Bioenergetics: The Role of ATP PDF

S E C T I O N Bioenergetics III 11 C...

S E C T I O N Bioenergetics III 11 C H A P T E R Bioenergetics: The Role of ATP Kathleen M. Botham, PhD, DSc & Peter A. Mayes, PhD, DSc O B J EC T IVES State the first and second laws of thermodynamics and understand how they apply to biologic systems. After studying this chapter, Explain what is meant by the terms free energy, entropy, enthalpy, exergonic, you should be able to: and endergonic. Appreciate how reactions that are endergonic may be driven by coupling to those that are exergonic in biologic systems. Explain the role of group transfer potential, adenosine triphosphate (ATP), and other nucleotide triphosphates in the transfer of free energy from exergonic to endergonic processes, enabling them to act as the “energy currency” of cells. BIOMEDICAL IMPORTANCE FREE ENERGY IS THE USEFUL Bioenergetics, or biochemical thermodynamics, is the study ENERGY IN A SYSTEM of the energy changes accompanying biochemical reactions. Gibbs change in free energy (ΔG) is that portion of the Biologic systems are essentially isothermic and use chemical total energy change in a system that is available for doing energy to power living processes. The way in which an ani- work—that is, the useful energy, also known as the chemical mal obtains suitable fuel from its food to provide this energy potential. is basic to the understanding of normal nutrition and metab- olism. Death from starvation occurs when available energy reserves are depleted, and certain forms of malnutrition are Biologic Systems Conform to the associated with energy imbalance (marasmus). Thyroid hor- General Laws of Thermodynamics mones control the metabolic rate (rate of energy release), and The first law of thermodynamics states that the total energy disease results if they malfunction. Excess storage of surplus of a system, including its surroundings, remains constant. energy causes obesity, an increasingly common disease of It implies that within the total system, energy is neither lost Western society which predisposes to many diseases, includ- nor gained during any change. However, energy may be ing cardiovascular disease and diabetes mellitus type 2, and transferred from one part of the system to another, or may be lowers life expectancy. transformed into another form of energy. In living systems, 105 106 SECTION III Bioenergetics chemical energy may be transformed into heat or into electri- A cal, radiant, or mechanical energy. The second law of thermodynamics states that the total Heat Ex entropy of a system must increase if a process is to occur er go D spontaneously. Entropy is the extent of disorder or random- ni c ness of the system and becomes maximum as equilibrium is Free energy approached. Under conditions of constant temperature and pressure, the relationship between the free-energy change (ΔG) of a reacting system and the change in entropy (ΔS) is Chemical expressed by the following equation, which combines the two n ic energy laws of thermodynamics: rgo de En ∆G = ∆H −T∆S where ΔH is the change in enthalpy (heat) and T is the abso- C B lute temperature. A+C B + D + Heat In biochemical reactions, since ΔH is approximately equal to the total change in internal energy of the reaction or ΔE, FIGURE 11–1 Coupling of an exergonic to an endergonic the above relationship may be expressed in the following way: reaction. ∆G = ∆E −T∆S If ΔG is negative, the reaction proceeds spontaneously with may be represented as shown in Figure 11–1. The conversion loss of free energy, that is, it is exergonic. If, in addition, ΔG is of metabolite A to metabolite B occurs with release of free of great magnitude, the reaction goes virtually to completion energy and is coupled to another reaction in which free energy and is essentially irreversible. On the other hand, if ΔG is posi- is required to convert metabolite C to metabolite D. The terms tive, the reaction proceeds only if free energy can be gained, exergonic and endergonic, rather than the normal chemical that is, it is endergonic. If, in addition, the magnitude of ΔG is terms “exothermic” and “endothermic,” are used to indicate great, the system is stable, with little or no tendency for a reac- that a process is accompanied by loss or gain, respectively, of tion to occur. If ΔG is zero, the system is at equilibrium and no free energy in any form, not necessarily as heat. In practice, an net change takes place. endergonic process cannot exist independently, but must be a When the reactants are present in concentrations of component of a coupled exergonic–endergonic system where 1.0 mol/L, ΔG0 is the standard free-energy change. For bio- the overall net change is exergonic. The exergonic reactions chemical reactions, a standard state is defined as having a pH are termed catabolism (generally, the breakdown or oxidation of 7.0. The standard free-energy change at this standard state of fuel molecules), whereas the synthetic reactions that build is denoted by ΔG0′. up substances are termed anabolism. The combined catabolic The standard free-energy change can be calculated from and anabolic processes constitute metabolism. the equilibrium constant Keq. If the reaction shown in Figure 11–1 is to go from left to right, then the overall process must be accompanied by loss ∆G 0′ = − RT ln K eq ′ of free energy as heat. One possible mechanism of coupling where R is the gas constant and T is the absolute temperature could be envisaged if a common obligatory intermediate (I) (see Chapter 8). It is important to note that the actual ΔG may took part in both reactions, that is, be larger or smaller than ΔG0′ depending on the concentra- A+C→1→B+D tions of the various reactants, including the solvent, various ions, and proteins. Some exergonic and endergonic reactions in biologic systems In a biochemical system, an enzyme only speeds up the are coupled in this way. This type of system has a built-in mech- attainment of equilibrium; it never alters the final concentra- anism for biologic control of the rate of oxidative processes tions of the reactants at equilibrium. since the common obligatory intermediate allows the rate of utilization of the product of the synthetic path (D) to deter- mine by mass action the rate at which A is oxidized. Indeed, ENDERGONIC PROCESSES these relationships supply a basis for the concept of respiratory PROCEED BY COUPLING control, the process that prevents an organism from burning out of control. An extension of the coupling concept is pro- TO EXERGONIC PROCESSES vided by dehydrogenation reactions, which are coupled to The vital processes—for example, synthetic reactions, muscu- hydrogenations