1110–6. PMID 16904626. The final electron acceptor oxygen, which is also called the terminal electron acceptor, is reduced to water in this step. Both the direct pumping of protons and the consumption of matrix protons in the reduction of oxygen contribute to the proton gradient. The reaction catalyzed is the oxidation of cytochrome c and the reduction of oxygen:
For a detailed discussion of the mechanism of reduction of oxygen, and a better figure illustrating the redox centers in the protein, see the linked article Cytochrome c oxidase.
Many eukaryotic organisms have electron transport chains that differ from the well-studied mammalian enzymes described above. For example, in plants, alternative NADH oxidases exist that oxidize NADH in the cytosol, rather than the mitochondrial matrix, and pass these electrons to the ubiquinone pool.Rasmusson AG, Soole KL, Elthon TE (2004). "Alternative NAD(P)H dehydrogenases of plant mitochondria". Annual review of plant biology 55: 23–39. PMID 15725055. These enzymes do not transport protons and therefore reduce ubiquinone without altering the electrochemical gradient across the inner membrane.Menz RI, Day DA (1996). "Purification and characterization of a 43-kDa rotenone-insensitive NADH dehydrogenase from plant mitochondria". J. Biol. Chem. 271 (38): 23117–20. PMID 8798503.
Another example of a divergent electron transport chain is the alternative oxidase, which is found in plants, as well as some fungi, protists, and possibly some animals.McDonald A, Vanlerberghe G (2004). "Branched mitochondrial electron transport in the Animalia: presence of alternative oxidase in several animal phyla". IUBMB Life 56 (6): 333–41. PMID 15370881. Sluse FE, Jarmuszkiewicz W (1998). "Alternative oxidase in the branched mitochondrial respiratory network: an overview on structure, function, regulation, and role". Braz. J. Med. Biol. Res. 31 (6): 733–47. PMID 9698817. This enzyme transfers electrons directly from ubiquinol to oxygen.Moore AL, Siedow JN (1991). "The regulation and nature of the cyanide-resistant alternative oxidase of plant mitochondria". Biochim. Biophys. Acta 1059 (2): 121–40. PMID 1883834.
The electron transport pathways produced by these alternative NADH and ubiquinone oxidases have lower ATP yields than the full pathway. The advantages produced by a shortened pathway are not entirely clear. However, the alternative oxidase is produced in response to stresses such as cold, reactive oxygen species and infection by pathogens, as well as other factors that inhibit the full electron transport chain.Vanlerberghe GC, McIntosh L (1997). "Alternative oxidase: From Gene to Function" 48: 703–34. PMID 15012279. Ito Y, Saisho D, Nakazono M, Tsutsumi N, Hirai A (1997). "Transcript levels of tandem-arranged alternative oxidase genes in rice are increased by low temperature". Gene 203 (2): 121–9. PMID 9426242. Alternative pathways might therefore enhance an organisms\' resistance to injury, by reducing oxidative stress.Maxwell DP, Wang Y, McIntosh L (1999). "The alternative oxidase lowers mitochondrial reactive oxygen production in plant cells". Proc. Natl. Acad. Sci. U.S.A. 96 (14): 8271–6. PMID 10393984.
