The Pyruvate Dehydrogenase Complex

Acetyl-CoA is produced from fatty acids, proteins, and carbohydrates and is a central and major compound in intermediary metabolism. The mechanism of its formation from the degradation of fatty acids and proteins is discussed in Chaps. 13 and 15, respectively here, the means whereby carbohydrates form this most important molecule will be presented. The glycolytic pathway can yield pyruvate from all degradable sugars, and this can be converted to acetyl-CoA. Pyruvate enters the mitochondrial matrix and is the substrate for the multienzyme complex pyruvate dehydrogenase. 

Question What is the overall reaction catalyzed by the pyruvate dehydrogenase complex  

Control of the activity of the pyruvate dehydrogenase complex is exerted by the phosphorylation of pyruvate decarboxylase (E[), which renders it inactive. This process is catalyzed by pyruvate dehydrogenase kinase, which is always tightly bound to E[. The kinase is activated by high-energy conditions, and it requires ATP to accomplish the phosphorylation step. Another enzyme, phosphoprotein phosphatase, is weakly bound to E, and reactivates the system by removing the inhibitory phosphate group (Fig. 12-8). 

The pyruvate dehydrogenase complex is not directly a part of the reactions that constitute the citric acid cycle. It is the link between glycolysis and the citric acid cycle, and its activity is controlled by the energy status of the mitochondria. 

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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 ail 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. 

The conversion occurs through a multistep sequence of reactions catalyzed by a complex of enzymes and cofactors called the pyruvate dehydrogenase complex. The process occurs in three stages, each catalyzed by one of the enzymes in the complex, as outlined in Figure 29.11 on page 1152. Acetyl CoA, the ultimate product, then acts as fuel for the final stage of catabolism, the citric acid cycle. All the steps have laboratory analogies. 

Several enzymes of the intermediary metabolism require thiaminpyrophosphate (TPP, Fig. 1) as coenzyme, e.g., enzymes of the pyruvate dehydrogenase complex, a-ketoglutarate dehydrogenase complex, or pentose phosphate pathway. 

Many metabolic fuels are oxidized in the mitochondrial matrix. Pyruvate is oxidatively decarboxylated to acetyl-CoA by the pyruvate dehydrogenase complex (PDH)... 

The pyruvate dehydrogenase complex consists of a number of polypeptide chains of each of the three component enzymes, all organized in a regular spatial configuration. Movement of the individual enzymes appears to be restricted, and the metabofic intermediates do not dissociate freely but remain bound to the enzymes. Such a complex of enzymes, in which the sub-... 

Figure 17-5. Oxidative decarboxylation of pyruvate by the pyruvate dehydrogenase complex. Lipoic acid is joined by an amide link to a lysine residue of the transacetylase component of the enzyme complex. It forms a long flexible arm, allowing the lipoic acid prosthetic group to rotate sequentially between the active sites of each of the enzymes of the complex. (NAD nicotinamide adenine dinucleotide FAD, flavin adenine dinucleotide TDP, thiamin diphosphate.)...

Metabolic Transitions and the Role of the Pyruvate Dehydrogenase Complex During Development of Ascaris suum... 

Mitochondria from body wall muscle and probably the pharynx lack a functional TCA cycle and their novel anaerobic pathways rely on reduced organic acids as terminal electron acceptors, instead of oxygen (Saz, 1971 Ma et al, 1993 Duran et al, 1998). Malate and pyruvate are oxidized intramitochondrially by malic enzyme and the pyruvate dehydrogenase complex, respectively, and excess reducing power in the form of NADH drives Complex II and [3-oxidation in the direction opposite to that observed in aerobic organelles (Kita, 1992 Duran et al, 1993 Ma et al,... 

This chapter focuses on the developmental regulation of the pyruvate dehydrogenase complex (PDC). The PDC plays diverse and pivotal roles in the entry of glycolytically generated carbon into the TCA cycle in aerobic stages and the metabolism of mitochondrially generated pyruvate in anaerobic stages (Fig. 14.1). 

