Biochemistry and Molecular Biology - Belyasova N.A. 2002

Metabolism. Processes Leading to Energy Storage
Tricarboxylic Acid Cycle
Chemistry of the TCA Cycle

In eukaryotes, the Reactions of the tricarboxylic acid (TCA) cycle take place in the Cell/35.html">Mitochondria, precisely where the systems supplying "fuel" for the TCA cycle function: fatty acid ß-Oxidation and Pyruvate oxidative decarboxylation. It is believed that the Enzymes participating in the TCA cycle are organized into a specific catalytic complex located between The inner mitochondrial membranes. The Structure of this complex is such that intermediate products of the cycle are sequentially transferred from one enzyme to another without being released into the matrix. This significantly increases the process rate.

All TCA cycle reactions can be conditionally divided into 4 stages: 1) acceptance of acetyl-CoA and formation of isocitrate; 2) Oxidative Decarboxylation of isocitrate; 3) oxidative decarboxylation of a-ketoglutarate; 4) regeneration of the acetyl-CoA acceptor.

The acceptor of acetyl-CoA is oxaloacetate (oxaloacetic acid, OAA). This four-carbon compound condenses with the two-carbon component of acetyl-CoA with the participation of the enzyme citrate synthase. This aldol Condensation reaction proceeds stepwise with The formation of an intermediate product, citryl-CoA, the Hydrolysis of which shifts the reaction equilibrium toward the formation of citrate (Fig. 11.2).

For The oxidative decarboxylation of the six-carbon compound to proceed, citrate (citric acid) must be isomerized into isocitrate. This process is catalyzed by aconitase and represents a dehydration reaction followed by Hydration.

The oxidative decarboxylation of isocitrate is catalyzed by isocitrate dehydrogenase, which uses NAD as a coenzyme. Isocitrate is oxidized to the ß-keto acid oxalosuccinate, which rapidly loses its carboxyl group while bound to the enzyme and is converted into a-ketoglutarate (Fig. 11.3).

At the Third Stage (Fig. 11.4), a-ketoglutarate also undergoes oxidative decarboxylation. This reaction is catalyzed by an organized enzyme assembly, the a-ketoglutarate dehydrogenase complex. The composition of this assembly is similar to the complex catalyzing the oxidative decarboxylation of pyruvate, and both processes share many common features. The reaction product is a four-carbon activated compound, succinyl-CoA. The high-energy bond of the succinyl thioester is cleaved with the participation of succinate thiokinase, which is coupled with the phosphorylation of GDP (in mammals) or ATP (in Bacteria and higher plants). The Formation of the high-energy GTP or ATP bond during succinyl-CoA hydrolysis is the only instance of substrate-level phosphorylation that occurs in the TCA cycle.

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Fig. 11.2. First stage of the TCA cycle: formation of isocitrate

The succinate (succinic acid) formed in the previous stage is converted into oxaloacetate through a series of reactions (oxidation, hydration, oxidation) (Fig. 11.5). This sequence of transformations represents a process of ß-oxidation, which was already discussed with respect to Fatty acids in Chapter 9. The reactions of The final stage of the TCA cycle constitute the regeneration of oxaloacetate. The oxidation of succinate is catalyzed by succinate dehydrogenase. Unlike other TCA cycle enzymes, this enzyme is directly linked to the Electron Transport Chain and Functions as an integral protein of the inner mitochondrial membrane.

The oxidation product of succinate—fumarate—is hydrated by fumarase. A stereospecific addition of hydrogen and hydroxyl groups takes place, yielding L-malate (malic acid). Finally, L-malate is oxidized to oxaloacetate by malate dehydrogenase.

The fully cyclic reaction System of the TCA cycle is illustrated in Fig. 11.6. Many TCA cycle intermediates serve as precursors in biosynthetic reactions. For instance, oxaloacetate and a-ketoglutarate are utilized in Amino acid Biosynthesis. Oxaloacetate is also incorporated into Gluconeogenesis (the Synthesis of glucose from non-carbohydrate substrates). Most of the carbon atoms in Porphyrins originate from succinyl-CoA. However, the withdrawal of TCA cycle intermediates for biosynthetic purposes must be continuously replenished; otherwise, the cycle will be interrupted. Under normal conditions, a steady-state dynamic equilibrium exists between the reactions that deplete and those that replenish these intermediates in the cycle.

Fig. 11.4. Third stage of the TCA cycle: oxidative decarboxylation of a-ketoglutarate

Fig. 11.5. Final stage of the TCA cycle: regeneration of oxaloacetate

Fig. 11.6. Tricarboxylic Acid Cycle. Dashed lines indicate the reactions of The Glyoxylate cycle

Reactions that replenish the pool of TCA cycle intermediates are termed anaplerotic. One of the primary Anaplerotic reactions in mammalian Cells is the carboxylation of pyruvate to form oxaloacetate. This conversion is catalyzed by the allosteric enzyme pyruvate carboxylase, which is activated by the TCA cycle substrate acetyl-CoA. When acetyl-CoA accumulates, active carboxylation of pyruvate is stimulated, increasing the supply of oxaloacetate, which serves as the immediate acceptor for acetyl-CoA.

In bacterial and plant cells, the replenishment of oxaloacetate is carried out by a different enzyme: phosphoenolpyruvate carboxylase.