BIOCHEMISTRY: A TEXTBOOK FOR MEDICAL UNIVERSITIES - E. S. Severin - 2004

SECTION 6. ENERGY METABOLISM

Living organisms exist in a constant and inseparable relationship with their environment. This interaction is maintained through METABOLISM. Metabolic processes comprise three stages: the uptake of substances into the Organism, Intermediary Metabolism, and The excretion of end products from the organism.

Substances enter the organism through Respiration (oxygen) and Nutrition. Food products are digested within the gastrointestinal tract, where they are broken down into simpler substances. Digestion involves the Hydrolysis of polymers (Proteins, Polysaccharides, and other complex organic molecules) into monomers, which are then absorbed into the bloodstream and integrated into intermediary metabolism.

Intermediary metabolism (intracellular metabolism) includes Two Types of reactions: Catabolism and anabolism (Fig. 6-1).

Class="center">Fig. 6-1. General scheme of energy and substance metabolism. 1 — digestion; 2 — catabolism; 3 — anabolism; 4 — breakdown of Structural and functional Cell components; 5 — exergonic reactions; 6, 7 — endergonic reactions; 8 — elimination from the organism.

Catabolism — The process of breaking down organic molecules into end products. In animals and humans, the End products of organic substance transformations are CO₂, H₂O, and urea. Catabolic processes involve metabolites produced both during digestion and through The breakdown of structural and functional cellular components.

Catabolic reactions are accompanied by the release of energy (exergonic reactions).

Anabolism encompasses BIOSYNTHETIC PROCESSES IN which simple building blocks are assembled into complex macromolecules essential for the organism. Anabolic reactions utilize The energy released during catabolism (endergonic reactions).

I. Biological Oxidation

Catabolic processes in animal Cells are accompanied by the consumption of oxygen, which is required for oxidation reactions. These reactions release the energy that organisms need to perform various types of work necessary for life processes. While non-biological systems can perform work using thermal energy, biological systems function under isothermal conditions and rely on chemical energy to sustain life. The Study of energy transformations accompanying Chemical Reactions is the domain of bioenergetics, or biochemical Thermodynamics.

A. Free energy and the Laws of Thermodynamics

From a thermodynamic standpoint, Living organisms are open systems. Energy exchange can occur between the System and Its environment in accordance with the laws of thermodynamics.

1. Laws of Thermodynamics

The First Law is the law of conservation of energy, which can be formulated as follows: the total energy of a system and its surroundings is constant.

Within the system under consideration, energy can be transferred from one part to another or transformed from one form into another.

The Second Law states that all Physical and Chemical processes in a system tend toward the irreversible conversion of useful energy into a chaotic, uncontrollable form. The measure of this transition or system disorder is a quantity called Entropy (S), which reaches its maximum when the system attains true equilibrium with its environment.

2. Free Energy

Every organic compound entering the organism from the outside or comprising living matter possesses a certain store of internal energy (E). A portion of this internal energy can be used to perform useful work. Such energy of the system is referred to as free energy (G).

At constant Temperature and pressure, the relationship between The change in the Free energy of the system (∆G) and the change in entropy (∆S) can be expressed by the following equation:

∆G = ∆Н — Т х S, where ∆Н is the change in enthalpy (internal energy or heat contained within the system), and Т is absolute temperature. Under the conditions in which biochemical reactions take place, ∆Н is approximately equal to ∆Е (the change in the internal energy of the system resulting from the reaction). In biological systems, free energy is typically measured under standard conditions: pH 7.0, temperature 25 °C, all solutions at a concentration of 1 mol/L, and all gases at a pressure of 1 atm.

Under standard conditions, all Functions are denoted as ∆G°′, ∆S°′, and ∆H°′. The change in Standard Free Energy (∆G°′) can be calculated by knowing the Equilibrium Constant (К'_eq) of a chemical reaction.

3. Endergonic and Exergonic Reactions

The direction of a chemical reaction is determined by the value of ∆G. If this value is negative, the reaction proceeds spontaneously and is accompanied by a decrease in free energy. Such reactions are called exergonic. If the absolute value of ∆G is large, the reaction goes virtually to completion and can be considered irreversible.

If ∆G is positive, the reaction will proceed only if free energy is supplied from an external source; such reactions are referred to as endergonic.

If the absolute value of ∆G is large, the system is stable, and the reaction practically does not take place. When ∆G is zero, the system is at equilibrium (Table 6-1).