by an intermediate carrier (Figure 11–2). lar contraction, nerve impulse conduction, and active trans- An alternative method of coupling an exergonic to an end- port—obtain energy by chemical linkage, or coupling, to ergonic process is to synthesize a compound of high-energy oxidative reactions. In its simplest form, this type of coupling potential in the exergonic reaction and to incorporate this  CHAPTER 11 Bioenergetics: The Role of ATP 107 AH2 Carrier BH2 adenosine diphosphate (ADP), and inorganic phosphate (Pi) in glycolysis (see Chapter 17). A Carrier H2 B The Intermediate Value for the Free FIGURE 11–2 Coupling of dehydrogenation and hydrogena- tion reactions by an intermediate carrier. Energy of Hydrolysis of ATP Has Important Bioenergetic Significance The standard free energy of hydrolysis of a number of bio- new compound into the endergonic reaction, thus effecting chemically important phosphates is shown in Table 11–1. An a transference of free energy from the exergonic to the ender- estimate of the comparative tendency of each of the phosphate gonic pathway. The biologic advantage of this mechanism is groups to transfer to a suitable acceptor may be obtained that the compound of high potential energy, ~ , unlike I in from the ΔG0′ of hydrolysis at 37°C. This is termed the group the previous system, need not be structurally related to A, B, transfer potential. The value for the hydrolysis of the termi- C, or D, allowing to serve as a transducer of energy from a nal phosphate of ATP divides the list into two groups. Low- wide range of exergonic reactions to an equally wide range of energy phosphates, having a low group transfer potential, endergonic reactions or processes, such as biosyntheses, mus- exemplified by the ester phosphates found in the intermedi- cular contraction, nervous excitation, and active transport. In ates of glycolysis, have G0′ values smaller than that of ATP, the living cell, the principal high-energy intermediate or car- while in high-energy phosphates, with a more negative G0′, rier compound is ATP (Figure 11–3). the value is higher than that of ATP. The components of this latter group, including ATP, are usually anhydrides (eg, the 1-phosphate of 1,3-bisphosphoglycerate), enol phosphates HIGH-ENERGY PHOSPHATES (eg, phosphoenolpyruvate), and phosphoguanidines (eg, cre- PLAY A CENTRAL ROLE IN ENERGY atine phosphate, arginine phosphate). CAPTURE AND TRANSFER The symbol ~ indicates that the group attached to the bond, on transfer to an appropriate acceptor, results in trans- In order to maintain living processes, all organisms must obtain fer of the larger quantity of free energy. Thus, ATP has a high supplies of free energy from their environment. Autotrophic group transfer potential, whereas the phosphate in adenosine organisms utilize simple exergonic processes; for example, the energy of sunlight (green plants), the reaction Fe2+ → Fe3+ (some bacteria). On the other hand, heterotrophic organisms TABLE 11–1 Standard Free Energy of Hydrolysis of obtain free energy by coupling their metabolism to the break- Some Organophosphates of Biochemical Importance down of complex organic molecules in their environment. In ΔG0′ all these organisms, ATP plays a central role in the transference of free energy from the exergonic to the endergonic processes. Compound kJ/mol kcal/mol ATP is a nucleotide consisting of the nucleoside adenosine Phosphoenolpyruvate –61.9 –14.8 (adenine linked to ribose) and three phosphate groups (see Chapter 32). In its reactions in the cell, it functions as the Mg2+ Carbamoyl phosphate –51.4 –12.3 complex (Figure 11–3). 1,3-Bisphosphoglycerate –49.3 –11.8 The importance of phosphates in intermediary metabo- (to 3-phosphoglycerate) lism became evident with the discovery of the role of ATP, Creatine phosphate –43.1 –10.3 ATP → AMP + PPi –32.2 –7.7 NH2 ATP → ADP + Pi –30.5 –7.3 N N Glucose-1-phosphate –20.9 –5.0 Mg2+ PPi –19.2 –4.6 N N Fructose-6-phosphate –15.9 –3.8 O– O– O– –O Glucose-6-phosphate –13.8 –3.3 P O P O P O CH2 O O O O Glycerol-3-phosphate –9.2 –2.2 C C H H Abbreviations: PPi, pyrophosphate; Pi, inorganic orthophosphate. ATP H H Note: All values are taken from Jencks WP: Free energies of hydrolysis and decarbox- ylation. In: Handbook of Biochemistry and Molecular Biology, vol 1. Physical and OH OH Chemical Data. Fasman GD (editor). CRC Press, 1976:296-304, except that for PPi which is from Frey PA, Arabshahi A: Standard free-energy change for the hydrolysis of the alpha, beta-phosphoanhydride bridge in ATP. Biochemistry 1995;34:11307. FIGURE 11–3 Adenosine triphosphate (ATP) is shown as the Values differ between investigators, depending on the precise conditions under magnesium complex. which the measurements were made. 108 SECTION III Bioenergetics O– O– O– Phosphoenolpyruvate 1,3-Bisphosphoglycerate Succinyl- Oxidative Adenosine O P O P O P O– CoA phosphorylation O O O Creatine P P or Adenosine P P P P ) (Store of Adenosine triphosphate (ATP) Creatine ATP Adenosine P P Adenosine P ATP/ADP cycle Adenosine diphosphate (ADP) Adenosine monophosphate (AMP) P Glucose-6-phosphate ADP FIGURE 11–4 Structure of ATP, ADP, and AMP showing the Other phosphorylations, position and the number of high-energy phosphates (~ ). activations, Glucose-1,6- and endergonic Glycerol-3-phosphate bisphosphate processes monophosphate (AMP) is of the low-energy type since it is a FIGURE 11–6 Role of ATP/ADP cycle in transfer of high- normal ester link (Figure 11–4). In energy transfer reactions, energy phosphate. ATP may be converted to ADP and Pi or, in reactions requir- ing a greater energy input, to AMP + PPi (Table 11–1). The intermediate position of ATP allows it to play an ATP ACTS AS THE “ENERGY important role in energy transfer. The high free-energy CURRENCY” OF THE CELL change on hydrolysis of ATP is due to relief of charge repul- The high group transfer potential of ATP enables it to act as sion of adjacent negatively charged oxygen atoms and to sta- a donor of high-energy phosphate to form those compounds bilization of the reaction products, especially phosphate, as below it in Table 11–1. Likewise, with the necessary enzymes, resonance hybrids (Figure 11–5). Other “high-energy com- ADP can accept phosphate groups to form ATP from those pounds” are thiol esters involving coenzyme A (eg, acetyl- compounds above ATP in the table. In effect, an ATP/ADP CoA), acyl carrier protein, amino acid esters involved in cycle connects those processes that generate ~ to those pro- protein synthesis, S-adenosylmethionine (active methionine), cesses that utilize ~ (Figure 11–6), continuously consuming uridine diphosphate glucose (UDPGlc), and 5-phosphoribosyl- and regenerating ATP. This occurs at a very rapid rate since 1-pyrophosphate (PRPP). the total ATP/ADP pool is extremely small and sufficient to maintain an active tissue for only a few seconds. ADENOSINE There are three major sources of ~ taking part in energy conservation or energy capture: O– O– O– – O P O P O P O CH2 O 1. Oxidative phosphorylation is the greatest quantitative source of ~ in aerobic organisms. ATP is generated O O O C C H H in the mitochondrial matrix as O2 is reduced to H2O by ATP 4– H H electrons passing down the respiratory chain (see Chap- OH OH ter 13). 