Lipoyl transacetylase

Oxoglutarate undergoes oxidative decarboxylation to succinyl-CoA, via multi-enzyme reaction similar to the reaction pattern of pyruvate. The multi-enzyme complex (mw about 2 x 10 ) is an octamer of an elementary unit containing each of the three contributing enzyme proteins oxoglutarate decarboxylase, dihydro-lipoyl transacetylase, and dihydrolipoyl dehydrogenase. The overall reaction involves thiamine pyrophosphate, lipoic acid, CoASH and NAD succinyl-CoA is the end product ... 

The aim of the present experiment was to study the PDHc activity in the livers and brains of developing rats born to females kept on a fat-free diet from 10 days after mating. The question arises in the mechanism of action of the supplemental linoleic acid given to the progeny after weaning on PDHc activity. Any effect could obviously be either direct, due to variation in the PDHc environment, or could be secondary and due to the stimulation of lipoic acid synthesis needed for further activation of the lipoyl transacetylase and lipoamide dehydrogenase. ... 

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 PDH complex is composed of multiple copies of three enzymes pyruvate dehydrogenase, Ei (with its bound cofactor TPP) dihydrolipoyl transacetylase, E2 (with its covalently bound lipoyl group) and dihydrolipoyl dehydrogenase, E3 (with its cofactors FAD and NAD). 

In E. coli, the complex has a mass of about 4 x 106 Da and consists of 60 polypeptide chains. In the center of the complex there is a core of eight trimers of E2 arranged in cubic symmetry (Fig. 5-10). Dimers of E3 are bound to the six faces of the cube. Finally, pairs of Et bind to each edge of the cube encircling the dehydrogenase dimers. The central position of the transacetylase (E2) allows the flexible lipoyl arms to transfer reactants from Ej to E3 or to CoA. 

The lipoamide of dihydrolipoyl transacetylase constitutes a long arm which may now move the acetyl group from the active site of pyruvate DH to its own active site where the lipoamide is exchanged for Coenzyme A-SH. (On the mammalian enzyme the 60 subunits of the transacetylase seem to form a pool of lipoyl groups among which the acetyl groups are freely exchanged). 

A detailed mechanism of the first two reactions is shown in Figure 13-8. In the third step, catalyzed by dihydrolipoyl transacetylase, the acetyl group attached to E2 is transferred to CoASH, with formation of the dithiol form of the lipoyl group and acetyl-CoA. 

In the second step, also catalyzed by pyruvate dehydrogenase, the hydroxyethyl group is transferred to the oxidized form of the lipoyl-lysyl prosthetic group of dihydrolipoyl transacetylase (E2) ... 

Schematic representation of the relationship between the three enzymes of the pyruvate dehydrogenase complex. The lipoyl-lysyl moiety of the transacetylase delivers the acetyl group to CoA and the reducing equivalents to FAD. TPP = Thiamine pyrophosphate.

The acyl-generation reaction, Eq. (8), has been visualized as a reductive acylation of protein-bound lipoic acid. As will be seen below, this reaction is now belitwod to consist of two steps an oxidation of the 2-hydroxyalkyl-thiamine pyrophcjsphatc to 2-aoylthiaminc pyrophosphate with a concomitant reduction of bound lipoic acid, and a transfer of (he acyl group of 2-acylthiamine pyrophosphate to the bound dihydrolipoic acid (Das el al., 19(il). An enzymatic component which contains bound lipoic acid and apparently catalyzes reactions (8) and (9) has been isolated from the E. mli pyruvate dehydrogenation complex (Koike and Reed, 1961). This component, designated lipoyl-Ea in Fig. 1, has been tentatively named lipoic reductase-transacetylase. 

Fig. 2. Mechanism of the oxidative decarboxylation of 2-oxoglutarate by the 2- oxoglutarate dehydrogenase complex EC 1.2.4.2). Enzyme, = 2-oxoglutarate decarboxylase. Enzym = lipoyl-redurtase-transacetylase (lipoyl reductase-t-transsuccinylase). Enzymej = dihydrolipoyl dehydrogenase, tpp-thiamin pyrophosphate. HSCoA = coenzyme A.

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