Table 6-1. Relationship between K'_eq and ∆G°′ values and the direction of chemical reactions

4. Coupling of exergonic and endergonic processes in the body

In biological systems, thermodynamically unfavorable (endergonic) reactions can proceed only at the expense of energy released by exergonic reactions. Such reactions are known as energy-coupled reactions. Many of these processes occur with the participation of adenosine triphosphate (ATP), which acts as a coupling factor.

Let us examine the energetics of coupled reactions in more detail using glucose phosphorylation as an example.

The phosphorylation of glucose by free phosphate to form glucose-6-phosphate is an endergonic reaction:

(1) Glucose + H₃PO₄ —> Glucose-6-phosphate + H₂O (∆G = +13.8 kJ/mol).

For such a reaction to proceed toward The formation of glucose-6-phosphate, it must be coupled with another reaction whose free energy change exceeds the energy required for glucose phosphorylation.

(2) ATP —> ADP + H₃PO₄ (∆G = -30.5 kJ/mol).

When processes (1) and (2) are coupled in a reaction catalyzed by hexokinase (see Section 7), glucose phosphorylation proceeds readily under physiological conditions; the reaction equilibrium is strongly shifted to the right and is practically irreversible:

(3) Glucose + ATP —> Glucose-6-phosphate + ADP (∆G = -16.7 kJ/mol).

B. Characteristics of high-energy phosphates. The ATP-ADP cycle

Living organisms contain a distinct group of organic phosphates whose hydrolysis releases a large amount of free energy. Such compounds are called high-energy phosphates (Table 6-2).

Table 6-2. Free energy of hydrolysis for selected organic phosphates

As can be seen from Table 6-2, various phosphorylated compounds possess differing stores of free energy. In addition to ATP, the high-energy phosphate group includes enol phosphates, anhydrides, and phosphoguanidines. Compounds located at the bottom of the table constitute the low-energy phosphate group. ATP occupies a central position among these compounds (Fig. 6-2).

Fig. 6-2. Adenosine triphosphate (ATP). The ATP molecule contains two high-energy (macroergic) bonds, β and y, indicated in the figure by the tilde symbol (~).

ATP is an energy-rich molecule because it contains two phosphoanhydride bonds (β, y).

Hydrolysis of the terminal phosphoanhydride bond converts ATP into ADP and inorganic phosphate P_i. The associated change in free energy is -7.3 kcal/mol. Under normal cellular conditions (pH 7.0, temperature 37 °C), the actual ∆G°′ value for hydrolysis is approximately -12 kcal/mol. The free energy of ATP hydrolysis enables its formation from ADP via Phosphate group transfer from high-energy phosphates such as phosphoenolpyruvate or 1,3-bisphosphoglycerate. Conversely, ATP can drive endergonic reactions such as the phosphorylation of glucose or glycerol. ATP serves as an energy donor in the endergonic reactions of many anabolic processes. Certain biosynthetic reactions in the body can proceed with the participation of other nucleoside triphosphates, which are analogues of ATP, including guanosine triphosphate (GTP), uridine triphosphate (UTP), and cytidine triphosphate (CTP). All of these NUCLEOTIDES are, in turn, generated utilizing the free energy of the terminal phosphate group of ATP. Finally, the free energy of ATP powers Various Forms of work underlying vital cellular functions, such as Muscle contraction or active Transport of substances.

Thus, ATP is the primary, directly utilized donor of free energy in biological systems. Within The Cell, an ATP molecule is consumed within a minute of its formation. In humans, an amount of ATP equivalent to the total body weight is synthesized and degraded every 24 hours.

The utilization of ATP as an energy source is possible only if ATP is continuously resynthesized from ADP using the energy derived from The oxidation of Organic compounds (Fig. 6-3). The ATP-ADP cycle is the core mechanism of energy exchange in biological systems, and ATP serves as the universal "energy currency".

Fig. 6-3. The ATP-ADP cycle.

B. Oxidation-Reduction Reactions. Oxidation-Reduction Potential

Oxidation is defined as the loss of electrons, whereas reduction is the gain of electrons. The oxidation of an electron donor is always accompanied by the reduction of an electron acceptor. This principle of redox processes also applies to biochemical systems. Any oxidation-reduction reaction involves an electron acceptor (oxidizing agent) and an electron donor (reducing agent). For example,

(1) Сu + О —> Сu^{2 +} O^2-.

The overall reaction (1) can be conventionally divided into 2 half-reactions (2) and (3):

(2) Сu — 2е —> Сu²⁺.

(3) О + 2е —> О^2-.

