Active site, lipoamide dehydrogenase

Achromobacter fischeri, nitrite reductase, physical properties, 277-279 Active site, lipoamide dehydrogenase, 105 Acyl hydrazides, catalase and, 379 Acyltransferase activity, glyceraldehyde-... 

Figure 2. Mechanism of PDH. The three different subunits of the PDH complex in the mitochondrial matrix (E, pyruvate decarboxylase E2, dihydrolipoamide acyltrans-ferase Ej, dihydrolipoamide dehydrogenase) catalyze the oxidative decarboxylation of pyruvate to acetyl-CoA and CO2. E, decarboxylates pyruvate and transfers the acetyl-group to lipoamide. Lipoamide is linked to the group of a lysine residue to E2 to form a flexible chain which rotates between the active sites of E, E2, and E3. E2 then transfers the acetyl-group from lipoamide to CoASH leaving the lipoamide in the reduced form. This in turn is oxidized by E3, which is an NAD-dependent (low potential) flavoprotein, completing the catalytic cycle. PDH activity is controlled in two ways by product inhibition by NADH and acetyl-CoA formed from pyruvate (or by P-oxidation), and by inactivation by phosphorylation of Ej by a specific ATP-de-pendent protein kinase associated with the complex, or activation by dephosphorylation by a specific phosphoprotein phosphatase. The phosphatase is activated by increases in the concentration of Ca in the matrix. The combination of insulin with its cell surface receptor activates PDH by activating the phosphatase by an unknown mechanism.

Glutathione reductase, thioredoxin reductase, and lipoamide dehydrogenase are members of a group of flavoproteins that contain an active site disulfide as well as FAD. They catalyze the NAD(P)H dependent reduction of a disulfide... 

Figure 17.9 Reactions of the pyruvate dehydrogenase complex. At the top (center), the enzyme (represented by a yellow, a green, and two red spheres) is unmodified and ready for a catalytic cycle. (1) Pyruvate is decarboxylated to form hydroxyethyl-TPR (2) The lipoamide arm of Ej moves into the active site of E], (3) Ei catalyzes the transfer of the two-carbon group to the lipoamide group to form the acetyl-lipoamide complex.

Fox, B.S., and C.T. Walsh. 1983. Mercuric reductase Homology to glutathione reductase and lipoamide dehydrogenase. lodoacetamide alkylation and sequence of the active site peptide. Biochemistry 22(17) 4082-4088. 

Scheme 6 (a)Domain structure of the E2 component of the 2-oxoacid dehydrogenase complexes, (b) Schematic layout of domains in the 1-lipolyl E2ec. The lipoyl domain (LD) with the appended lipoamide visits the active sites of all three components of PDHc-ec. 

As for reduction processes, C02 free radicals were shown to react specifically with disulfide bonds (122). They were extensively used to study the redox properties of disulfide bonds, thiyl and disulfide free radicals in proteins. This is discussed in paragraph 5. However, they do react with thiol functions also (37). For proteins containing a prosthetic group, the reduction concerns also oxidized valencies of metals and flavins. Flavin adenine dinucleotide (FAD) or Flavin Mononucleotide (FMN). The proportion of reduced disulfide/reduced prosthetic group varies considerably with the protein. For instance, lipoamide dehydrogenase contains one disulfide bond close to a flavin (FAD). Free radicals can reduce only the flavin, although both are in the active site (123). In chicken egg white riboflavin binding protein, competitive formation of both disulfide and semireduced flavin is observed (124). 

Fig. 11-15 The pyruvate dehydrogenase reaction takes piace via three enzymes in a complex Pyruvate is decarboxyiated by the pyruvate decarboxylase) component of the enzyme complex a key cofactor is thiamine pyrophosphate (TPP) that transfers the hydroxyethyl moiety to one of the sulfur atoms on oxidized lipoamide that is covalently bound to Ej(c//hyc/ro//poy/ transacetylase). When this transfer takes place, the 2-carbon moiety is oxidized to an acetyl moiety and then the acetyl moiety is transferred to CoA to yield acetyl-CoA which is then released from the active site of Ej. The reduced lipoamide moiety is recycled back to its oxidized form by donating hydrogen atoms to FAD in E3 lipoamide dehydrogenase), and the reaction cycle begins over again.

Adamson SR, Robinson JA, Stevenson KJ (1984) Inhibition of pymvate dehydrogenase multienzyme complex from Escherichia coli with a radiolabeled bifunctional arsenoxide evidence for essential histidine residue at the active site of lipoamide dehydrogenase. Biochemistry 23 1269-1274... 

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