LEHNINGER PRINCIPLES OF BIOCHEMISTRY - VOL. 2. BIOENERGETICS AND METABOLISM - 2014

PART II. BIOENERGETICS AND METABOLISM

19. OXIDATIVE PHOSPHORYLATION AND PHOTOPHOSPHORYLATION

19.3. Regulation of Oxidative Phosphorylation

The bulk of the ATP required by aerobic Cells is synthesized by Oxidative Phosphorylation. For every molecule of glucose completely oxidized to CO₂, 30 to 32 molecules of ATP are formed (Table 19-5). The Complete oxidation of palmitoyl (16:0)-CoA, which also takes place in the mitochondrial matrix, yields 108 molecules of ATP per molecule of palmitoyl-CoA (see Table 17-1). Similar calculations of ATP yield can be made for The oxidation of each of the Amino Acids, as discussed in detail in Chapter 18. By comparison, in Glycolysis under anaerobic conditions—that is, during Lactic acid Fermentation—only 2 molecules of ATP are formed per oxidized glucose molecule. These Examples show that the evolutionary development of oxidative phosphorylation led to a dramatic increase in the energetic efficiency of catabolic processes. Oxidation processes in aerobic cells involve The transfer of electrons along the mitochondrial Cell/36.html">Respiratory Chain and are accompanied by oxidative phosphorylation; it is critically important to be able to determine The amount of ATP formed and to understand the mechanism regulating its production in response to the changing needs of The Cell.

Class="center">Table 19-5. ATP Yield from Complete Oxidation of Glucose

* The number of ATP molecules formed depends on the type of shuttle system used to transfer reducing equivalents into the Mitochondria.

ATP Production in Oxidative phosphorylation is Regulated by Cellular Energy Needs

The rate of mitochondrial Respiration—that is, the rate of O₂ consumption—is regulated with high precision in cells. In general, the rate of oxygen consumption is limited by the concentration of ADP, the phosphorylation of which in mitochondria yields ATP. The dependence of the rate of oxygen consumption on the concentration of ADP (the phosphate acceptor) is known as acceptor control of respiration. The acceptor control ratio is The ratio of maximal oxygen consumption in the presence of ADP to that in the resting state, and in various animal Tissues this ratio is 10 or greater.

One characteristic of the cellular energy state is the intracellular concentration of ADP, which can be expressed through the mass-action ratio of the ATP-ADP system: [ATP] / [ADP] [Pi]. Typically, this ratio is very high, meaning that the ATP-ADP system is almost fully phosphorylated. If the rate of any cellular process requiring Energy Expenditure (ATP consumption), such as Protein Synthesis, increases, the rate of cellular ATP Cleavage to ADP and phosphate rises, causing the mass-action ratio to decrease. The resulting increase in ADP concentration automatically leads to an elevation in the rates of electron transfer and OXIDATIVE PHOSPHORYLATION AND, consequently, to an enhanced regeneration of ATP from ADP. This continues until the mass-action ratio returns to its normal high value, at which point respiration slows down again. The rate of cellular fuel oxidation is usually regulated with such sensitivity and precision that in most tissues the [ATP] / [ADP] [P_i] ratio fluctuates within very narrow limits, even when energy demands change. In other words, ATP is produced at a rate just sufficient to compensate for its consumption in energy-requiring processes.

