Biochemistry and Molecular Biology - Belyasova N.A. 2002

Metabolism. Processes Leading to Energy Storage
Catabolic Pathways
Carbohydrate Catabolism

CARBOHYDRATES serve as the primary sources of carbon and energy—and consequently, as nutritional substrates—for the vast majority of organisms (excluding plants). Among them, Cellulose, its derivatives, and starch are the most widespread on the planet. Besides these Polysaccharides, Glycogen, inulin, Chitin, xylans, pectin substances, and others are of great importance. Most of these listed polysaccharides are cleaved by specific Enzymes into Monosaccharides, with hexoses and pentoses predominating. Often, the Cleavage of poly- and Oligosaccharides is mediated by phosphorylases, resulting in phosphorylated products.

A characteristic feature of sugars is the presence of an oxygen atom at each carbon atom, which enables chemical attack of these substrates at virtually any point in the molecule. Furthermore, monosaccharides, and primarily their phosphorylated forms, are capable of isomerization: carbonyl groups and hydrogen atoms can easily migrate to an adjacent position or alter their spatial arrangement within the molecule with the aid of isomerases. Thus, a transition from any hexose or pentose to any other isomeric one becomes possible. For this reason, despite the diversity and complexity of Carbohydrate METABOLISM, several typical pathways of their conversion—particularly Catabolism—can be distinguished, possessing pronounced distinctive features. Such pathways include Glycolysis, the Pentose Phosphate Pathways, and the Entner–Doudoroff pathway.

A regular feature of monosaccharide catabolism is the mandatory initial activation stage of free monoses, which is accomplished via phosphorylation. As a result, phosphoric esters of monosaccharides are formed, which are capable of entering further transformations.

Glycolysis. This mode of sugar catabolism is otherwise known as the fructose-1,6-bisphosphate pathway (named after the key intermediate) or the Embden–Meyerhof–Parnas pathway (after its investigators).

Glycolysis is considered the most universal and energetically favorable pathway of hexose catabolism. The process was discovered in 1897 by the Buchner brothers, and its name derives from two Greek roots: glicos (sugar) and lysis (to dissolve). During glycolysis, a multi-step conversion of hexoses into Pyruvate occurs without the requirement for molecular gas, accompanied by The formation of ATP and reducing equivalents.

The reactions of The Glycolytic Pathway take place in the Cytosol. All intermediates are in a phosphorylated form. The substrate-level phosphorylation mechanism is utilized for energy storage.

The conversion of glucose into pyruvate (Fig. 9.3) requires the participation of ten enzymes and proceeds through the following stages: preparation for the Cleavage of the hexose chain, chain cleavage and Formation of glyceraldehyde-3-phosphate, and the First and Second substrate-level phosphorylations.

The process begins with the phosphorylation of glucose (molecule activation) involving ATP (as a phosphate group donor) and the enzyme hexokinase. The resulting glucose-6-phosphate is isomerized in the next reaction by glucosephosphate isomerase into fructose-6-phosphate, which undergoes a second phosphorylation driven by ATP to yield fructose-1,6-bisphosphate.

Fructose-1,6-bisphosphate is a key intermediate of the glycolytic pathway: it is this specific compound that distinguishes glycolysis from other sugar catabolism pathways. Furthermore, the Regulation of the rate of the entire glycolytic process occurs at the level of this substance. The activity of The enzyme catalyzing this reaction (Phosphofructokinase) is inhibited by high concentrations of ATP, which simultaneously decreases the affinity of phosphofructokinase for its substrate, fructose-6-phosphate. In addition, phosphofructokinase is inhibited by citrate, an early intermediate of The Tricarboxylic Acid Cycle. Activation of phosphofructokinase is also known: it is brought about by ADP and inorganic phosphate. Thus, phosphofructokinase is most active under conditions where cellular ATP is low (ADP is high) and building blocks are scarce.

At the next stage of glycolysis, fructose bisphosphate is cleaved by fructose-bisphosphate aldolase into two triose phosphates: dihydroxyacetone phosphate and glyceraldehyde triphosphate. These products are isomers and readily interconvert under the action of Triosephosphate isomerase. However, the direction of this reaction is shifted toward the formation of glyceraldehyde-3-phosphate because the latter is continually removed from the reaction mixture as it undergoes further transformations. Glyceraldehyde-3-phosphate serves as the substrate for the first substrate-level phosphorylation.

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Fig. 9.3. The glycolytic pathway of hexose catabolism. Zigzag lines indicate high-energy chemical bonds.