3– Od – Hydrolysis of ATP4– to 2. Glycolysis. A net formation of two ~ results from Od – P Od – the formation of lactate from one molecule of glucose, ADP3– relieves charge Od – repulsion generated in two reactions catalyzed by phosphoglyc- The phosphate released +H + erate kinase and pyruvate kinase, respectively (see Fig- is stabilised by the formation of a ure 17–2). resonance hybrid in which the ADP3– three negative charges are shared ADENOSINE 3. The citric acid cycle. One ~ is generated directly in the between the four O atoms – – cycle at the succinate thiokinase step (see Figure 16–3). O O – O P O P O CH2 O Phosphagens act as storage forms of group transfer poten- tial and include creatine phosphate, which occurs in vertebrate O O C C H H skeletal muscle, heart, spermatozoa, and brain, and arginine H H phosphate, which occurs in invertebrate muscle. When ATP is rapidly being utilized as a source of energy for muscular contrac- OH OH tion, phosphagens permit its concentrations to be maintained, FIGURE 11–5 The free-energy change on hydrolysis of ATP but when the ATP/ADP ratio is high, their concentration can to ADP. increase to act as an energy store (Figure 11–7).  CHAPTER 11 Bioenergetics: The Role of ATP 109 H Creatine Adenylyl kinase is important for the maintenance of energy N kinase H 2N P homeostasis in cells because it allows: C NH C NH H3C N H3C N 1. The group transfer potential in ADP to be used in the syn- ADP ATP thesis of ATP. CH2 CH2 – (∆G 0′ = –12.6 kJ/mol) 2. The AMP formed as a consequence of activating reactions COO COO– involving ATP to rephosphorylated to ADP. Creatine phosphate Creatine 3. AMP to increase in concentration when ATP becomes FIGURE 11–7 Transfer of high-energy phosphate between depleted so that it is able to act as a metabolic (allosteric) ATP and creatine. signal to increase the rate of catabolic reactions, which in turn lead to the generation of more ATP (see Chapter 14). When ATP acts as a phosphate donor to form com- pounds of lower free energy of hydrolysis (Table 11–1), When ATP Forms AMP, Inorganic the phosphate group is invariably converted to one of low Pyrophosphate (PPi) Is Produced energy. For example, the phosphorylation of glycerol to form ATP can also be hydrolyzed directly to AMP, with the release glycerol-3-phosphate: of PPi (Table 11–1). This occurs, for example, in the activation of long-chain fatty acids (see Chapter 22). GLYCEROL KINASE ACYL-CoA Glycerol + Adenosine SYNTHETASE P P P ATP + CoA SH + R COOH AMP + PPi + R CO — SCoA Glycerol P + Adenosine P P This reaction is accompanied by loss of free energy as heat, which ensures that the activation reaction will go to the right, ATP Allows the Coupling of and is further aided by the hydrolytic splitting of PPi, catalyzed Thermodynamically Unfavorable by inorganic pyrophosphatase, a reaction that itself has a large ΔG0′ of –19.2 kJ/mol. Note that activations via the pyro- Reactions to Favorable Ones phosphate pathway result in the loss of two ~ rather than Endergonic reactions cannot proceed without an input of one, as occurs when ADP and Pi are formed. free energy. For example, the phosphorylation of glucose to glucose-6-phosphate, the first reaction of glycolysis (see INORGANIC Figure 17–2): PYROPHOSPHATASE PPi + H2O 2Pi Glucose + Pi → Glucose-6-phosphate + H2O A combination of the above reactions makes it possible for (ΔG0′) = +13.8 kJ/mol (1) phosphate to be recycled and the adenine nucleotides to inter- is highly endergonic and cannot proceed under physiologic change (Figure 11–8). conditions. Thus, in order to take place, the reaction must be coupled with another—more exergonic—reaction such as the Inorganic hydrolysis of the terminal phosphate of ATP. pyrophosphatase 2Pi ATP → ADP + Pi (ΔG = –30.5 kJ/mol) 0′ (2) Pi PPi When (1) and (2) are coupled in a reaction catalyzed by hexokinase, phosphorylation of glucose readily proceeds in a Acyl-CoA highly exergonic reaction that under physiologic conditions synthetase, etc is irreversible. Many “activation” reactions follow this pattern. ATP Adenylyl Kinase (Myokinase) Interconverts Adenine Nucleotides X2 This enzyme is present in most cells. It catalyzes the following ADP AMP reaction: Adenylyl kinase ADENYLYL KINASE FIGURE 11–8 Phosphate cycles and interchange of adenine ATP + AMP 2ADP nucleotides. 110 SECTION III Bioenergetics Other Nucleoside Triphosphates SUMMARY Participate in Group Transfer Potential Biologic systems use chemical energy to power living processes. By means of the nucleoside diphosphate (NDP) kinases, Exergonic reactions take place spontaneously with loss of free UTP, GTP, and CTP can be synthesized from their diphos- energy (ΔG is negative). Endergonic reactions require the gain phates, for example, UDP reacts with ATP to form UTP. of free energy (ΔG is positive) and occur only when coupled to exergonic reactions. NUCLEOSIDE ATP acts as the “energy currency” of the cell, transferring free DIPHOSPHATE KINASE energy derived from substances of higher energy potential to ATP + UDP ADP + UTP those of lower energy potential. (uridine triphosphate) All of these triphosphates take part in phosphorylations in the REFERENCES cell. Similarly, specific nucleoside monophosphate (NMP) Haynie D: Biological Thermodynamics. Cambridge University Press, kinases catalyze the formation of NDP from the correspond- 2008. ing monophosphates. Nicholls DG, Ferguson S: Bioenergetics, 4th ed. Academic Press, Thus, Adenylyl kinase is a specialized NMP kinase. 2013.  