13.2.      Anaplerotic reactions

Anaplerotic means replenishing or “filling-up” reactions. Many intermediates of the citric acid cycle are used for the synthesis of other substances. Anaplerosis is defined as any reaction that can restore the concentration of a crucial but depleted intermediate. Several biosynthetic pathways utilize citric acid cycle intermediates as starting materials. The citric acid cycle is therefore amphibolic (both anabolic and catabolic).

The TCA cycle in the liver is often called an “open cycle” because there is such a high efflux of intermediates. After a high carbohydrate meal, citrate efflux and cleavage to acetyl CoA provides acetyl units for cytosolic fatty acid synthesis. During fasting, gluconeogenic precursors are converted to malate, which leaves the mitochondria for cytosolic gluconeogenesis.

Some important anaplerotic reactions of the citric acid cycle intermediate are used for the synthesis of following substrate.

Pyruvate carboxylase is a major anaplerotic enzyme. The citric acid cycle operates when oxaloacetate is available. In animal tissues especially, liver and kidney, oxaloacetate is formed by the carboxylation of pyruvate by pyruvate carboxylase in  mitochondria. It is a readily reversible reaction. This enzyme is absent in mammals.

Pyruvate carboxylase is homotetramer. Each subunit of Pyruvate carboxylase contains an active site that has tightly-bound Mn+2 (or Mg+2) and covalently-bound biotin and also an allosteric site, which binds acetyl-CoA. The lysine residue of active site is covalently attached with biotin. Pyruvate carboxylase also requires a monovalent cation (K+) apart from a divalent cation (Mg+2 or Mn+2).

Pyruvate carboxylase catalyzes the addition of CO2 to pyruvate to form oxaloacetate.

The  biotin of pyruvate carboxylase enzymes forms a covalent intermediate with CO2 in a reaction requiring ATP and Mg+2. The activated CO2 is then transferred to pyruvate to form the carboxyl group of Oxaloacetate.

Pyruvate carboxylase is activated by acetyl CoA and inhibited by high concentrations of many acyl CoA derivatives. As the concentration of oxaloacetate is depleted through the efflux of TCA cycle intermediates, the rate of the citrate synthase reaction decreases and acetyl CoA concentration rises. The acetyl CoA then activates pyruvate carboxylase to synthesize more oxaloacetate.

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