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Under anaerobic conditions,  pyruvate is converted into lactic acid or ethanol, depending on the organism. Under aerobic conditions, pyruvate is transported into mitochondria by a proton symporter (Figure-1). In the mitochondrial matrix, pyruvate is oxidatively decarboxylated by the pyruvate dehydrogenase complex to form acetyl CoA.

 Pyruvate dehydrogenase complex catalyzed reaction

 This irreversible reaction is the link between glycolysis and the citric acid cycle. In the preparation of the glucose derivative pyruvate for the citric acid cycle, an oxidative decarboxylation takes place and high transfer- potential electrons in the form of NADH are captured.

 Pyruvate symport

Figure-1- pyruvate symport

Components of Pyruvate dehydrogenase complex

1)  Enzymes- The pyruvate dehydrogenase complex is a large, highly integrated complex of 2 types of enzymes-

A)- Catalytic enzymes

a) Pyruvate dehydrogenase (E1)

b) Dihydrolipoyl transacetylase (E2)

c) Dihydrolipoyl dehydrogenase (E3)

B)- Regulatory Enzymes

a) PDH Kinase

b) PDH Phosphatase

The pyruvate dehydrogenase complex consists of a number of polypeptide chains of each of the three component  catalytic enzymes, and the intermediates do not dissociate, but remain bound to the enzymes.

2) Coenzymes of PDH complex

Five coenzymes: thiamine pyrophosphate (TPP), lipoic acid, CoASH, FAD and NAD+ participate in the overall reaction (Figure-2)

Pyruvate dehydrogenase is a member of a family of homologous complexes that includes the citric acid cycle enzyme α- ketoglutarate dehydrogenase, a branched-chain α-ketoacid dehydrogenase, and acetoin dehydrogenase, found in certain prokaryotes. These complexes are large, with molecular masses ranging from 4 to 10 million daltons.

Reaction catalyzed by PDH Complex

 The conversion of pyruvate into acetyl CoA consists of three steps: decarboxylation, oxidation, and transfer of the resultant acetyl group to CoA.

 Pyruvate to Acetyl co A

Reaction sequence in PDH complex catalyzed reaction

These steps must be coupled to preserve the free energy derived from the decarboxylation step to drive the formation of NADH and acetyl CoA.

Reaction steps

 i) Pyruvate is decarboxylated by the pyruvate dehydrogenase component of the enzyme complex to a hydroxyethyl derivative of the thiazole ring of enzyme-bound thiamine pyrophosphate (Figure-2)

ii) Hydroxyethyl TPP in turn reacts with oxidized lipoamide, the prosthetic group of dihydrolipoyl transacetylase, to form acetyl lipoamide. Thiamine is vitamin B1 and in deficiency, glucose metabolism is impaired, and there is significant (and potentially life-threatening) lactic and pyruvic acidosis.

iii) Acetyl lipoamide reacts with coenzyme A to form acetyl-CoA and reduced lipoamide.

iv) The reaction is completed when the reduced lipoamide is reoxidized by a flavoprotein, dihydrolipoyl dehydrogenase, containing FAD.

v) Finally, the reduced flavoprotein is oxidized by NAD+, which in turn transfers reducing equivalents to the respiratory chain. (Figure-2)

 REACTION STEPS PDH complex

Figure-2- Reactions of PDH complex.During the oxidation of pyruvate to CO2 by pyruvate dehydrogenase the electrons flow from pyruvate to the lipoamide moiety of dihydrolipoyl transacetylase then to the FAD cofactor of dihydrolipoyl dehydrogenase and finally to reduction of NAD+ to NADH. The acetyl group is linked to coenzyme A (CoASH) in a high energy thioester bond. The acetyl-CoA then enters the TCA cycle for complete oxidation to CO2 and H2O.

Regulation of PDH complex

The reactions of the PDH complex serve to interconnect the metabolic pathways of glycolysis, gluconeogenesis and fatty acid synthesis to the TCA cycle. As a consequence, the activity of the PDH complex is highly regulated by a variety of allosteric effectors and by covalent modification.

Allosteric regulation- Pyruvate dehydrogenase is inhibited by its products, acetyl-CoA and NADH.

Covalent modification- It is also regulated by phosphorylation of three serine residues on the pyruvate dehydrogenase component of the multienzyme complex.

PDH exists in two forms-

i) PDH-a form which is active and dephosphorylated form

ii) PDH -b form which is inactive and phosphorylated form

PDH kinase, causes phosphorylation resulting in decreased activity, and

PDH phosphatase causes an increase in activity by dephosphorylation of the enzyme

Regulation of PDH Kinase

Positive effectors- NADH and acetyl-CoA are powerful positive effectors on PDH kinase, the enzyme thus inactivates PDH by converting it to the phosphorylated PDH-b form (Figure-3)

The kinase is activated by increases in the [ATP]/[ADP], [acetyl-CoA]/[CoASH], and [NADH]/[NAD+] ratios. Since NADH and acetyl-CoA accumulate when the cell energy charge is high, it is not surprising that high ATP levels also up-regulate PDH kinase activity, reinforcing down-regulation of PDH activity in energy-rich cells.