Each of these involves the oxidized and reduced forms of a single compound, which are referred to as a conjugate pair, or a redox pair.

Table 6-3. Standard oxidation-reduction potentials of some conjugate pairs

Different redox pairs have varying affinities for electrons. Those with a lower affinity transfer electrons to those with a higher affinity. The measure of a redox pair's electron affinity is the oxidation-reduction potential, or redox potential (Е₀'), the value of which is directly related to the change in free energy. The value of Е₀' is expressed in volts; the lower (more negative) it is, the lower the substance's affinity for electrons. Conversely, the higher the affinity, the greater the reduction potential.

Redox potentials Е₀' are related to the change in free energy by the Nernst equation:

∆G₀' = - nF∆E₀'

where n is the number of electrons transferred in the reaction; F is the Faraday constant (23,061 kcal V^-1mol^-1); and ∆Е₀' is the difference in redox potentials between the electron-donor and electron-acceptor pairs.

The value ∆Е₀' represents the standard oxidation-reduction potential, determined under standard conditions where the concentrations of all substances are 1 M, gas pressure is 1 atm, and pH is 7.0 (Table 6-3).

C. MAIN STAGES OF Energy Transformation in Catabolic Processes

Energy is released during the enzymatic oxidation of metabolites by specific dehydrogenases. In dehydrogenation reactions, electrons and protons are transferred from organic substrates to the Coenzymes of NAD- and FAD-dependent dehydrogenases. High-energy electrons are passed from the reduced coenzymes NADH and FADH₂ to oxygen via a chain of carriers localized in The inner mitochondrial membrane. The reduction of the О₂ molecule occurs As a result of The transfer of 4 electrons. With each addition of 2 electrons (delivered to oxygen via the carrier chain) from the matrix, 2 protons are consumed, resulting in the formation of a molecule of Н₂О.

The oxidation of organic substances in cells, accompanied by oxygen consumption and Water synthesis, is called tissue respiration, and the Electron Transport Chain (ETC) is referred to as the Respiratory Chain.

As electrons entering the ETC move from one carrier to the next, they lose free energy. A significant portion of this energy is conserved in the form of ATP, while some is dissipated as heat. Additionally, high-energy electrons generated during the oxidation of various substrates can be utilized in biosynthetic reactions, which require reducing equivalents such as NADPH in addition to ATP.

D. Enzymes and Coenzymes Involved in Oxidation-Reduction Reactions

The transfer of electrons from oxidizable substrates to oxygen occurs in several stages. It involves A large number of intermediate carriers, each capable of accepting electrons from the preceding component and passing them on to the next. This forms a chain of oxidation-reduction reactions, ultimately leading to the reduction of О₂ and the synthesis of Н₂О. The mitochondrial respiratory chain includes a large number of carriers (Fig. 6-4).

Fig. 6-4. The Mitochondrial Electron Transport chain. Complex I contains FMN and at least five iron-sulfur (FeS) proteins. Complex III includes two different forms of cytochrome b (with absorption maxima at 562 and 566 nm), one FeS protein, and cytochrome с₁. Complex IV contains Cytochromes а and а₃ and two copper ions. Complex II (succinate dehydrogenase) is not shown in the figure (see Fig. 6-13). Complex V is ATP synthase.

With the exception of ubiquinone (CoQ), all Components of the ETC are proteins. These proteins contain various non-protein moieties: FMN, iron in iron-sulfur proteins and porphyrin rings, and Cu ions.

1. Primary Hydrogen Acceptors

Primary hydrogen acceptors in oxidation-reduction reactions belong to two types of dehydrogenases: nicotinamide-dependent, containing nicotinic acid derivatives as coenzymes, and flavin-dependent, containing riboflavin derivatives (see section 2).

Nicotinamide-dependent dehydrogenases contain NAD⁺ or NADP⁺ as coenzymes (see section 2). NAD⁺ and NADP⁺ are derivatives of Vitamin PP. These coenzymes are components of the active sites of dehydrogenases, but they can reversibly dissociate from the apoenzyme complex and become incorporated into the enzyme during the reaction. The substrates of NAD- and NADP-dependent dehydrogenases are located in the mitochondrial matrix and Cytosol. The functional part of nicotinamide coenzymes is nicotinamide (Fig. 6-5).