12 C H A P T E R Biologic Oxidation Kathleen M. Botham, PhD, DSc & Peter A. Mayes, PhD, DSc O B J EC T IVES Explain the meaning of redox potential and how it can be used to predict the direction of flow of electrons in biologic systems. After studying this chapter, Identify the four classes of enzymes (oxidoreductases) involved in oxidation you should be able to: and reduction reactions. Describe the action of oxidases and provide examples of where they play an important role in metabolism. Indicate the two main functions of dehydrogenases and explain the importance of nicotinamide adenine dinucleotide (NAD)- and riboflavin-linked dehydrogenases in metabolic pathways such as glycolysis, the citric acid cycle, and the respiratory chain. Identify the two types of enzymes classified as hydroperoxidases; indicate the reactions they catalyze and explain why they are important. Give the two steps of reactions catalyzed by oxygenases and identify the two subgroups of this class of enzymes. Appreciate the role of cytochrome P450 in drug detoxification and steroid synthesis. Describe the reaction catalyzed by superoxide dismutase and explain how it protects tissues from oxygen toxicity. BIOMEDICAL IMPORTANCE class, known as the cytochrome P450 system. Administration of oxygen can be lifesaving in the treatment of patients with Chemically, oxidation is defined as the removal of electrons respiratory or circulatory failure. and reduction as the gain of electrons. Thus, oxidation of a molecule (the electron donor) is always accompanied by reduction of a second molecule (the electron acceptor). This FREE ENERGY CHANGES principle of oxidation–reduction applies equally to biochemi- cal systems and is an important concept underlying under- CAN BE EXPRESSED IN TERMS standing of the nature of biologic oxidation. Note that many OF REDOX POTENTIAL biologic oxidations can take place without the participation In reactions involving oxidation and reduction, the free of molecular oxygen, for example, dehydrogenations. The life energy change is proportionate to the tendency of reactants of higher animals is absolutely dependent on a supply of oxy- to donate or accept electrons. Thus, in addition to express- gen for respiration, the process by which cells derive energy ing free energy change in terms of ΔG0′ (see Chapter 11), it is in the form of ATP from the controlled reaction of hydrogen possible, in an analogous manner, to express it numerically as with oxygen to form water. In addition, molecular oxygen is an oxidation–reduction or redox potential (E′0). Chemi- incorporated into a variety of substrates by enzymes desig- cally, the redox potential of a system (E0) is usually com- nated as oxygenases; many drugs, pollutants, and chemical pared with the potential of the hydrogen electrode (0.0 V at carcinogens (xenobiotics) are metabolized by enzymes of this pH 0.0). However, for biologic systems, the redox potential 111 112 SECTION III Bioenergetics TABLE 12–1 Some Redox Potentials of Special Interest AH2 1 /2O2 AH2 O2 in Mammalian Oxidation Systems (Red) Oxidase Oxidase System E′0 Volts A H2O A H2O2 H+/H2 –0.42 (Ox) NAD+/NADH –0.32 A B Lipoate; ox/red –0.29 FIGURE 12–1 Oxidation of a metabolite catalyzed by an oxi- Acetoacetate/3-hydroxybutyrate –0.27 dase (A) forming H2O and (B) forming H2O2. Pyruvate/lactate –0.19 Oxaloacetate/malate –0.17 heme, heme a, in a single protein and so is also termed cyto- chrome aa3. It contains two molecules of heme, each having Fumarate/succinate +0.03 one Fe atom that oscillates between Fe3+ and Fe2+ during oxi- Cytochrome b; Fe3+/Fe2+ +0.08 dation and reduction. Furthermore, two atoms of copper are Ubiquinone; ox/red +0.10 present, one associated with each heme unit. Cytochrome c1; Fe3+/Fe2+ +0.22 Other Oxidases Are Flavoproteins Cytochrome a; Fe3+/Fe2+ +0.29 Flavoprotein enzymes contain flavin mononucleotide (FMN) Oxygen/water +0.82 or flavin adenine dinucleotide (FAD) as prosthetic groups. FMN and FAD are formed in the body from the vitamin ribo- flavin (see Chapter 44). FMN and FAD are usually tightly— (E′0) is normally expressed at pH 7.0, at which pH the elec- but not covalently—bound to their respective apoenzyme trode potential of the hydrogen electrode is –0.42 V. The proteins. Metalloflavoproteins contain one or more metals as redox potentials of some redox systems of special interest in essential cofactors. Examples of flavoprotein oxidases include mammalian biochemistry are shown in Table 12–1. The rela- l-amino acid oxidase, an enzyme found in kidney with gen- tive positions of redox systems in the table allow prediction eral specificity for the oxidative deamination of the naturally of the direction of flow of electrons from one redox couple occurring l-amino acids; xanthine oxidase, which contains to another. molybdenum and plays an important role in the conversion Enzymes involved in oxidation and reduction are called of purine bases to uric acid (see Chapter 33), and is of par- oxidoreductases and are classified into four groups: oxidases, ticular significance in uricotelic animals (see Chapter 28); dehydrogenases, hydroperoxidases, and oxygenases. and aldehyde dehydrogenase, an FAD-linked enzyme pres- ent in mammalian livers, which contains molybdenum and nonheme iron and acts on aldehydes and N-heterocyclic sub- OXIDASES USE OXYGEN AS strates. The mechanisms of oxidation and reduction of these A HYDROGEN ACCEPTOR enzymes are complex. Evidence suggests a two-step reaction as shown in Figure 12–2. Oxidases catalyze the removal of hydrogen from a substrate using oxygen as a hydrogen acceptor.∗ They form water or hydrogen peroxide as a reaction product (Figure 12–1). DEHYDROGENASES PERFORM Cytochrome Oxidase Is a Hemoprotein TWO MAIN FUNCTIONS There are a large number of enzymes in the dehydrogenase Cytochrome oxidase is a hemoprotein widely distributed in class. Their two main functions are as follows: many tissues, having the typical heme prosthetic group pres- ent in myoglobin, hemoglobin, and other cytochromes (see 1. Transfer of hydrogen from one substrate to another in a Chapter 6). It is the terminal component of the chain of respi- coupled oxidation–reduction reaction (Figure 12–3). ratory carriers found in mitochondria (see Chapter 13) and These dehydrogenases often utilize common coenzymes transfers electrons resulting from the oxidation of substrate or hydrogen carriers, for example, nicotinamide adenine molecules by dehydrogenases to their final acceptor, oxygen. dinucleotide (NAD+). This type of reaction in which one The action of the enzyme is blocked by carbon monoxide, substrate is oxidized/reduced at the expense of another is cyanide, and hydrogen sulfide, and this causes poisoning freely reversible, enabling reducing equivalents to be trans- by preventing cellular respiration. The cytochrome oxidase ferred within the cell and oxidative processes to occur in enzyme complex comprises heme a3 combined with another the absence of oxygen, such as during the anaerobic phase of glycolysis (see Figure 17–2). ∗ The term “oxidase” is sometimes used collectively to denote all 2. Transfer of electrons in the respiratory chain of electron enzymes that catalyze reactions involving molecular oxygen. transport from substrate to oxygen (see Figure 13–3).  