Acetyl lipoic acid

El + E2 hydroxyethyl-TPP -1- lipoic acid acetyl-lipoic acid -hTPP... 

E2 acetyl-lipoic acid -1- Coenzyme A acetyl-CoA + E2 lipoic aCidreduced... 

The formation of acetyl lipoic acid-enzyme complex by internal oxido-reduction and rearrangement (see equation 3 on p. 36) is a process which is similar in principle to the enolase reaction. 

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 first step of this reaction, decarboxylation of pyruvate and transfer of the acetyl group to lipoic acid, depends on accumulation of negative charge on the carbonyl carbon of pyruvate. This is facilitated by the quaternary nitrogen on the thiazolium group of thiamine pyrophosphate. As shown in (c), this cationic... 

Hydroxyetliyl group is transferred to lipoic acid and oxidized to form acetyl dihydro lipoamide... 

The reaction of hydroxyethyl-TPP with the oxidized form of lipoic acid yields the energy-rich thiol ester of reduced lipoic acid and results in oxidation of the hydroxyl-carbon of the two-carbon substrate unit (c). This is followed by nucleophilic attack by coenzyme A on the carbonyl-carbon (a characteristic feature of CoA chemistry). The result is transfer of the acetyl group from lipoic acid to CoA. The subsequent oxidation of lipoic acid is catalyzed by the FAD-dependent dihydrolipoyl dehydrogenase and NAD is reduced. 

The PDHC catalyzes the irreversible conversion of pyruvate to acetyl-CoA (Fig. 42-3) and is dependent on thiamine and lipoic acid as cofactors (see Ch. 35). The complex has five enzymes three subserving a catalytic function and two subserving a regulatory role. The catalytic components include PDH, El dihydrolipoyl trans-acetylase, E2 and dihydrolipoyl dehydrogenase, E3. The two regulatory enzymes include PDH-specific kinase and phospho-PDH-specific phosphatase. The multienzyme complex contains nine protein subunits, including... 

ATP and magnesium were required for the activation of acetate. Acetylations were inhibited by mercuric chloride suggesting an SH group was involved in the reaction either on the enzyme or, like lipoic acid, as a cofactor. Experiments from Lipmann s laboratory then demonstrated that a relatively heat-stable coenzyme was needed—a coenzyme for acetylation—coenzyme A (1945). The thiol-dependence appeared to be associated with the coenzyme. There was also a strong correlation between active coenzyme preparations and the presence in them of pantothenic acid—a widely distributed molecule which was a growth factor for some microorganisms and which, by 1942-1943, had been shown to be required for the oxidation of pyruvate. 

Direct evidence of the reaction of PAN with sulfhydryl compounds has since been obtained (PAN at 115 ppm for 1-10 min). - In the reaction with glutathione, the major products are oxidized glutathione (disulfide) and 5-acetylglutathione. Other sulfhydryl compounds (e.g., coenzyme A, lipoic acid, and cysteine) yield only oxidation products, with no evidence of 5-acetylation. However, acetylation reactions have been observed with alcohols and amines. Sulfur compounds other than thiols can undergo oxidation by PAN methionine is converted to methionine sulfoxide, and oxidized lipoic acid (disulfide) is converted to sulfoxide. 

Now this reaction is effectively a repeat of the pyruvate acetyl-CoA oxidative decarboxylation we saw at the beginning of the Krebs cycle. It similarly requires thiamine diphosphate, lipoic acid, coenzyme A and NAD+. A further feature in common with that reaction is that 2-oxoglutarate dehydrogenase is also an enzyme complex comprised of three separate enzyme activities. 2-Oxoglutarate is thus transformed into succinyl-CoA, with the loss of... 

Dihydrolipoyl transacetylase transfers the acetyl CoA to its lipoic acid... 

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.

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.

FIGURE 16-4 Lipoic acid (lipoate) in amide linkage with a Lys residue. The lipoyllysyl moiety is the prosthetic group of dihydrolipoyl transacetylase (E2 of the PDH complex). The lipoyl group occurs in oxidized (disulfide) and reduced (dithiol) forms and acts as a carrier of both hydrogen and an acetyl (or other acyl) group. 