During Hypoxia, Cellular ATP Hydrolysis Is Blocked by a Protein Inhibitor

As shown in previous sections (see Fig. 11-39), the enzyme ATP synthase can be regarded as a pump that extrudes protons from the mitochondrial matrix into the intermembrane space, driven by ATP energy and catalyzing the reverse of ATP synthesis. If a cell enters a state of hypoxia—that is, oxygen deprivation, as occurs during a Heart attack or stroke—the flow of electrons to oxygen along the mitochondrial respiratory chain ceases, and consequently, proton extrusion from the matrix is halted. The proton-motive force thus dissipates. Under these conditions, one might expect the ATP synthase enzyme to catalyze the reverse reaction—ATP hydrolysis—to supply energy for pumping protons out of the matrix. However, this would deplete cellular ATP to lethal levels. Therefore, no ATP hydrolysis occurs; the ATPase (hydrolytic) activity of ATP synthase under these conditions is blocked by a small 84-amino-acid protein inhibitor, IF₁ (Fig. 19-31). The IF₁ inhibitor binds simultaneously to two ATP synthase molecules. The activity of IF₁ is manifested at pH < 6.5, when it exists as a dimer. In the absence of oxygen, ATP is produced in cells primarily via glycolysis, a process that also yields pyruvic and lactic acids, which lower the pH of the Cytosol and the mitochondrial matrix. Thus, when oxygen supply to cells is cut off, conditions are created for The formation of the dimeric form of the IF₁ protein, which blocks the hydrolytic activity of ATP synthase, thereby preventing futile and wasteful ATP hydrolysis. Upon the resumption of aerobic metabolic processes, the concentration of pyruvic acid decreases and the cytosolic pH rises, causing the IF₁ protein complex dimer to dissociate and restoring the Functions of the ATP synthase enzyme.

Fig. 19-31. Structure OF THE regulatory protein IF₁ complex with F₁-ATPase isolated from bovine cells (PDB ID 1OHH). The protein inhibitor IF₁ (red helix) binds to two molecules of the F₁ enzyme (for its structure, see Fig. 19-25, d) via binding sites located between adjacent α,β-subunits, in the α-ADP and β-ADP Conformations (respectively) within the F₁ "HEAD". Light helices represent PARTS OF THE IF₁ inhibitor that are easily resolved in crystals of isolated IF₁, but cannot be resolved in crystals of its complex with the F₁ enzyme. The IF₁-F₁ enzyme complex exists only at low cytosolic pH—that is, during glycolytic ATP production. Upon the resumption of aerobic metabolic processes, the cytosolic pH increases, the inhibitory effect ceases, and ATP synthase functions are restored.

Hypoxia Leads to ROS Generation and Certain Adaptive Responses

Under hypoxic conditions, the balance between the influx of electrons from the oxidation of fuel molecules in the mitochondrial matrix and the transfer of electrons to molecular oxygen is disrupted in cells, resulting in accelerated Generation of reactive oxygen species (ROS). Along with Glutathione peroxidase (Fig. 19-18), cells possess two other lines of defense against oxygen radicals (Fig. 19-32). The first involves the Regulation of the pyruvate dehydrogenase (PDH) complex, which supplies acetyl-CoA to The Citric Acid cycle (Chapter 16). Under hypoxic conditions, pyruvate dehydrogenase kinase phosphorylates mitochondrial PDH, inactivating it and slowing the supply of FADH₂ and NADH from the citric acid cycle to the respiratory chain. The second pathway preventing ROS generation consists in replacing one subunit of complex IV, known as cytochrome c oxidase subunit 4-1 (COX4-1), with another subunit, cytochrome c oxidase subunit 4-2 (COX4-2), which is better adapted to hypoxic conditions. The catalytic properties of complex IV containing COX4-1 are optimal for respiration at normal oxygen concentrations, whereas in the presence of COX4-2, complex IV is optimized for hypoxic conditions.

Fig. 19-32. Hypoxia-inducible factor HIF-1 regulates the expression of genes that reduce the formation of reactive oxygen species (ROS). Under oxygen deficiency (hypoxia), HIF-1 is synthesized in large quantities and acts as a METABOLISM/31.html">Transcription factor that upregulates the synthesis of a glucose transporter, glycolytic Enzymes, pyruvate dehydrogenase kinase, Lactate dehydrogenase, the cytochrome c oxidase subunit COX4-2, and a protease that degrades the cytochrome c oxidase subunit COX4-1. These changes prevent ROS generation by reducing the synthesis of NADH and FADH₂, as well as by optimizing the action of complex IV cytochrome c oxidase. Bold gray arrows indicate reactions stimulated by HIF-1, and thin dashed arrows indicate reactions inhibited by HIF-1.

The alteration of PDH activity and the replacement of COX4-1 with COX4-2 within complex IV occur with the participation of hypoxia-inducible factor 1 (HIF-1). During hypoxia, this substance accumulates in cells and acts as a transcription factor, enhancing the synthesis of pyruvate dehydrogenase kinase, COX4-2, and proteinases that degrade COX4-1. Recall that HIF-1 also mediates changes in glucose transport and glycolytic enzymes, leading to the Pasteur Effect (a decrease in the rate of glucose consumption and the cessation of lactate accumulation in the presence of oxygen; see Box 14-1).