CHAPTER 12 Biologic Oxidation 113 R R R H H H3C N N O H3C N N O H3C N N O Oxidized Substrate + + NH NH NH substrate H3C N H3 C N H3C N O O H O H H Oxidized flavin Semiquinone (H+ + e–) (H+ + e–) Reduced flavin (FAD) intermediate (FADH2) FIGURE 12–2 Oxidoreduction of isoalloxazine ring in flavin nucleotides via a semiquinone intermediate. In oxidation reactions, the flavin (eg, FAD) accepts two electrons and two H+ in two steps, forming the semiquinone intermediate followed by the reduced flavin (eg, FADH2) and the substrate is oxidized. In the reverse (reduction) reaction, the reduced flavin gives up two electrons and two H+ so that it becomes oxidized (eg, to FAD) and the substrate is reduced. Many Dehydrogenases Depend on leads to the formation of ATP (see Chapter 13). NADP-linked dehydrogenases are found characteristically in biosynthetic Nicotinamide Coenzymes pathways where reductive reactions are required, as in the extra- These dehydrogenases use NAD+ or nicotinamide adenine mitochondrial pathway of fatty acid synthesis (see Chapter 23) dinucleotide phosphate (NADP+)—or both—which are and steroid synthesis (see Chapter 26)—and also in the pentose formed in the body from the vitamin niacin (see Chapter 44). phosphate pathway (see Chapter 20). The structure of NAD+ is shown in Figure 12–4. NADP+ has a phosphate group esterified to the 2′ hydroxyl of its adenosine H O moiety, but otherwise is identical to NAD+. The oxidized forms H H O of both nucleotides have a positive charge on the nitrogen atom NH2 NH2 O– of the nicotinamide moiety as indicated in Figure 12–4. The + coenzymes are reduced by the specific substrate of the dehy- O P O N N drogenase and reoxidized by a suitable electron acceptor. They O H+ are able to freely and reversibly dissociate from their respective R apoenzymes. Generally, NAD-linked dehydrogenases catalyze oxido- O OH OH NH2 reduction reactions of the type: N OH O N C + NAD+ C + NADH + H+ O P O N N O– O H When a substrate is oxidized, it loses two hydrogen atoms and two electrons. One H+ and both electrons are accepted by NAD+ OH OH to form NADH and the other H+ is released (Figure 12–4). Many such reactions occur in the oxidative pathways of metab- olism, particularly in glycolysis (see Chapter 17) and the citric Oxidized substrate/product acid cycle (see Chapter 16). NADH is generated in these path- OH O ways via the oxidation of fuel molecules, and NAD+ is regener- ated by the oxidation of NADH as it transfers the electrons to NAD+ + C C + NADH + H+ O2 via the respiratory chain in mitochondria, a process which H Reduced substrate/product AH2 Carrier BH2 (Red) (Ox) (Red) FIGURE 12–4 Oxidation and reduction of nicotinamide coenzymes. Nicotinamide coenzymes consist of a nicotinamide ring linked to an adenosine via a ribose and a phosphate group, forming A Carrier–H2 B a dinucleotide. NAD+/NADH are shown, but NADP+/NADPH are iden- (Ox) (Red) (Ox) tical except that they have a phosphate group esterified to the 2′ OH Dehydrogenase Dehydrogenase of the adenosine. An oxidation reaction involves the transfer of two specific for A specific for B electrons and one H+ from the substrate to the nicotinamide ring of NAD+ forming NADH and the oxidized product. The remaining hydro- FIGURE 12–3 Oxidation of a metabolite catalyzed by cou- gen of the hydrogen pair removed from the substrate remains free as pled dehydrogenases. a hydrogen ion. NADH is oxidized to NAD+ by the reverse reaction. 114 SECTION III Bioenergetics Other Dehydrogenases Depend on metabolism, but can be damaging if they accumulate. They are believed to contribute to the causation of diseases such as can- Riboflavin cer and atherosclerosis, as well as the aging process in general The flavin groups such as FMN and FAD are associated with (see Chapters 21, 44, 54). dehydrogenases as well as with oxidases as described above. FAD is the electron acceptor in reactions of the type: Peroxidases Reduce Peroxides Using H Various Electron Acceptors C C + FAD C C + FADH2 Peroxidases are found in milk and in leukocytes, platelets, and H H H other tissues involved in eicosanoid metabolism (see Chapter 23). Their prosthetic group is protoheme. In the reaction catalyzed by FAD accepts two electrons and two H + in the reaction peroxidase, hydrogen peroxide is reduced at the expense of sev- (Figure 12–2), forming FADH2. Flavin groups are generally eral substances that act as electron acceptors, such as ascorbate more tightly bound to their apoenzymes than are the nico- (vitamin C), quinones, and cytochrome c. The reaction catalyzed tinamide coenzymes. Most of the riboflavin-linked dehydro- by peroxidase is complex, but the overall reaction is as follows: genases are concerned with electron transport in (or to) the respiratory chain (see Chapter 13). NADH dehydrogenase PEROXIDASE acts as a carrier of electrons between NADH and the com- H2O2 + AH2 2H2O + A ponents of higher redox potential (see Figure 13–3). Other dehydrogenases such as succinate dehydrogenase, acyl-CoA In erythrocytes and other tissues, the enzyme glutathione per- dehydrogenase, and mitochondrial glycerol-3-phosphate oxidase, containing selenium as a prosthetic group, catalyzes dehydrogenase transfer reducing equivalents directly from the destruction of H2O2 and lipid hydroperoxides through the the substrate to the respiratory chain (see Figure 13–5). conversion of reduced glutathione to its oxidized form, pro- Another role of the flavin-dependent dehydrogenases is in tecting membrane lipids and hemoglobin against oxidation by the dehydrogenation (by dihydrolipoyl dehydrogenase) of peroxides (see Chapter 21). reduced lipoate, an intermediate in the oxidative decarboxyl- ation of pyruvate and α-ketoglutarate (see Figures 13–5 and Catalase Uses Hydrogen Peroxide as 17–5). The electron-transferring flavoprotein (ETF) is an Electron Donor & Electron Acceptor intermediary carrier of electrons between acyl-CoA dehydro- Catalase is a hemoprotein containing four heme groups. It genase and the respiratory chain (see Figure 13–5). can act as a peroxidase, catalyzing reactions of the type shown above, but it is also able to catalyze the breakdown of H2O2 Cytochromes May Also Be formed by the action of oxygenases to water and oxygen: Regarded as Dehydrogenases CATALASE The cytochromes are iron-containing hemoproteins in which 2H2O2 2H2O + O2 the iron atom oscillates between Fe3+ and Fe2+ during oxida- tion and reduction. Except for cytochrome oxidase (previously This reaction uses one molecule of H2O2 as a substrate electron described), they are classified as dehydrogenases. In the respi- donor and another molecule of H2O2 as an oxidant