The acetyl group, bound as a thioester to the side chain of lipoic acid, is transferred to CoA. I... 

The conversion of pyruvate to acetyl CoA and C02 A. is reversible. B. involves the participation of lipoic acid. C. is activated when pyruvate dehydrogenase complex is phosphorylated by a protein kinase in the pres ence of ATP. D. occurs in the cytosol. E. depends on the coenzyme biotin. Correct answer = B. Lipoic acid is an intermedi ate acceptor of the acetyl group formed in the reaction. Pyruvate dehydrogenase complex cat alyzes an irreversible reaction that is inhibited when the enzyme is phosphorylated. The enzyme is located in the mitochondrial matrix. 

Reactions of the TCA cycle Enzyme that oxidatively decarboxylates pyruvate, its coenzymes, activators, and inhibitors REACTIONS OF THE TRICARBOXYLIC ACID CYCLE (p. 107) Pyruvate is oxidatively decarboxylated by pyruvate dehydrogenase complex producing acetyl CoA, which is the major fuel for the tricarboxylic acid cycle (TCA cycle). The irreversible set of reactions catalyzed by this enzyme complex requires five coenzymes thiamine pyrophosphate, lipoic acid, coenzyme A (which contains the vitamin pantothenic acid), FAD, and NAD. The reaction is activated by NAD, coenzyme A, and pyruvate, and inhibited by ATP, acetyl CoA, and NADH. 

Pyruvate oxidase. The soluble flavoprotein pyruvate oxidase, which was discussed briefly in Chapter 14 (Fig. 14-2, Eq. 14-22), acts together with a membrane-bound electron transport system to convert pyruvate to acetyl phosphate and C02.319 Thiamin diphosphate is needed by this enzyme but lipoic acid is not. The flavin probably dehydrogenates the thiamin-bound intermediate to 2-acetylthiamin as shown in Eq. 15-34. The electron acceptor is the bound FAD and the reaction may occur in two steps as shown with a thiamin diphosphate radical intermediate.3193 Reaction with inorganic phosphate generates the energy storage metabolite acetyl phosphate. 

Tire enzyme does not require lipoic acid. It seems likely that a thiamin-bound enamine is oxidized by an iron-sulfide center in the oxidoreductase to 2-acetyl-thiamin which then reacts with CoA. A free radical intermediate has been detected318 321 and the proposed sequence for oxidation of the enamine intermediate is that in Eq. 15-34 but with the Fe-S center as the electron acceptor. Like pyruvate oxidase, this enzyme transfers the acetyl group from acetylthiamin to coenzyme A. Cleavage of the resulting acetyl-CoA is used to generate ATR An indolepyruvate ferredoxin oxidoreductase has similar properties 322... 

Pyruvate formate-lyase reaction. Anaerobic cleavage of pyruvate to acetyl-CoA and formate (Eq. 15-37) is essential to the energy economy of many cells, including those of E. coli. No external oxidant is needed, and the reaction does not require lipoic acid. 

The conversion of pyruvate to acetyl-CoA. The reactions are catalyzed by the enzymes of the pyruvate dehydrogenase complex. This complex has three enzymes pyruvate decarboxylase, dihydrolipoyl transacetylase, and dihydrolipoyl dehydrogenase. In addition, five coenzymes are required thiamine pyrophosphate, lipoic acid, CoASH, FAD, and NAD+. Lipoic acid is covalently attached to... 

Answer TPP thiazolium ring adds to a carbon of pyruvate, then stabilizes the resulting car-banion by acting as an electron sink. Lipoic acid oxidizes pyruvate to level of acetate (acetyl-CoA), and activates acetate as a thioester. CoA-SH activates acetate as thioester. FAD oxidizes lipoic acid. NAD+ oxidizes FAD. (See Fig. 16-6.)... 

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