Chapter Summary, Study Questions

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Chapter: Biochemistry : Tricarboxylic Acid Cycle and Pyruvate Dehydrogenase Complex

Pyruvate is oxidatively decarboxylated by pyruvate dehydrogenase (PDH) complex, producing acetyl coenzyme A (CoA), which is the major fuel for the tricarboxylic acid cycle.


CHAPTER SUMMARY

Pyruvate is oxidatively decarboxylated by pyruvate dehydrogenase (PDH) complex, producing acetyl coenzyme A (CoA), which is the major fuel for the tricarboxylic acid cycle ([TCA cycle] Figure 9.9). This multienzyme complex requires five coenzymes: thiamine pyrophosphate, lipoic acid, flavin adenine dinucleotide (FAD), nicotinamide adenine dinucleotide (NAD+), and CoA. PDH complex is regulated by covalent modification of E1 (pyruvate decarboxylase) b y PDH kinase and PDH phosphatase: phosphorylation inhibits E1. PDH kinase is allosterically activated by ATP, acetyl CoA, and NADH and inhibited by pyruvate. The phosphatase is activated by Ca2+. PDH complex deficiency is the most common biochemical cause of congenital lactic acidosis. The central nervous system is particularly affected in this X-linked dominant disorder. Arsenic poisoning causes inactivation of the PDH complex by binding to lipoic acid. Citrate is synthesized from oxaloacetate and acetyl CoA by citrate synthase. This enzyme is subject to product inhibition by citrate. Citrate is isomerized to isocitrate by aconitase (aconitate hydratase) . Isocitrate is oxidatively decarboxylated by isocitrate dehydrogenase to α-ketoglutarate, producing CO2 and NADH. The enzyme is inhibited by ATP and NADH, and activated by ADP and Ca2+. α-Ketoglutarate is oxidatively decarboxylated to succinyl CoA by the α-ketoglutarate dehydrogenase complex, producing CO2 and NADH. The enzyme is very similar to the PDH complex and uses the same coenzymes. α-Ketoglutarate dehydrogenase complex is activated by Ca+2 and inhibited by NADH and succinyl CoA but is not covalently regulated. Succinyl CoA is cleaved by succinate thiokinase (also called succinyl CoA synthetase), producing succinate and GTP. This is an example of substrate-level phosphorylation. Succinate is oxidized to fumarate by succinate dehydrogenase, producing FADH2.

Fumarate is hydrated to malate by fumarase (fumarate hydratase), and malate is oxidized to oxaloacetate by malate dehydrogenase, producing NADH. Three NADH, one FADH2, and one GTP (whose terminal phosphate can be transferred to ADP by nucleoside diphosphate kinase, producing ATP) are produced by one round of the TCA cycle. The generation of acetyl CoA by the oxidation of pyruvate via the PDH complex also produces an NADH. Oxidation of the NADH and FADH2 by the electron transport chain yields 14 ATP. An additional ATP (GTP) comes from substrate level phosphorylation in the TCA cycle. Therefore, a total of 15 ATP are produced from the complete mitochondrial oxidation of pyruvate to CO2.


Figure 9.9 Key concept map for the tricarboxylic acid (TCA) cycle. CoA = coenzyme A; NAD(H) = nicotinamide adenine dinucleotide; FAD(H2) = flavin adenine dinucleotide; GDP = guanosine diphosphate; GTP = guanosine triphosphate; ADP = adenosine diphosphate; Pi = inorganic phosphate.

 

Study Questions

Choose the ONE best answer.

 

9.1 The conversion of pyruvate to acetyl coenzyme A and CO2:

A. involves the participation of lipoic acid.

B. is activated when pyruvate decarboxylase of the pyruvate dehydrogenase (PDH) complex is phosphorylated by PDH kinase in the presence of ATP.

C. is reversible.

D. occurs in the cytosol.

E. requires the coenzyme biotin.

Correct answer = A. Lipoic acid is an intermediate acceptor of the acetyl group formed in the reaction. Pyruvate dehydrogenase complex catalyzes an irreversible reaction that is inhibited when the decarboxylase component is phosphorylated. The enzyme complex is located in the mitochondrial matrix. Biotin is utilized by carboxylases, not decarboxylases.

 

9.2 Which one of the following conditions decreases the oxidation of acetyl coenzyme A by the citric acid cycle?

A. A high availability of calcium

B. A high acetyl CoA/CoA ratio

C. A low ATP/ADP ratio

D. A low NAD+/NADH ratio

Correct answer = D. A low NAD+/NADH ratio limits the rates of the NAD+-requiring dehydrogenases. High availability of calcium and substrate (acetyl CoA), and a low ATP/ADP ratio stimulates the cycle.

 

9.3 The following is the sum of three steps in the citric acid cycle.

A+B+FAD+H2O → C+FADH2+NADH

Choose the lettered answer that corresponds to the missing “A,” “B,” and “C” in the equation.


Correct answer = B. Succinate + NAD+ + FAD + H2O → oxaloacetate + NADH + FADH2

 

9.4 A 1-month-old male shows neurologic problems and lactic acidosis. Enzyme assay for pyruvate dehydrogenase (PDH) complex activity on extracts of cultured skin fibroblasts showed 5% of normal activity with a low concentration of thiamine pyrophosphate (TPP), but 80% of normal activity when the assay contained a thousand-fold higher concentration of TPP. Which one of the following statements concerning this patient is correct?

A. Administration of thiamine is expected to reduce his serum lactate level and improve his clinical symptoms.

B. A high carbohydrate diet would be expected to be beneficial for this patient.

C. Citrate production from aerobic glycolysis is expected to be increased.

D. PDH kinase, a regulatory enzyme of the PDH complex, is expected to be active.

Correct answer = A. The patient appears to have a thiamine-responsive pyruvate dehydrogenase (PDH) complex deficiency. The pyruvate decarboxylase (E1) component of the PDH complex fails to bind thiamine pyrophosphate at low concentration, but shows significant activity at a high concentration of the coenzyme. This mutation, which affects the Km of the enzyme for the coenzyme, is present in some, but not all, cases of PDH complex deficiency. Because the PDH complex is an integral part of carbohydrate metabolism, a diet low in carbohydrates would be expected to blunt the effects of the enzyme deficiency. Aerobic glycolysis generates pyruvate, the substrate of the PDH complex. Decreased activity of the complex decreases production of acetyl coenzyme A, a substrate for citrate synthase. PDH kinase is allosterically inhibited by pyruvate and, therefore, is inactive.

 

9.5 Which coenzyme-cosubstrate is used by the dehydrogenases of both glycolysis and the tricarboxylic acid cycle?

Oxidized nicotinamide adenine dinucleotide (NAD+) is used by glyceraldehyde 3-phosphate dehydrogenase of glycolysis and by isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, and malate dehydrogenase of the tricarboxylic acid cycle.


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