Fig. 14.2. Regulation of the pyruvate dehydrogenase complex (PDC) from adult A suum muscle. PDC, pyruvate dehydrogenase complex E1, pyruvate dehydrogenase subunit of the PDC PDK, pyruvate dehydrogenase kinase PDP, pyruvate dehydrogenase phosphatase.

Fig. 14.4. Expression of subunits of the pyruvate dehydrogenase complex during the development of A suum. Homogenates of different A suum larval stages and adult tissues were immunoblotted with polyclonal antisera prepared against individual subunits of the A suum PDC isolated from adult muscle, as described in detail in Klingbeil etal. (1996). UE, unembryonated egg M, adult body wall muscle p45, E3-binding protein (E3BP).

In summary, it is clear that A. suum undergoes a number of metabolic transitions during development and, in the case of the pyruvate dehydrogenase complex at least, is constantly fine-tuning the subunit-specific expression and function of the PDC during the course of development. 

Klingbeil, M.M., Walker, D.J., Arnette, R., Sidawy, E., Hayton, K, Komuniecki, P.R. and Komuniecki, R. (1996) Identification of a novel dihydrolipoyl dehydrogenase-binding protein in the pyruvate dehydrogenase complex of the anaerobic parasitic nematode, Ascaris suum. Journal of Biological Chemistry 271, 5451-5457. 

Ravindran, S., Radke, G.A., Guest, J.R. and Roche, T.E. (1996) Lipoyl domain-based mechanism for the integrated feedback control of the pyruvate dehydrogenase complex by enhancement of pyruvate dehydrogenase kinase activity. Journal of Biological Chemistry 271,653-562. 

Thissen, J., Desai, S., McCartney, P. and Komuniecki, R. (1986) Improved purification of the pyruvate dehydrogenase complex from Ascaris suum body wall muscle and characterization of PDHa kinase activity. Molecular and Biochemical Parasitology 21, 129-138. 

Fig. 3. Generation of propionyl-CoA from the isoleucine biosynthetic pathway. The intermediate 2-ketobutyrate can be decarboxylated by either the 2-oxoacid dehydrogenase complex or at low efficiency by the pyruvate dehydrogenase complex. Inhibition of the threonine deaminase by isoleucine and of the acetolactate synthase by herbicides are indicated with dashed arrows...

Lipoic acid (the other names are a-lipoic acid or thioctic acid) (Figure 29.9) is a natural compound, which presents in most kinds of cells. Lipoic acid (LA) is contained in many food products, in particular in meat, but it is also synthesized in human organism from fatty acids. Earlier, it has been shown that in humans lipoic acid functions as a component of the pyruvate dehydrogenase complex. However, later on, attention has been drawn to the possible antioxidant activity of the reduced form of lipoic acid, dihydrolipoic acid (DHLA) (Figure 29.9). 

The pyruvate dehydrogenase complex plays a key role in regulating oxidation of glucose 543... 

Similarly, the pyruvate dehydrogenase complex (PDC) can be activated directly by electrogenerated methyl viologen radical cations (MV +) as mediator. Thus, the naturally PDC-catalyzed oxidative decarboxylation of pyruvic acid in the... 

Thiamine pyrophosphate is a coenzyme for several enzymes involved in carbohydrate metabolism. These enzymes either catalyze the decarboxylation of oi-keto acids or the rearrangement of the carbon skeletons of certain sugars. A particularly important example is provided by the conversion of pyruvic acid, an oi-keto acid, to acetic acid. The pyruvate dehydrogenase complex catalyzes this reaction. This is the key reaction that links the degradation of sugars to the citric acid cycle and fatty acid synthesis (chapters 16 and 18) ... 

We know that anaerobic glycolysis of glncose yields pyruvate and/or lactate, interconvertable metabolites. Pyruvate is converted into acetyl-SCoAin the following reaction, catalyzed by the pyruvate dehydrogenase complex ... 