Negative effector

Pyruvate is a potent negative effector on PDH kinase, with the result that when pyruvate levels rise, PDH-a (active form) will be favored even with high levels of NADH and acetyl-CoA.

Concentrations of pyruvate which maintain PDH in the active form (PDH-a) are sufficiently high so that, in energy-rich cells, the allosterically down-regulated, high Km form of PDH is nonetheless capable of converting pyruvate to acetyl-CoA. With large amounts of pyruvate in cells having high energy charge and high NADH, pyruvate carbon will be directed to the 2 main storage forms of carbon (glycogen via gluconeogenesis and fat production via fatty acid synthesis) where acetyl-CoA is the principal carbon donor.

Regulation of PDH complex 

Figure -3-Regulation of PDH complex by covalent modification. Activation of PDH kinase causes inactivation of PDH, whereas activation of  PDH phosphatase bring about activation of enzyme

Regulation of PDH phosphatase

Although the regulation of PDH-b phosphatase is not well understood, it is likely to be regulated to maximize pyruvate oxidation under energy-poor conditions and to minimize PDH activity under energy-rich conditions. It is known that Mg2+ and Ca2+ activate the enzyme

Thus, pyruvate dehydrogenase, and therefore glycolysis, is inhibited both when there is adequate ATP (and reduced coenzymes for ATP formation) available, and also when fatty acids are being oxidized. In fasting, when free fatty acid concentrations increase, there is a decrease in the proportion of the enzyme in the active form, leading to a sparing of carbohydrate. In adipose tissue, glucose provides acetyl-CoA for lipogenesis, the enzyme is activated in response to insulin and in cardiac muscle PDH activity is increased by catecholamines.

Energetics of PDH complex

Two pyruvate molecules are obtained from one glucose molecule through glycolysis. Each of the pyruvate yields one NADH, thus there are two NADH molecules to be oxidized through the electron transport chain.

 Pyruvate dehydrogenase complex catalyzed reaction

Each of NADH yields 3 ATP molecules, thus a total of 6 ATP molecules are produced at the level of PDH complex.

Clinical Significance

PDH Complex deficiency

Pyruvate dehydrogenase complex deficiency (PDCD) is a rare disorder of carbohydrate metabolism caused by a deficiency of one or more enzymes in the pyruvate dehydrogenase complex. Dysfunctions in all 3 substrate-processing enzymes, regulatory proteins and thiamine dependence of the E1 alpha enzyme, have been described; however, dysfunction of the E1 alpha enzyme subunit is most common.

The age of onset and severity of disease depends on the activity level of the PDC enzymes. Individuals with PDCD beginning prenatally or in infancy usually die in early childhood. Those who develop PDCD later in childhood may have mental retardation and other neurological symptoms and usually survive into adulthood.

Pathophysiology

The following features are characteristic of this disease-

1) Energy Deficit-A deficiency in this enzymatic complex limits the production of citrate. Because citrate is the first substrate in the citric acid cycle, the cycle cannot proceed. Alternate metabolic pathways are stimulated in an attempt to produce acetyl-CoA; however, an energy deficit remains, especially in the CNS. The magnitude of the energy deficit depends on the residual activity of the enzyme.

2) Neurological deficit- Severe enzyme deficiencies may lead to congenital brain malformation because of a lack of energy during neural development. Underlying neuropathology is not usually observed in individuals whose onset of pyruvate dehydrogenase complex deficiency is in childhood.

Clinical Manifestations

The signs of poor neurological development or degenerative lesions are poor acquisition or loss of motor milestones, poor muscle tone, new onset seizures, and periods of incoordination (ie, ataxia) abnormal eye movements, poor response to visual stimuli, mental delay, psychomotor delays and growth retardation

Laboratory Diagnosis

High blood lactate and pyruvate levels with or without lactic acidemia suggest an inborn error of metabolism at the mitochondrial level.

Treatment

Cofactor supplementation with thiamine, carnitine, and Lipoic acid is the standard of care. Ketogenic diets (with restricted carbohydrate intake) have been used to control lactic acidosis with minimal success.

Prognosis

Correction of acidosis does not reverse all the symptoms. CNS damage is common and limits recovery of normal function.

Inhibitors of PDH Complex

Arsenite and mercuric ions react with the —SH groups of lipoic acid and inhibit pyruvate dehydrogenase complex .

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