| Abstract
| - (Methylenecyclopropyl)formyl-CoA (MCPF-CoA), a toxic metabolite of methylenecyclopropylglycine (MCPG), is known to have hypoglycemic activity due to its ability to interrupt the β-oxidation pathwayof fatty acid metabolism. Earlier experiments have shown that acetoacetyl-CoA thiolase, 3-ketoacyl-CoA thiolase,acyl-CoA dehydrogenases, and enoyl-CoA hydratase (ECH) are cellular targets that can be inhibited by MCPF-CoA and/or MCPG. To gain more insights with respect to the target specificity and the mode of action, wehave carried out a detailed investigation of the effects of MCPF-CoA on a variety of enzymes involved infatty acid metabolism. Our studies confirmed that MCPF-CoA is a potent inactivator for ECHs but showslittle effect on other β-oxidation enzymes tested in this study. Our results also revealed that MCPF-CoA manifestsdistinct modes of inhibition among ECHs isolated from different sources, being a competitive inhibitor for ratliver ECH and an irreversible inactivator for the bovine liver as well as pig kidney ECH. Given the highsequence homology of the mammalian ECH genes studied so far, the structures of these proteins are expectedto be similar. Thus, the effects of MCPF-CoA toward different ECHs must be governed in part by the interactionof MCPF-CoA with the active site of each ECH whose architecture may be subtly different. More importantly,the incubation results with bovine liver ECH established that MCPF-CoA inactivates this enzyme via amechanism involving the covalent trapping of an active site nucleophile by the methylenecyclopropane ring.Since MCPF-CoA is a rare irreversible inhibitor for ECHs, it could serve as a new lead for designing moreeffective agents for modulating ECH activity so as to control and/or regulate fatty acid metabolism.
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