To begin with, let us return to the aerobic catabolism of simple sugars such as glucose to yield two molecules of pyruvate -I- two molecules of ATP - - two molecules of NADH. We noted just above that coupling the oxidation of the two molecules of NADH to the electron transport chain yields an additional six molecules of ATP, three for each molecule of NADH, for a total of eight. Now let s ask what happens when we further metabolize the two molecules of pyruvate via the pyruvate dehydrogenase complex and the citric acid cycle. 

The intermediary metabolism has multienzyme complexes which, in a complex reaction, catalyze the oxidative decarboxylation of 2-oxoacids and the transfer to coenzyme A of the acyl residue produced. NAD" acts as the electron acceptor. In addition, thiamine diphosphate, lipoamide, and FAD are also involved in the reaction. The oxoacid dehydrogenases include a) the pyruvate dehydrogenase complex (PDH, pyruvate acetyl CoA), b) the 2-oxoglutarate dehydrogenase complex of the tricarboxylic acid cycle (ODH, 2-oxoglutarate succinyl CoA), and c) the branched chain dehydrogenase complex, which is involved in the catabolism of valine, leucine, and isoleucine (see p. 414). 

Pyruvate dehydrogenase (lipoamide) [EC 1.2.4.1], which requires thiamin pyrophosphate, catalyzes the reaction of pyruvate with lipoamide to produce 5-acetyldihydroli-poamide and carbon dioxide. It is a component of the pyruvate dehydrogenase complex (which also includes dihydrolipoamide dehydrogenase [EC 1.8.1.4] and dihy-drolipoamide acetyltransferase [EC 2.3.1.12]). Pyruvate dehydrogenase (cytochrome) [EC 1.2.2.2] catalyzes the... 

This enzyme [EC 3.1.3.43] catalyzes the hydrolysis of the phosphorylated form of pyruvate dehydrogenase (lipoamide) to generate the dephosphorylated dehydrogenase and orthophosphate. The enzyme is associated with the pyruvate dehydrogenase complex. 

Figure 7-1. Conversion of pyruvate to acetyl CoA by the pyruvate dehydrogenase complex. The three enzymes, pyruvate dehydrogenase, dihydrolipoyl transacetylase, and dihydrolipoyl dehydrogenase, exist in a complex associated with the mitochondrial matrix. Each enzyme requires at least one coenzyme that participates in the reaction. TPP, thiamine pyrophosphate Lip, lipoic acid CoA, coenzyme A.

High acetyl CoA levels from 3-oxidation of fatty acids in liver cells inhibit the pyruvate dehydrogenase complex and activate pyruvate carboxylase, which increases oxaloacetate synthesis. 

Figure 5.6 Self-organization in oligomeric proteins. (A) The transacetylase core of the pyruvate dehydrogenase complex. The core consists of 24 identical chains (12 can be seen in this view). (B) The aspartate transcarbamoylase, formed by six catalytic (lighter subunits) and six regulatory chains (darker subunits) (Bi) view showing the threefold symmetry (B2) a perpendicular view. (C) The helical assembly of several identical globular subunits in F-actin polymer. The helix repeats after 13 subunits. (All adapted from Stryer, 1975.)...

The pyruvate dehydrogenase complex from Escherichia coli is considerably more complex than tryptophan synthetase. It has a molecular weight of approximately 4.6 millon and contains three enzymes pyruvate dehydrogenase (Et), dihydrolipoyl transacetylase (E2), and dihydrolipoyl dehydrogenase (E3).82 The overall reaction catalyzed by the complex is... 

Fig. 9. A schematic drawing of a possible mechanism for the reaction catalyzed by the pyruvate dehydrogenase complex. The three enzymes Elf E2, and E3 are located so that lipoic acid covalently linked to E2 can rotate between the active sites containing thiamine pyrophosphate (TPP) and pyruvate (Pyr) on Elt CoA on E2, and FAD on E3. Acetyl-CoA and GTP are allosteric effectors of E, and NAD+ is an inhibitor of the overall reaction.

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