Fig. 6-5. Structural formulas of the functional part of NAD⁺ and NАDР⁺ coenzymes. In their oxidized form, nicotinamide coenzymes are designated as NAD⁺ and NАDР⁺ because they carry a positive charge on the nitrogen atom of the pyridine ring. In dehydrogenation reactions, out of the two hydrogen atoms split off from the oxidized substrate, the nicotinamide ring attaches a hydrogen ion and two electrons in the form of a hydride ion (: Н^-). The second ion passes into the medium. In the reverse reaction, NАDН (NАDРН) act as electron and proton Donors.

Most dehydrogenases that supply electrons to the ETC contain NAD⁺. They catalyze Reactions of the type:

R-CHOH-R₁+ NAD⁺<-> R-CO-R₁+ NADH + Н⁺.

Thus, by accepting protons and electrons from various substrates, NAD⁺ serves as the main collector of energy from oxidized substances and the primary source of high-potential electrons for the ETC.

NADPH is not a direct electron donor in the ETC, but is used almost exclusively in reductive biosyntheses (see section 8). However, electrons from NADPH can be introduced into the ETC through the action of pyridine nucleotide transhydrogenase, which catalyzes the reaction:

NADPH + NAD⁺<-> NADP⁺+ NADH.

Flavin dehydrogenases contain FAD or FMN as coenzymes. These coenzymes are synthesized in The Human Body from vitamin B₂ (see section 2). Flavin Coenzymes are tightly bound to apoenzymes. The functional part of FAD and FMN is an isoalloxazine conjugated ring system (Fig. 6-6).

FAD acts as an electron acceptor from many substrates in reactions of the type:

where E is the protein moiety of the enzyme.

Fig. 6-6. Structural formulas of the functional part of FAD and FMN coenzymes. During the reaction, FAD and FMN accept 2 electrons and, unlike NAD⁺, both protons lost by the substrate.

Most FAD-dependent dehydrogenases are soluble proteins localized in the mitochondrial matrix. An exception is succinate dehydrogenase, which is located in the inner mitochondrial membrane. FMN-containing enzymes include NADH dehydrogenase, which is also localized in the inner mitochondrial membrane; it oxidizes NADH generated in the mitochondrial matrix.

2. Electron transport chain from NADH and FADH₂ to oxygen

The transfer of electrons from NADH to O₂ involves a series of carriers localized in the inner mitochondrial membrane. With the exception of ubiquinone and cytochrome c, these are complex Structure/178.html">Protein Complexes.

NADH dehydrogenase (NADH-Q reductase, complex I) consists of several polypeptide chains. FMN plays The Role of the prosthetic group. The sole substrate of the enzyme is NADH, from which 2 electrons and a proton are transferred to FMN, forming FMNH₂. The second proton is taken up from the matrix. The reaction proceeds According to the equation:

NADH + Н⁺ + Е (FMN) —> NAD⁺ + Е (FMNH₂).

From FMNH₂, electrons are subsequently transferred to a series of iron-sulfur proteins (FeS), which act as a second prosthetic group in the NADH dehydrogenase molecule. The iron atoms in these proteins (non-heme iron) are grouped into several clusters, the so-called iron-sulfur centers. FeS centers are part of many proteins (Flavoproteins, cytochromes) involved in oxidation-reduction reactions. Three types of FeS centers are known (FeS, Fe₂S₂, Fe₄S₄), in which an iron atom is bound to sulfur atoms of Cysteine residues or inorganic sulfur. The structure of iron-sulfur centers is shown in Fig. 6-7.

Fig. 6-7. Structure of iron-sulfur centers. I — FeS center; the iron atom is coordinated to four sulfur atoms belonging to four cysteine residues in the protein. II — Fе₂S₂ center; each of the two iron atoms is coordinated to two inorganic sulfur atoms and two cysteine residues in the protein; III — Fе₄S₄ center; four iron atoms are bound to four sulfur atoms and four cysteine residues in the protein. The iron atoms in FeS centers can exist in an oxidized (Fe³⁺) or reduced (Fe²⁺) state.

NADH dehydrogenase contains several Fe₂S₂ and Fe₄S₄ type centers. The iron atoms in such centers can alternately accept and donate electrons, shifting between ferro- (Fe²⁺) and ferri- (Fe³⁺) states. From the iron-sulfur centers, electrons are transferred to coenzyme Q (ubiquinone) (Fig. 6-8).

Fig. 6-8. Structure of ubiquinone (coenzyme Q), where n is the number of isoprenoid units. Ubiquinone can accept one electron and be converted into semiquinone, or accept 2 electrons and be fully reduced to hydroquinone (ubiquinol).