The original model for how the respiratory chain complexes are organized was that they diffuse freely and independently in the mitochondrial membrane.Lenaz G (2001). "A critical appraisal of the mitochondrial coenzyme Q pool". FEBS Lett. 509 (2): 151-5. PMID 11741580. However, recent data suggest that the complexes might form higher-order structures called supercomplexes or "respirasomes".Heinemeyer J, Braun HP, Boekema EJ, Kouril R (2007). "A structural model of the cytochrome C reductase/oxidase supercomplex from yeast mitochondria". J. Biol. Chem. 282 (16): 12240–8. PMID 17322303. In this model, the various complexes exist as organized sets of interacting enzymes.Schägger H, Pfeiffer K (2000). "Supercomplexes in the respiratory chains of yeast and mammalian mitochondria". EMBO J. 19 (8): 1777–83. PMID 10775262. These associations might allow channeling of substrates between the various enzyme complexes, increasing the rate and efficiency of electron transfer.Schägger H (2002). "Respiratory chain supercomplexes of mitochondria and bacteria". Biochim. Biophys. Acta 1555 (1–3): 154–9. PMID 12206908. Within such mammalian supercomplexes, some components would be present in higher amounts than others, with a ratio between complexes I/II/III/IV and the ATP synthase of approximately 1:1:3:7:4.Schägger H, Pfeiffer K (2001). "The ratio of oxidative phosphorylation complexes I-V in bovine heart mitochondria and the composition of respiratory chain supercomplexes". J. Biol. Chem. 276 (41): 37861–7. PMID 11483615. However, the debate over this supercomplex model is not completely resolved, as some data do not appear to fit with this model.Gupte S, Wu ES, Hoechli L, et al (1984). "Relationship between lateral diffusion, collision frequency, and electron transfer of mitochondrial inner membrane oxidation-reduction components". Proc. Natl. Acad. Sci. U.S.A. 81 (9): 2606–10. PMID 6326133.
In contrast to the general similarity in structure and function of the electron transport chains in eukaryotes, bacteria and archaea possess a large variety of electron-transfer enzymes. These use an equally wide set of chemicals as substrates.Nealson KH (1999). "Post-Viking microbiology: new approaches, new data, new insights". Origins of life and evolution of the biosphere : the journal of the International Society for the Study of the Origin of Life 29 (1): 73–93. PMID 11536899. In common with eukaryotes, prokaryotic electron transport uses the energy released from the oxidation of a substrate to pump ions across a membrane and generate an electrochemical gradient. In the bacteria, oxidative phosphorylation in Escherichia coli is understood in most detail, while archaeal systems are at present poorly-understood.Schäfer G, Engelhard M, Müller V (1999). "Bioenergetics of the Archaea". Microbiol. Mol. Biol. Rev. 63 (3): 570-620. PMID 10477309.
The main difference between eukaryotic and prokaryotic oxidative phosphorylation is that bacteria and archaea use many different substances to donate or accept electrons. This allows prokaryotes to grow under a wide variety of environmental conditions.Ingledew WJ, Poole RK (1984). "The respiratory chains of Escherichia coli". Microbiol. Rev. 48 (3): 222–71. PMID 6387427. In E. coli, for example, oxidative phosphorylation can be driven by a large number of pairs of reducing agents and oxidizing agents, which are listed below. The midpoint potential of a chemical measures how much energy is released when it is oxidized or reduced, with reducing agents having negative potentials and oxidizing agents positive potentials.
As shown above, E. coli can grow with reducing agents such as formate, hydrogen or lactate as electron donors, and nitrate, DMSO or oxygen as acceptors. The larger the difference in midpoint potential between an oxidizing and reducing agent, the more energy is released when they react. Out of these compounds, the succinate/fumarate pair is unusual, as its midpoint potential is close to zero. Succinate can therefore be oxidized to fumarate if a strong oxidizing agent such as oxygen is available, or fumarate can be reduced to succinate using a strong reducing agent such as formate. These alternative reactions are catalyzed by succinate dehydrogenase and fumarate reductase, respectively.Cecchini G, Schröder I, Gunsalus RP, Maklashina E (2002). "Succinate dehydrogenase and fumarate reductase from Escherichia coli". Biochim. Biophys. Acta 1553 (1–2): 140–57. PMID 11803023.