or electron ratory chain, they are involved as carriers of electrons from fla- acceptor. It is one of the fastest enzyme reactions known, destroy- voproteins on the one hand to cytochrome oxidase on the other ing millions of potentially damaging H2O2 molecules per second. (see Figure 13–5). Several identifiable cytochromes occur in the Under most conditions in vivo, the peroxidase activity of catalase respiratory chain, that is, cytochromes b, c1, c, and cytochrome seems to be favored. Catalase is found in blood, bone marrow, oxidase (aa3). Cytochromes are also found in other locations, mucous membranes, kidney, and liver. Peroxisomes are found for example, the endoplasmic reticulum (cytochromes P450 in many tissues, including liver. They are rich in oxidases and and b5), and in plant cells, bacteria, and yeasts. in catalase. Thus, the enzymes that produce H2O2 are grouped with the enzyme that breaks it down. However, mitochondrial and microsomal electron transport systems as well as xanthine HYDROPEROXIDASES USE oxidase must be considered as additional sources of H2O2. HYDROGEN PEROXIDE OR AN ORGANIC PEROXIDE OXYGENASES CATALYZE AS SUBSTRATE THE DIRECT TRANSFER Two types of enzymes found both in animals and plants fall & INCORPORATION OF OXYGEN into the hydroperoxidase category: peroxidases and catalase. Hydroperoxidases play an important role in protecting INTO A SUBSTRATE MOLECULE the body against the harmful effects of reactive oxygen spe- Oxygenases are concerned with the synthesis or degrada- cies (ROS). ROS are highly reactive oxygen-containing mol- tion of many different types of metabolites. They catalyze the ecules such as peroxides, which are formed during normal incorporation of oxygen into a substrate molecule in two steps:  CHAPTER 12 Biologic Oxidation 115 (1) oxygen is bound to the enzyme at the active site and (2) the but are also found in the mitochondria in some tissues. The bound oxygen is reduced or transferred to the substrate. Oxy- cytochromes participate in an electron transport chain in genases may be divided into two subgroups, dioxygenases and which both NADH and NADPH may donate reducing equiv- monooxygenases. alents. Electrons are passed to cytochrome P450 in two types of reaction involving FAD or FMN. Class I systems consist of an FAD-containing reductase enzyme, an iron sulfur (Fe2S2) Dioxygenases Incorporate Both Atoms protein, and the P450 heme protein, while class II systems con- of Molecular Oxygen into the Substrate tain cytochrome P450 reductase, which passes electrons from The basic reaction catalyzed by dioxygenases is shown below: FADH2 to FMN (Figure 12–5). Class I and II systems are well characterized, but in recent years, other cytochromes P450, A + O2 → AO2 which do not fit into either category, have been identified. In Examples include the liver enzymes, homogentisate dioxy- the final step, oxygen accepts the electrons from cytochrome genase (oxidase) and 3-hydroxyanthranilate dioxygenase P450 and is reduced, with one atom being incorporated into (oxidase), which contain iron; and l-tryptophan dioxygenase H2O and the other into the substrate, usually resulting in its (tryptophan pyrrolase) (see Chapter 29), which utilizes heme. hydroxylation. This series of enzymatic reactions, known as the hydroxylase cycle, is illustrated in Figure 12–6. In the endo- plasmic reticulum of the liver, cytochromes P450 are found Monooxygenases (Mixed-Function together with another heme-containing protein, cytochrome b5 Oxidases, Hydroxylases) Incorporate (Figure 12–5) and together they have a major role in drug Only One Atom of Molecular Oxygen metabolism and detoxification. Cytochrome b5 also has an Into the Substrate important role as a fatty acid desaturase. Together, cytochromes P450 and b5 are responsible for about 75% of the modification The other oxygen atom is reduced to water, an additional elec- and degradation of drugs, which occurs in the body. The rate of tron donor or cosubstrate (Z) being necessary for this purpose: detoxification of many medicinal drugs by cytochromes P450 A — H + O2 +ZH2 → A — OH + H2O + Z determines the duration of their action. Benzpyrene, aminopy- rine, aniline, morphine, and benzphetamine are hydroxylated, Cytochromes P450 Are Monooxygenases increasing their solubility and aiding their excretion. Many Important in Steroid Metabolism & for drugs such as phenobarbital have the ability to induce the syn- thesis of cytochromes P450. the Detoxification of Many Drugs Mitochondrial cytochrome P450 systems are found in Cytochromes P450 are an important superfamily of heme- steroidogenic tissues such as adrenal cortex, testis, ovary, containing monooxygenases, and more than 50 such enzymes and placenta and are concerned with the biosynthesis of ste- have been found in the human genome. They are located roid hormones from cholesterol (hydroxylation at C22 and mainly in the endoplasmic reticulum in the liver and intestine, C20 in side-chain cleavage and at the 11β and 18 positions). Class I P450 REDUCTASE Fe2S2 P450 NAD(P)H FAD FADH2 Fe3+ Fe2+ O2+RH H2O+ROH Hydroxylation Class II P450 P450 REDUCTASE P450 NAD(P)H Hydroxylation FAD FMN FMNH2 O2+RH H2O+ROH Cytochrome b5 O2+Oleoyl CoA b5 REDUCTASE NADH b5 FAD FADH2 Stearoyl CoA + H2O Stearoyl CoA desaturase P450 REDUCTASE P450 Hydroxylation FAD FMN FMNH2 O2+RH H2O+ROH FIGURE 12–5 Cytochromes P450 and b5 in the endoplasmic reticulum. Most cytochromes P450 are class I or class II. In addition to cytochrome P450, class I systems contain a small FAD-containing reductase and an iron sulfur protein, and class II contains cytochrome P450 reductase, which incorporates FAD and FMN. Cytochromes P450 catalyze many steroid hydroxylation reactions and drug detoxification steps. Cytochrome b5 acts in conjunction with the FAD-containing cytochrome b5 reductase in the fatty acyl-CoA desaturase (eg, stearoyl-CoA desaturase) reaction and also works together with cytochromes P450 in drug detoxification. It is able to accept electrons from cytochrome P450 reductase via cytochrome b5 reductase and donate them to cytochrome P450. 