During the remaining stages of glycolysis, unlike the preceding ones, energy is released and stored in the form of ATP. One such reaction is The oxidation of glyceraldehyde-3-phosphate. The enzyme catalyzing this reaction (glyceraldehyde-3-phosphate dehydrogenase) uses NAD as a coenzyme and is characterized by a high content of sulfhydryl groups (SH-groups). Oxidation begins with the binding of glyceraldehyde-3-phosphate to the SH-group of the enzyme, forming an enzyme–substrate complex. Next, the enzyme catalyzes The transfer of hydrogen from the substrate to NAD⁺, and the reduced coenzyme dissociates. This dehydrogenation reaction serves as the oxidative reaction that supplies energy: a high-energy enzyme–acyl residue complex (thioester) arises. Then, phosphorolysis takes place—the transfer of the glyceraldehyde-3-phosphate residue along with the high-energy bond to phosphoric acid, leading to the formation of 1,3-diphosphoglyceric acid and the initial form of the enzyme. 1,3-Diphosphoglycerate represents a mixed anhydride of phosphoric and carboxylic acids and possesses a high Phosphate group transfer potential. The energy-rich phosphate group is transferred from 1,3-diphosphoglycerate to ADP with the participation of phosphoglycerate kinase, yielding the products 3-phosphoglycerate and ATP. This is how the first substrate-level phosphorylation in glycolysis is accomplished.

The final stage of glycolysis—the second substrate-level phosphorylation—begins with an intramolecular rearrangement during which 3-phosphoglycerate is isomerized to 2-phosphoglycerate (by the enzyme phosphoglycerate mutase). 2-Phosphoglycerate is dehydrated to phosphoenolpyruvate with the participation of enolase. As a result of this reaction, a compound characterized by a high phosphate group transfer potential is formed; thus, the elimination of a Water molecule from 2-phosphoglycerate is accompanied by the redistribution of energy within the molecule, and the phosphate bond at the C-2 carbon atom is converted from a low-energy to a high-energy one. The final reaction of glycolysis is catalyzed by pyruvate kinase, during which the phosphate group is transferred to an ADP molecule to produce the pathway's end product, pyruvate.

The glycolytic cleavage of a single glucose molecule leads to the formation of 4 ATP molecules (two for each glyceraldehyde-3-phosphate molecule), of which 2 are consumed in the Formation of fructose bisphosphate. Thus, a net total of only 2 ATP molecules is stored. In addition, 2 molecules of NADH are stored per glucose molecule in this process (upon the oxidation of 2 molecules of glyceraldehyde-3-phosphate). The glycolytic balance sheet is as follows:

1 С₆Н₁₂О₆ → 2 С₃Н₄О₃ + 2 АТР + 2NADH

The glycolytic process serves The Cell for storing energy and reducing equivalents, and also acts as a supplier of "building blocks" in the form of a three-carbon, partially oxidized compound—pyruvic acid—and certain intermediates, particularly glyceraldehyde-3-phosphate.

Pentose phosphate pathways. These pathways of sugar catabolism are quite diverse. Depending on the conditions and the species of the Organism, the outcome of these processes can yield various substances. The operation of the Pentose Phosphate Pathway reactions in reverse is utilized by Cells during CO₂ fixation. The primary distinguishing feature of these pathways is the formation of five-carbon sugars as intermediates, as well as the sequential removal of one carbon atom at a time from a six-carbon phosphorylated sugar, which is released into the environment as CO₂.

The pentose phosphate pathways (otherwise known as the Warburg–Dickens–Horecker scheme, the hexose monophosphate shunt, or the phosphogluconate pathway) are utilized by organisms less frequently than glycolysis and are most often found in bacterial cells.

Following the traditional glucose activation reaction via phosphorylation, the resulting glucose-6-phosphate undergoes dehydrogenation. This reaction is catalyzed by glucose-6-phosphate dehydrogenase, and its coenzyme, NADP⁺, accepts the reducing equivalents. 6-Phosphogluconolactone is formed (Fig. 9.4). This compound undergoes Hydrolysis (ring opening) with the participation of gluconolactonase, and the resulting 6-phosphogluconic acid undergoes a second dehydrogenation (phosphogluconate dehydrogenase), which is immediately followed by decarboxylation to yield ribulose-5-phosphate. Ribulose-5-phosphate is isomerized into two five-carbon sugars: xylulose-5-phosphate and ribose-5-phosphate (Fig. 9.4).

The next stage of the pentose phosphate pathways represents multiple intermolecular rearrangements and the isomerization of intermediates formed from ribulose-5-phosphate. These reactions are directed toward the formation of a six-carbon compound (glucose-6-phosphate), which can once again undergo oxidation and decarboxylation. As a result, 5 molecules of glucose-6-phosphate are produced from 6 molecules of ribulose-5-phosphate (Fig. 9.5).

