Enoyl CoA hydratase ECH

Enoyl-CoA hydratase (ECH commonly known as crotonase), that catalyzes the cofactor-independent hydration of conjugated enoyl-CoA esters in yS-oxidation, has been the subject of considerable debate regarding the timing of bond-making reactions and, therefore, the importance of a thioester enolate anion on the reaction coordinate. The active site contains Glu 144 and Glu 164 as the only possible acid-base catalysts. In the nonphysiological dehydration direction, the value of the pKa... 

Another enzyme that bears a close resemblance to bifunctional acid-base catalysts is enoyl-CoA hydratase (ECH). This enzyme is discussed here as an example of an enolate-stabilizing enzyme. The mechanism of ECH involves an enolate intermediate that is formed upon addition of a nucleophile, water molecule, to an a,p-unsaturated thioester. This reaction is the second step in the metabolic oxidation of fatty acids. The X-ray structures of enoyl hydratase enzymes complexed with various substrates analogs are available [29, 30]. A simplified view of the mechanism of ECH is presented in Scheme 6.14. The lifetime of the enolate intermediate is not... 

Scheme 6.14 The reaction mechanism and mode of cateilysis by enoyl-CoA hydratase (ECH)...

Further persuasive evidence in support of the expectation that the mechanism of the ECH-catalyzed reaction involves an Elcb mechanism with a stabilized thioester enolate anion intermediate is obtained from the membership of ECH in the mechanistically diverse enoyl-CoA hydratase superfamily [70]. Such superfamilies are derived from a common ancestor by divergent evolution the members of these share a partial reaction, usually formation of a common intermediate, e.g., an enolate anion. The reactions catalyzed by members of the enoyl-CoA hydratase superfamily (almost) always utilize acyl esters of CoA as substrates the reactions invariably can be rationalized with mechanisms that involve the formation of a thioester enolate anion intermediate, e.g., 1,3-proton transfer, 1,5-proton transfer, Dieckman and reverse Dieckman condensations, and yS-decarboxylation. Although mechanisms with thioester enolate anion intermediates are plausible for each of these reactions, as in the ECH-catalyzed reaction, evidence for their existence on the reaction coordinate is circumstantial because the intermediates do not accumulate, thereby avoiding spectroscopic detection. 

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