116 SECTION III Bioenergetics Substrate A-H P450-A-H Fe3+ e– P450 P450-A-H NADPH-Cyt P450 reductase Fe3+ Fe2+ NADP+ FADH2 2Fe2S23+ O2 e– – NADPH + H+ FAD 2Fe2S22+ CO 2H+ P450-A-H Fe2+ O2 H2O P450-A-H Fe2+ O2– A-OH FIGURE 12–6 Cytochrome P450 hydroxylase cycle. The system shown is typical of steroid hydroxylases of the adrenal cortex. Liver microsomal cytochrome P450 hydroxylase does not require the iron-sulfur protein Fe2S2. Carbon monoxide (CO) inhibits the indicated step. In addition, renal systems catalyzing 1α- and 24-hydroxylations In this reaction, superoxide acts as both oxidant and of 25-hydroxycholecalciferol in vitamin D metabolism—and reductant. Thus, superoxide dismutase protects aerobic cholesterol 7α-hydroxylase and sterol 27-hydroxylase involved organisms against the potential deleterious effects of super- in bile acid biosynthesis from cholesterol in the liver (see oxide. The enzyme occurs in all major aerobic tissues in the Chapters 26, 41)—are P450 enzymes. mitochondria and the cytosol. Although exposure of ani- mals to an atmosphere of 100% oxygen causes an adaptive increase in SOD, particularly in the lungs, prolonged exposure SUPEROXIDE DISMUTASE leads to lung damage and death. Antioxidants, for example, PROTECTS AEROBIC ORGANISMS α-tocopherol (vitamin E), act as scavengers of free radicals and reduce the toxicity of oxygen (see Chapter 44). AGAINST OXYGEN TOXICITY Transfer of a single electron to O2 generates the potentially damaging superoxide anion-free radical (O2−), which gives SUMMARY rise to free-radical chain reactions (see Chapter 21), amplifying In biologic systems, as in chemical systems, oxidation (loss of its destructive effects. The ease with which superoxide can be electrons) is always accompanied by reduction of an electron formed from oxygen in tissues and the occurrence of superoxide acceptor. dismutase (SOD), the enzyme responsible for its removal in all Oxidoreductases have a variety of functions in metabolism; aerobic organisms (although not in obligate anaerobes), indicate oxidases and dehydrogenases play major roles in respiration; that the potential toxicity of oxygen is due to its conversion to hydroperoxidases protect the body against damage by free superoxide. radicals; and oxygenases mediate the hydroxylation of drugs and steroids. Superoxide is formed when reduced flavins—present, for example, in xanthine oxidase—are reoxidized univalently by Tissues are protected from oxygen toxicity caused by the molecular oxygen: superoxide free radical by the speci%c enzyme superoxide dismutase. Enz − Flavin − H2 + O2 → Enz − Flavin − H + O2−. + H+ Superoxide can reduce oxidized cytochrome c REFERENCES − 3+ 2+ O2 + Cyt c (Fe ) → O2 + Cyt c(Fe )  Nelson DL, Cox MM: Lehninger Principles of Biochemistry, 6th ed. Macmillan Higher Education, 2013. or be removed by superoxide dismutase, which catalyzes the Nicholls DG, Ferguson SJ: Bioenergetics, 4th ed. Academic Press, conversion of to oxygen and hydrogen peroxide. 2013.  13 C H A P T E R The Respiratory Chain & Oxidative Phosphorylation Kathleen M. Botham, PhD, DSc & Peter A. Mayes, PhD, DSc O B J EC T IVES Describe the double membrane structure of mitochondria and indicate the location of various enzymes. After studying this chapter, Appreciate that energy from the oxidation of fuel substrates (fats, you should be able to: carbohydrates, amino acids) is almost all generated in mitochondria via a process termed electron transport in which electrons pass through a series of complexes (the respiratory chain) until they are finally reacted with oxygen to form water. Describe the four protein complexes involved in the transfer of electrons through the respiratory chain and explain the roles of flavoproteins, iron-sulfur proteins, and coenzyme Q. Explain how coenzyme Q accepts electrons from NADH via Complex I and from FADH2 via Complex II. Indicate how electrons are passed from reduced coenzyme Q to cytochrome c via Complex III in the Q cycle. Explain the process by which reduced cytochrome c is oxidized and oxygen is reduced to water via Complex IV. Describe how electron transport generates a proton gradient across the inner mitochondrial membrane, leading to the buildup of a proton motive force that generates ATP by the process of oxidative phosphorylation. Describe the structure of the ATP synthase enzyme and explain how it works as a rotary motor to produce ATP from ADP and Pi. Explain that oxidation of reducing equivalents via the respiratory chain and oxidative phosphorylation are tightly coupled in most circumstances, so that one cannot proceed unless the other is functioning. Indicate examples of common poisons that block respiration or oxidative phosphorylation and identify their site of action. Explain, with examples, how uncouplers may act as poisons by dissociating oxidation via the respiratory chain from oxidative phosphorylation, but may also have a physiologic role in generating body heat. Explain the role of exchange transporters present in the inner mitochondrial membrane in allowing ions and metabolites to pass through while preserving electrochemical and osmotic equilibrium. 117 118 SECTION III Bioenergetics BIOMEDICAL IMPORTANCE inner membrane together with the enzymes of the respiratory chain, ATP synthase, and various membrane transporters. Aerobic organisms are able to capture a far greater propor- tion of the available free energy of respiratory substrates than anaerobic organisms. Most of this takes place inside THE RESPIRATORY CHAIN mitochondria, which have been termed the “powerhouses” of the cell. Respiration is coupled to the generation of the OXIDIZES REDUCING high-energy intermediate, ATP (see Chapter 11), by oxida- EQUIVALENTS & ACTS tive phosphorylation. A number of drugs (eg, amobarbital) AS A PROTON PUMP and poisons (eg, cyanide, carbon monoxide) inhibit oxida- tive phosphorylation, usually with fatal consequences. Sev- Most of the energy liberated during the oxidation of carbo- eral inherited defects of mitochondria involving components hydrate, fatty acids, and amino acids is made available within of the respiratory chain and oxidative phosphorylation have mitochondria as reducing equivalents (—H or electrons) been reported. Patients present with myopathy and encepha- (Figure 13–2). The enzymes of the citric acid cycle and lopathy and often have lactic acidosis. β-oxidation (see Chapters 22 and 16), the respiratory chain complexes, and the machinery for oxidative phosphorylation are all found in mitochondria. The respiratory chain collects SPECIFIC ENZYMES and transports reducing equivalents, directing them to their ARE ASSOCIATED WITH final reaction with oxygen to form water, and oxidative phos- phorylation is the process by which the liberated free energy is COMPARTMENTS SEPARATED trapped as high-energy phosphate. BY THE MITOCHONDRIAL MEMBRANES Components of the Respiratory Chain The mitochondrial matrix (the internal compartment) is Are Contained in Four Large Protein enclosed by a double membrane. The outer membrane is Complexes Embedded in the Inner permeable to most metabolites and the inner membrane is selectively permeable (Figure 13–1). The outer mem- Mitochondrial Membrane brane is characterized by the presence of various enzymes, Electrons