The sugar structural rearrangement system includes two enzymes—transketolase and transaldolase—which catalyze the transfer of two- and three-carbon fragments by cleaving C—C bonds at two positions: adjacent to the carbonyl carbon (a) and at the carbon atom adjacent to the carbonyl group (ß) (in Fig. 9.5, the sites where bonds undergo cleavage are indicated by a dashed line).

Upon Complete oxidation of glucose via the pentose phosphate pathways, the glyceraldehyde-3-phosphate molecule formed at the final stage undergoes isomerization, mediated by triosephosphate isomerase, into dihydroxyacetone phosphate, which then enters an aldol Condensation reaction with a second molecule of glyceraldehyde-3-phosphate to yield fructose bisphosphate. This reaction is the reverse of the fructose bisphosphate cleavage that takes place in glycolysis (Fig. 9.3), and the enzyme fructose bisphosphate aldolase catalyzes both forward and reverse transformations. A molecule of fructose-1,6-bisphosphate is dephosphorylated to yield fructose-6-phosphate (by the enzyme phosphatase), which, in turn, is isomerized to glucose-6-phosphate and returns to the initial dehydrogenation stage.

Fig. 9.4. Formation and isomerization of ribulose-5-phosphate during the pentose phosphate pathways. Since 6 molecules of CO₂ can be formed from a single hexose molecule, for the convenience of balancing, the process is presented per 6 molecules of glucose-6-phosphate.

Thus, the complete oxidation of a single glucose molecule via the pentose phosphate pathway consumes 1 ATP molecule and yields 6 molecules of СО₂ and 12 molecules of NADPH. Therefore, The primary function of the pentose phosphate pathways is to supply the cell with NADPH, which is utilized in biosynthetic processes.

Complete oxidation of glucose does not necessarily occur during the pentose phosphate pathway. In this case, glyceraldehyde-3-phosphate undergoes alternative transformations, such as substrate-level phosphorylation catalyzed by glycolytic enzymes.

The pentose phosphate pathways play a crucial role in biosynthetic processes by generating intermediates that serve as precursors for numerous compounds. Specifically, erythrose-4-phosphate is utilized by Bacteria and plants for the synthesis of aromatic Amino Acids. Ribose-5-phosphate acts as a substrate for the synthesis of nitrogenous bases and Certain amino acids.

Fig. 9.5. Conversions of ribulose-5-phosphate isomers during the pentose phosphate pathway upon complete glucose oxidation (per 6 molecules of glucose-6-phosphate). Explained in the text

The Entner–Doudoroff pathway (2-keto-3-deoxy-6-phosphogluconate pathway). This process represents another mechanism of hexose catabolism and is predominantly found in microbial cells. This pathway is of particular importance for gluconate degradation. The Initial Stages of glucose-6-phosphate conversion are identical to those of the pentose phosphate pathway, up to the formation of 6-phosphogluconic acid. Subsequently, however, 6-phosphogluconate undergoes dehydration rather than oxidation, mediated by the enzyme 6-phosphogluconate dehydratase. This yields the key intermediate of the pathway, 2-keto-3-deoxy-6-phosphogluconate. The dehydration step proceeds via an intermediate enol form, which undergoes tautomeric rearrangement to form 2-keto-3-deoxy-6-phosphogluconate (Fig. 9.6).

Next, aldolase catalyzes the cleavage of this key intermediate into pyruvate and glyceraldehyde-3-phosphate. The latter can then enter the glycolytic pathway and undergo further transformations.

Fig. 9.6. The Entner–Doudoroff pathway and associated processes

As indicated by the Stoichiometry of the Entner–Doudoroff pathway, this process is energetically less efficient than glycolysis: only 1 ATP molecule is conserved per glucose molecule (out of two synthesized, one is consumed in the initial phosphorylation of glucose).

An Overview of the principal Catabolic pathways reveals that The breakdown of fuel molecules via incomplete oxidation yields two primary metabolites: pyruvic acid and acetyl-CoA. These compounds can either serve as building blocks in biosynthetic pathways or undergo further reactions to ensure cellular energy storage. Since this section focuses on energy-yielding processes, it should be noted that the metabolic fate of pyruvate and acetyl-CoA depends primarily on the availability of molecular oxygen within the cell and its enzymatic repertoire. For instance, in anaerobic microorganisms and Tissues of higher organisms, pyruvate and acetyl-CoA can undergo Various Forms of Fermentation, which complete the catabolic breakdown of substrates. In aerobic and facultatively anaerobic cells, in the presence of molecular oxygen, pyruvate can undergo oxidative decarboxylation, and the resulting acetyl-CoA enters the tricarboxylic acid cycle, where it is fully oxidized to СО₂ and Н₂О, while the generated reduction equivalents enter the Respiratory Chain. These processes will be discussed in detail in the following chapters.