Some prokaryotes use redox pairs that have only a small difference in midpoint potential. For example, nitrifying bacteria such as Nitrobacter oxidize nitrite to nitrate, donating the electrons to oxygen. The small amount of energy released in this reaction is enough to pump protons and generate ATP, but not enough to produce NADH or NADPH directly for use in anabolism.Freitag A, Bock E (1990). "Energy conservation in Nitrobacter". FEMS Microbiology Letters 66 (1–3): 157&ndash:62. doi:10.1111/j.1574-6968.1990.tb03989.x. This problem is solved by using a nitrite oxidoreductase to produce enough proton-motive force to run part of the electron transport chain in reverse, causing complex I to generate NADH.Starkenburg SR, Chain PS, Sayavedra-Soto LA, et al (2006). "Genome sequence of the chemolithoautotrophic nitrite-oxidizing bacterium Nitrobacter winogradskyi Nb-255". Appl. Environ. Microbiol. 72 (3): 2050–63. PMID 16517654. Yamanaka T, Fukumori Y (1988). "The nitrite oxidizing system of Nitrobacter winogradskyi". FEMS Microbiol. Rev. 4 (4): 259–70. PMID 2856189.
Prokaryotes control their use of these electron donors and acceptors by varying which enzymes are produced, in response to environmental conditions.Iuchi S, Lin EC (1993). "Adaptation of Escherichia coli to redox environments by gene expression". Mol. Microbiol. 9 (1): 9–15. PMID 8412675. This flexibility is possible because different oxidases and reductases use the same ubiquinone pool. This allows many combinations of enzymes to function together, linked by the common ubiquinol intermediate. These respiratory chains therefore have a modular design, with easily interchangeable sets of enzyme systems.
In addition to this metabolic diversity, prokaryotes also possess a range of isozymes – different enzymes that catalyze the same reaction. For example, in E. coli there are two different types of ubiquinol oxidase using oxygen as an electron acceptor. Under highly-aerobic conditions, the cell uses an oxidase with a low affinity for oxygen that can transport two protons per electron. However, if levels of oxygen fall, they switch to an oxidase that only transfers one proton per electron, but has a high affinity for oxygen.Calhoun MW, Oden KL, Gennis RB, de Mattos MJ, Neijssel OM (1993). "Energetic efficiency of Escherichia coli: effects of mutations in components of the aerobic respiratory chain". J. Bacteriol. 175 (10): 3020–5. PMID 8491720.
ATP synthase, also called complex V, is the final enzyme in the oxidative phosphorylation pathway. This enzyme is found in all forms of life and functions in the same way in both prokaryotes or eukaryotes.Boyer PD (1997). "The ATP synthase--a splendid molecular machine". Annu. Rev. Biochem. 66: 717–49. PMID 9242922. The enzyme uses the energy stored in a proton gradient across a membrane to drive the synthesis of ATP from ADP and phosphate (Pi). Estimates of the number of protons required to synthesise one ATP have ranged from three to four,Van Walraven HS, Strotmann H, Schwarz O, Rumberg B (1996). "The H+/ATP coupling ratio of the ATP synthase from thiol-modulated chloroplasts and two cyanobacterial strains is four". FEBS Lett. 379 (3): 309-13. PMID 8603713. Yoshida M, Muneyuki E, Hisabori T (2001). "ATP synthase--a marvellous rotary engine of the cell". Nat. Rev. Mol. Cell Biol. 2 (9): 669-77. PMID 11533724. with some suggesting cells can vary this ratio, to suit different conditions.Schemidt RA, Qu J, Williams JR, Brusilow WS (1998). "Effects of carbon source on expression of F0 genes and on the stoichiometry of the c subunit in the F1F0 ATPase of Escherichia coli". J. Bacteriol. 180 (12): 3205-8. PMID 9620972.
This phosphorylation reaction is an equilibrium, which can be shifted by altering the proton-motive force. In the absence of a proton-motive force, the ATP synthase reaction will run from right to left, hydrolyzing ATP and pumping protons out of the matrix across the membrane. However, when the proton-motive force is high, the reaction is forced to run in the opposite direction; it proceeds from left to right, allowing protons to flow down their concentration gradient and turning ADP into ATP. Indeed, in the closely related vacuolar type H+-ATPases the same reaction is used to acidify cellular compartments, by pumping protons and hydrolysing ATP.Nelson N, Perzov N, Cohen A, Hagai K, Padler V, Nelson H (2000). "The cellular biology of proton-motive force generation by V-ATPases". J. Exp. Biol. 203 (Pt 1): 89–95. PMID 10600677.