If these aforementioned defense mechanisms against ROS prove insufficient (which may be due to genetic Mutations in one of the protective Proteins or to extremely high rates of ROS generation), mitochondrial function is impaired. It is possible that mitochondrial dysfunction contributes to the Aging process, The Development of heart disease, certain rare forms of diabetes (see below), and maternally inherited genetic diseases affecting The Nervous system. ■

All Stages of ATP Synthesis in Carbohydrate Catabolism Are Coordinated by Interconnected Regulatory Mechanisms

The major catabolic stages possess their own interconnected regulatory mechanisms, thanks to which all stages of catabolism function in a unified, economical, and self-regulating regime. The processes of ATP formation and certain intermediates used as precursors in the biosynthetic reactions of other cellular components proceed in a coordinated manner. The relative concentrations of ATP and ADP—in other words, the mass-action ratio of the ATP-ADP system—determine not only the rate of electron transfer and oxidative phosphorylation, but also the rates of the citric acid cycle, pyruvate oxidation, and glycolysis (Fig. 19-33). When ATP consumption increases—that is, its concentration decreases—the concentrations of ADP and phosphate rise, and the rates of electron transfer and oxidative phosphorylation increase. Simultaneously, the rate of pyruvate oxidation via the citric acid cycle increases, thereby enhancing the flux of electrons into the respiratory chain. These events in turn lead to an increased rate of glycolysis, ensuring an enhanced production of pyruvate. Conversely, when the ADP concentration drops to a low level As a result of its conversion to ATP, electron transfer and oxidative phosphorylation slow down. The citric acid cycle and glycolysis also slow down because ATP acts as an allosteric inhibitor of the enzymes Phosphofructokinase-1 (see Fig. 15-14) and pyruvate dehydrogenase (see Fig. 16-18).

Fig. 19-33. Regulation of ATP-producing pathways. The interdependent Regulation of glycolysis, pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation, determined by the relative concentrations of ATP, ADP, AMP, and NADH. At high ATP concentrations and, accordingly, low ADP and AMP concentrations, the rates of glycolysis, pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation are minimal. If cellular ATP consumption increases sharply—and thus ADP, AMP, and phosphate concentrations rise—all four of these processes are accelerated. The interconnection between glycolysis and the citric acid cycle, mediated by citrate inhibiting glycolysis, complements the regulatory action of the adenine nucleotide system. In addition, elevated concentrations of NADH and acetyl-CoA inhibit the Oxidation of Pyruvate to acetyl-CoA, whereas a high [ATP] / [ADP] ratio inhibits dehydrogenase reactions in the citric acid cycle (see Fig. 16-18).

Phosphofructokinase-1 is also inhibited by citrate, the first intermediate of the citric acid cycle. During a futile cycle, citrate accumulates in the mitochondria and is then released into the cytosol. When ATP and citrate concentrations begin to exceed their normal levels, they act in concert to cause allosteric inhibition of phosphofructokinase-1, with The Effect of this dual inhibition being greater than the sum of the individual effects.

Summary of Section 19.3 Regulation of Oxidative Phosphorylation

■ Oxidative phosphorylation is regulated by the energy demands of the cell. The intracellular ADP concentration and the mass-action ratio [ATP] / [ADP] [P_i] reflect the energetic state of cells.

■ During hypoxia, i.e., when oxygen supply to cells is inadequate, a protein inhibitor blocks the enzyme ATP synthase, which under such conditions would otherwise catalyze ATP hydrolysis, thereby preventing the ATP concentration from dropping to levels dangerous to the cell.

■ Adaptation to hypoxic conditions, mediated by HIF-1, involves slowing down Electron transport along the respiratory chain and optimizing complex IV to function efficiently under oxygen-deficient conditions.

■ The rates of glycolysis, the citric acid cycle, and oxidative phosphorylation are coordinated with one another. This coordination is achieved through interconnected regulatory mechanisms that respond to ATP and ADP concentrations.