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Another common type of C—C cleavage is a-cleavage of an «-hydroxy-ketone:

Cleavage

(We see this type of cleavage in the transketolase reaction described in Chapter 23.)

Neither of these cleavage strategies is suitable for acetate. It has no ¡-carbon, and the second method would require hydroxylation—not a favorable reaction for acetate. Instead, living things have evolved the clever chemistry of condensing acetate with oxaloacetate and then carrying out a ¡-cleavage. The TCA cycle combines this cleavage reaction with oxidation to form CO2, regenerate oxaloacetate, and capture the liberated metabolic energy in NADH and ATP.

20.3 • The Bridging Step: Oxidative Decarboxylation of Pyruvate

Pyruvate produced by glycolysis is a significant source of acetyl-CoA for the TCA cycle. Because, in eukaryotic cells, glycolysis occurs in the cytoplasm, whereas the TCA cycle reactions and all subsequent steps of aerobic metabolism take place in the mitochondria, pyruvate must first enter the mitochondria to enter the TCA cycle. The oxidative decarboxylation of pyruvate to acetyl-CoA,

Pyruvate + CoA + NAD+-> acetyl-CoA + CO2 + NADH + H+

is the connecting link between glycolysis and the TCA cycle. The reaction is catalyzed by pyruvate dehydrogenase, a multienzyme complex.

The pyruvate dehydrogenase complex (PDC) is a noncovalent assembly of three different enzymes operating in concert to catalyze successive steps in the conversion of pyruvate to acetyl-CoA. The active sites of all three enzymes are not far removed from one another, and the product of the first enzyme is passed directly to the second enzyme and so on, without diffusion of substrates and products through the solution. The overall reaction (see A Deeper Look: "Reaction Mechanism of the Pyruvate Dehydrogenase Complex") involves a total of five coenzymes: thiamine pyrophosphate, coenzyme A, lipoic acid, NAD+, and FAD.

20.4 • Entry into the Cycle: The Citrate Synthase Reaction

The first reaction within the TCA cycle, the one by which carbon atoms are introduced, is the citrate synthase reaction (Figure 20.5). Here acetyl-CoA reacts with oxaloacetate in a Perkin condensation (a carbon-carbon condensation between a ketone or aldehyde and an ester). The acyl group is activated in two ways in an acyl-CoA molecule: the carbonyl carbon is activated for attack by nucleophiles, and the Ca carbon is more acidic and can be deprotonated to form a carbanion. The citrate synthase reaction depends upon the latter mode of activation. As shown in Figure 20.5, a general base on the enzyme accepts a proton from the methyl group of acetyl-CoA, producing a stabilized a-carban-ion of acetyl-CoA. This strong nucleophile attacks the a-carbonyl of oxaloacetate, yielding citryl-CoA. This part of the reaction has an equilibrium constant

20.4 • Entry into the Cycle: The Citrate Synthase Reaction

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