ATP synthase is a massive protein complex with a mushroom-like shape. The mammalian enzyme complex contains 16 subunits and has a mass of approximately 600 kilodaltons.Rubinstein JL, Walker JE, Henderson R (2003). "Structure of the mitochondrial ATP synthase by electron cryomicroscopy". EMBO J. 22 (23): 6182–92. PMID 14633978. The portion embedded within the membrane is called FO and contains a ring of c subunits and the proton channel. The stalk and the ball-shaped headpiece is called F1 and is the site of ATP synthesis. The ball-shaped complex at the end of the F1 portion contains six proteins of two different kinds (three α subunits and three β subunits), whereas the "stalk" consists of one protein: the γ subunit, with the tip of the stalk extending into the ball of α and β subunits.Leslie AG, Walker JE (2000). "Structural model of F1-ATPase and the implications for rotary catalysis". Philos. Trans. R. Soc. Lond., B, Biol. Sci. 355 (1396): 465–71. PMID 10836500. Both the α and β subunits bind nucleotides, but only the β subunits catalyze the ATP synthesis reaction. Reaching along the side of the F1 portion and back into the membrane is a long rod-like subunit that anchors the α and β subunits into the base of the enzyme.
As protons cross the membrane through the channel in the base of ATP synthase this causes the FO proton-driven motor to rotate.Noji H, Yoshida M (2001). "The rotary machine in the cell, ATP synthase". J. Biol. Chem. 276 (3): 1665-8. PMID 11080505. Rotation might be caused by changes in the ionization of amino acids in the ring of c subunits causing electrostatic interactions that propel the ring of c subunits past the proton channel.Capaldi R, Aggeler R (2002). "Mechanism of the F(1)F(0)-type ATP synthase, a biological rotary motor". Trends Biochem Sci 27 (3): 154–60. PMID 11893513. This rotating ring in turn drives the rotation of the central axle (the γ subunit stalk) within the α and β subunits. The α and β subunits are prevented from rotating themselves by the side-arm, which acts as a stator. This movement of the tip of the γ subunit within the ball of α and β subunits provides the energy for the active sites in the β subunits to undergo a cycle of movements that produces and then releases ATP.Dimroth P, von Ballmoos C, Meier T (2006). "Catalytic and mechanical cycles in F-ATP synthases. Fourth in the Cycles Review Series". EMBO Rep 7 (3): 276–82. PMID 16607397.
This ATP synthesis reaction is called the binding change mechanism and involves the active site of a β subunit cycling between three states.Gresser MJ, Myers JA, Boyer PD (1982). "Catalytic site cooperativity of beef heart mitochondrial F1 adenosine triphosphatase. Correlations of initial velocity, bound intermediate, and oxygen exchange measurements with an alternating three-site model". J. Biol. Chem. 257 (20): 12030–8. PMID 6214554. In the "open" state, ADP and phosphate enter the active site (shown in brown in the diagram). The protein then closes up around the molecules and binds them loosely – the "loose" state (shown in red). The enzyme then changes shape again and forces these molecules together, with the active site in the resulting "tight" state (shown in pink) binding the newly-produced ATP molecule with very high affinity. Finally, the active site cycles back to the open state, releasing ATP and binding more ADP and phosphate, ready for the next cycle.
In some bacteria and archaea, ATP synthesis is driven by the movement of sodium ions through the cell membrane, rather than the movement of protons.Dimroth P (1994). "Bacterial sodium ion-coupled energetics". Antonie Van Leeuwenhoek 65 (4): 381–95. PMID 7832594. Becher B, Müller V (1994). "Delta mu Na+ drives the synthesis of ATP via a delta mu Na(+)-translocating F1F0-ATP synthase in membrane vesicles of the archaeon Methanosarcina mazei Gö1". J. Bacteriol. 176 (9): 2543–50.