The designation of this fat-soluble quinone originates from the first letter of the English word "quinone," while the name ubiquinone reflects its widespread distribution in nature (ubiquitous). Depending on the source from which they are isolated, ubiquinone molecules vary in the length of their hydrocarbon chain, which in mammals contains 10 isoprenoid units and is designated as Q₁₀. During the transfer of electrons from NADH dehydrogenase through FeS to ubiquinone, it is reversibly converted into hydroquinone. Ubiquinone performs a collector function by accepting electrons from NADH dehydrogenase and other flavin-dependent dehydrogenases, particularly succinate dehydrogenase. Ubiquinone participates in reactions of the following type:

Е (FMNH₂) + Q —> Е (FMN) + QH₂.

Cytochromes, or Hemoproteins, are present in all types of organisms. In Eukaryotic cells, they are localized in the mitochondrial membranes and the ER. Approximately 30 different cytochromes are known. All cytochromes contain a heme group as their prosthetic group (see Section 1). Their diversity is due to:

✵ differences in the side chains of the heme structure;

✵ differences in the structure of The polypeptide chains;

✵ differences in the mode of attachment of the polypeptide chains to the heme.

Depending on their ability to absorb light in a specific region of the spectrum, all cytochromes are divided into groups a, b, and c. Within each group, individual species with unique spectral properties are designated by numerical indices (b, b₁, b₂, etc.).

The Structural Features of Different types of cytochromes determine the differences in their oxidation-reduction potentials. The electron transport chain (ETC) involves 5 types of cytochromes (a, a₃, b, c, c₁). With the exception of cytochrome c, all cytochromes are located in the inner mitochondrial membrane as part of complex Protein Assemblies (Table 6-4).

QH₂ dehydrogenase (coenzyme Q-cytochrome c reductase, complex III) consists of 2 types of cytochromes (b₁ and b₂) and cytochrome c₁. QH₂ dehydrogenase transfers electrons from ubiquinol to cytochrome c. Within complex III, electrons are passed from cytochromes b to FeS centers, to cytochrome c₁, and then to cytochrome c. Heme groups, like FeS centers, transfer only one electron at a time. Thus, 2 electrons are transferred from a QH₂ molecule to 2 molecules of cytochrome b. The formation of a semiquinone free radical is possible as an intermediate product in these Electron transfer reactions. In type b cytochromes, the heme is not covalently bound to the protein, whereas in Cytochromes c₁ and c, it is attached to the protein via thioether bonds (Fig. 6-9). These bonds are formed by The addition of 2 cysteine residues to the vinyl groups of the heme.

Fig. 6-9. STRUCTURE OF THE heme in cytochromes b, c, c₁.

Cytochrome c is a peripheral water-soluble membrane protein with a Molecular Weight of 12,500 D, featuring a single polypeptide chain of 100 amino acid residues and a heme molecule covalently linked to the polypeptide.

Cytochrome c oxidase (complex IV) consists of 2 cytochromes of the aa₃ type, each of which possesses an oxygen-binding site. Cytochromes a and a₃ contain a characteristic iron-porphyrin prosthetic group called heme A, which differs from the heme of cytochromes c and c₁ (Fig. 6-10). It contains a formyl group instead of one of the methyl groups and a hydrocarbon chain instead of one of the vinyl groups.

Fig. 6-10. Structure of heme A.

Another feature of the a-a₃ complex is the presence of copper ions bound to the protein moiety in the so-called Cu_A centers. Electron transfer by the a-a₃ complex involves the reactions:

Сu⁺ <-> Сu²⁺ + е,

Fе²⁺ <->Fе³⁺ + е.

The cytochrome a-a₃ complex reacts directly with molecular oxygen. Some CHARACTERISTICS OF THE ETC components are given in Table 6-4.

Table 6-4. Components of the mitochondrial electron transport chain

E. Organization OF THE respiratory chain in Mitochondria

The main electron carriers are embedded in the inner mitochondrial membrane and organized into 4 complexes arranged in a specific (vectorial) sequence. In this sequence, their standard redox potentials become progressively more positive as they approach oxygen (Table 6-3, Fig. 6-11).

Each link in this chain is specific regarding its electron donor and acceptor.

At The First stage, dehydrogenases catalyze the removal of hydrogen from various substrates. If the substrates are α-hydroxy acids such as malate, isocitrate, or 3-hydroxybutyrate, hydrogen is transferred to NAD⁺. The resulting NADH is, in turn, oxidized by NADH dehydrogenase (complex I) in the respiratory chain.