flow through the respiratory chain through a redox span including acyl-CoA synthetase (see Chapter 22) and glycerol of 1.1 V from NAD+/NADH to O2/2H2O (see Table 12–1), passing phosphate acyltransferase (see Chapter 24). Other enzymes, through three large protein complexes: NADH-Q oxidoreduc- including adenylyl kinase (see Chapter 11) and creatine tase (Complex I), where electrons are transferred from NADH kinase (see Chapter 51) are found in the intermembrane to coenzyme Q (Q) (also called ubiquinone); Q-cytochrome c space. The phospholipid cardiolipin is concentrated in the oxidoreductase (Complex III), which passes the electrons on to cytochrome c; and cytochrome c oxidase (Complex IV), which completes the chain, passing the electrons to O2 and causing it to be reduced to H2O (Figure 13–3). Some substrates with more positive redox potentials than NAD+/NADH (eg, succinate) pass electrons to Q via a fourth complex, succinate-Q reduc- Matrix Enzymes of inner membrane include: tase (Complex II), rather than Complex I. The four complexes Electron carriers (complexes I–IV) ATP synthase are embedded in the inner mitochondrial membrane, but Q and Membrane transporters cytochrome c are mobile. Q diffuses rapidly within the mem- brane, while cytochrome c is a soluble protein. Enzymes of the mitochondrial matrix include: Citric acid cycle enzymes Inter- β-Oxidation enzymes Flavoproteins & Iron-Sulfur Pyruvate dehydrogenase membrane space Cristae Proteins (Fe-S) Are Components of the Respiratory Chain Complexes Flavoproteins (see Chapter 12) are important components Inner of Complexes I and II. The oxidized flavin nucleotide (flavin membrane mononucleotide [FMN] or flavin adenine dinucleotide [FAD]) can be reduced in reactions involving the transfer of two elec- Outer membrane Enzymes in the outer membrane trons (to form FMNH2 or FADH2), but they can also accept include: Acyl CoA synthetase one electron to form the semiquinone (see Figure 12–2). Iron- Glycerolphosphate acyl transferase sulfur proteins (nonheme iron proteins, Fe-S) are found in Complexes I, II, and III. These may contain one, two, or four Fe FIGURE 13–1 Structure of the mitochondrial membranes. atoms linked to inorganic sulfur atoms and/or via cysteine-SH Note that the inner membrane contains many folds or cristae. groups to the protein (Figure 13–4). The Fe-S take part in single  CHAPTER 13 The Respiratory Chain & Oxidative Phosphorylation 119 Food ATP Digestion and absorption Fat Fatty acids + β-Oxidation Glycerol O2 Citric Carbohydrate Glucose, etc Acetyl – CoA acid 2H H2O cycle Respiratory chain Protein Amino acids Mitochondrion ADP Extramitochondrial sources of reducing equivalents FIGURE 13–2 Role of the respiratory chain of mitochondria in the conversion of food energy to ATP. Oxidation of the major foodstuffs leads to the generation of reducing equivalents (2H) that are collected by the respiratory chain for oxidation and coupled generation of ATP. Succinate Fumarate Complex II succinate-Q reductase NADH + H+ 1/ O 2 2 + 2H+ Q Cyt c NAD H2O Complex I Complex III Complex IV NADH-Q Q-cyt c Cyt c oxidoreductase oxidoreductase oxidase FIGURE 13–3 Overview of electron flow through the respiratory chain. (cyt, cytochrome; Q, coenzyme Q or ubiquinone.) Pr Pr Cys Cys S S Fe Pr S S Cys Cys Cys S Pr Pr S Fe A Pr Cys S Fe S Pr Pr Cys Cys Fe S S S S S S Fe Fe Fe Cys S S S S Cys Pr Cys Cys Pr C Pr Pr B FIGURE 13–4 Iron-sulfur proteins (Fe-S). (A) The simplest Fe-S with one Fe bound by four cysteines. (B) 2Fe-2S center. (C) 4Fe-4S center. (Cys, cysteine; Pr, apoprotein; , inorganic sulfur.) 120 SECTION III Bioenergetics electron transfer reactions in which one Fe atom undergoes oxi- Fe atoms is linked to two histidine residues rather than two doreduction between Fe2+ and Fe3+. cysteine residues) (Figure 13–5) and is known as the Q cycle (Figure 13–6). Q may exist in three forms: the oxidized qui- Q Accepts Electrons via Complexes I & II none, the reduced quinol, or the semiquinone (Figure 13–6). NADH-Q oxidoreductase or Complex I is a large L-shaped The semiquinone is formed transiently during the cycle, one multisubunit protein that catalyzes electron transfer from turn of which results in the oxidation of 2QH2 to Q, releas- NADH to Q, and during the process four H+ are transferred ing 4H+ into the intermembrane space, and the reduction of across the membrane into the intermembrane space: one Q to QH2, causing 2H+ to be taken up from the matrix (Figure 13–6). Note that while Q carries two electrons, the NADH + Q + 5H+matrix → NAD + QH2 + 4H+intermembrance space cytochromes carry only one, thus the oxidation of one QH2 is coupled to the reduction of two molecules of cytochrome c Electrons are transferred from NADH to FMN initially, then via the Q cycle. to a series of Fe-S centers, and finally to Q (Figure 13–5). In Complex II (succinate-Q reductase), FADH2 is formed during the conversion of succinate to fumarate in the citric acid cycle Molecular Oxygen Is Reduced (see Figure 16–3) and electrons are then passed via several to Water via Complex IV Fe-S centers to Q (Figure 13–5). Glycerol-3-phosphate (gener- Reduced cytochrome c is oxidized by Complex IV (cyto- ated in the breakdown of triacylglycerols or from glycolysis, chrome c oxidase), with the concomitant reduction of O2 to see Figure 17–2) and acyl-CoA also pass electrons to Q via two molecules of water: different pathways involving flavoproteins (Figure 13–5). 4Cyt creduced + O2 + 8H+matrix → The Q Cycle Couples Electron Transfer 4Cyt coxidized + 2H2O + 4H+intermembrane space to Proton Transport in Complex III Electrons are passed from QH2 to cytochrome c via Complex Four electrons are transferred from cytochrome c to O2 via III (Q-cytochrome c oxidoreductase): two heme groups, a and a3, and Cu (Figure 13–5). Electrons are passed initially to a Cu center (CuA), which contains 2Cu QH2 + 2Cyt coxidized + 2H+matrix → atoms linked to two protein cysteine-SH groups (resembling Q + 2Cyt creduced + 4H+intermembrane space an Fe-S), then in sequence to heme a, heme a3, a second Cu center, CuB, which is linked to heme a3, and finally to O2. The process is believed to involve cytochromes c1, bL, and Eight H+ are removed from the matrix, of which four are used bH and a Rieske Fe-S (an unusual Fe-S in which one of the to form two water molecules and four are pumped into the Glycerol-3-phosphate + + + + 4H 4H 2H 4H Intermembrane FAD space Cyt c Cyt c Complex I Complex II Inner Fe-S Q Cyt b Cyt b Q Fe-S mitochondrial Heme a + a3 membrane FMN Cyt c1 CuACuB Cyt c1 FAD Fe-S Complex III Complex IV Complex III Complex III Mitochondrial matrix NADH + H+ NAD ETF Fumarate Succinate 1/ O +