Chem*4520 Metabolic Processes Fall Semester 2000 Modified August 2000
schematic view of the enzyme citrate synthase, with bound acetyl-CoA analog in green	Department of Chemistry and Biochemistry Home Page Back to Chem*4520 Home Page

Regulation of pyruvate entry to the TCA cycle

The initial substrate for the TCA cycle is acetyl CoA. Acetyl CoA may be made by pyruvate dehydrogenase, or by beta oxidation of fatty acids or by various amino acid breakdown pathways. Pyruvate in turn may derive from glycolysis or amino acid breakdown.

The TCA cycle is what is known as a non catalytic cycle. That means that one mole of acetyl CoA requires one mole of oxaloacetate making one mole of citrate which is converted to one mole of oxaloacetate. Thus the pathway can do no more than replace the oxaloacetate initially used. No matter how much acetyl CoA is fed in, it's not possible to produce more oxaloacetate than you started with. The ability to oxidize acetyl CoA depends on the availability of oxaloacetate.

Pyruvate carboxylase provides a way to jump start the TCA cycle, by replenishing or increasing the overall available oxaloacetate supply. Because pyruvate dehydrogenase is irreversible, oxaloacetate can't be made from acetyl CoA.

The two enzymes, pyruvate carboxylase and pyruvate dehydrogenase, control the use of pyruvate and are reciprocally regulated. Acetyl CoA activates carboxylase, and inhibits the dehydrogenase. High energy states favour carboxylase by weighting the equilibrium in favour of CO2 activation, but inhibit the dehydrogenase. Low energy states favour the dehydrogenase.

In addition, pyruvate dehydrogenase is inactivated by a specific protein kinase, and activated by a phosphatase. The phosphatase is Ca²⁺ sensitive, so elevated Ca²⁺ levels increase pyruvate dehydrognase activity.

Although some texts refer to Ca²⁺ release as a signal for muscle contraction and enhanced glycogenolysis in the cytoplasm, bear in mind that the TCA and pyruvate enzymes are in the mitochondrion and are partly isolated from cytoplasmic signals. (The section in Mathews, p.504-505 is probably more correct than Voet in this context.) Divalent cation concentrations can rise in mitochondria when ATP depleted and replaced by ADP, since the metal binding affinity of the diphosphate is significantly less than triphosphate.

NADH and acetyl CoA are activators of the pyruvate dehydrogenase (protein) kinase, which in turn reduces the dehydrogenase activity by phosphorylating key serine residues.

Although some texts show malic enzyme (malate dehydrogenase decarboxylating) as interconverting malate and pyruvate with the suggestion that this can replenish TCA cycle intermediates, the thermodynamics favour the reaction malate to pyruvate, and not the other way around.

malate^2- + NADP⁺ pyruvate^- + CO₂ + NADPH

= -2.1 kJ/mol, which is modest, but the CO₂ release makes a big negative contribution to .

The glyoxylate cycle

Bacteria and some species of higher plants express the enzymes of the glyoxylate cycle or glyoxylate shunt. This allows the conversion of acetyl CoA to result in net increase in malate or oxaloacetate, which is not possible with the TCA cycle alone. Two acetyl CoA are input per cycle with no loss of CO2, making possible net synthesis of a 4-carbon product. The two additional enzymes of the glyoxylate cycle are isocitrate lyase and malate synthase.

Isocitrate lyase splits isocitrate into glyoxylate and succinate. The reaction is not particularly favourable, so the concentration of glyoxylate product will be low. This bypasses the oxidative CO2 releasing steps of the normal TCA cycle.

The malate synthase reaction closely parallels the citrate synthase reaction, incorporating a second acetyl CoA to produce another malate independently of the succinate produced by isocitrate lyase. The reaction is energetically very favourable, and proceeds even with low concentrations of glyoxylate.

For each oxaloacetate consumed by citrate synthase, two malates can be produced. One goes on to replace the oxaloacetate originally used, and the other represents a net gain that can be made available for gluconeogenesis, or simply to boost the rate of citrate synthesis.

Glyoxysomes

Some higher plants such as soybeans store metabolic reserves in seeds as oils. On germination, oils are used as a carbon source for growth as well as for energy metabolism. The glyoxylate cycle allows for gluconeogenesis from fatty acids, a process that's impossible in animals.

The enzymes of the TCA cycle and the glyoxylate cycle are physically segragated, and glyoxylate cycle enzymes are localized in a specialized organelle called the glyoxysome. The glyoxysome lacks means for reoxidizing NADH, so has none of the dehydrogenases.

Glyoxysomes import fatty acids and aspartate. Fatty acids provide the source of acetyl CoA. Aspartate transaminase (aspartate aminotransferase) converts aspartate into oxaloacetate, allowing for incorporation of acetyl CoA into citrate via citrate synthase. Glyoxysomal aconitase is present, but isocitrate lyase is found in the glyoxysome in place of isocitrate dehydrogenase. The succinate produced by the lyase is exported back to the mitochondria since there is no glyoxysomal succinate dehydrogenase. Mitochondria oxidize succinate to oxaloacetate, and transminase converts it back to aspartate, to maintain the cycle. Meanwhile, the glyoxysomes incorporate a second acetyl CoA to make malate, which is exported to the cytoplasm for gluconeogenesis.

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