CYCLOPROPYL METABOLITES, ENZYMATIC INHIBITION/FORMATION

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The cyclopropyl groups has been found to be an important structural feature in a wide variety of natural products. Recently, many novel enzyme inhibitors and mechanistic probes have been designed based upon the intrinsic ring strain and associated properties of this moiety. Intrigued by the unique biological activity of cyclopropyl metabolites and their potential use as therapeutic agents, we have undertaken an investigation into the mechanistic details of cyclopropanoid-induced enzyme inactivation. In the past few years we have focused our efforts on studying the inactivation mechanism of acyl-CoA dehydrogenases by (methylenecyclo-propyl) acetyl-CoA (MCPA-CoA), the causative agent of Jamaican vomiting sickness. Our initial results appear to support a radical mechanism, and we have recently begun to explore whether a strong hydrogen bond is involved in the activation of the alpha-H (pKa approximately 16) of the inhibitor. We will also continue to investigate the inactivation of crotonase by (methylenecyclopropyl) formyl-CoA (MCPF-CoA). Both studies are particularly relevant towards the development of class-selective inhibitors as possible therapeutic elements in the regulation of fatty acid metabolism to treat hyperglycemia. Another system we are studying is 1-aminocyclopropane-1- carboxylate (ACC) deaminase, which is a PLP-dependent enzyme catalyzing the ring cleavage of a variety of cyclopropyl amino acids, a reaction unique to vitamin B6 catalysts. To enhance our overall understanding of the bioprocessing of cyclopropyl metabolites, we have initiated a study of the biosynthesis of cyclopropane fatty acids (CFAs) by CFA synthase. This enzyme catalyzes a the cyclopropanation of cis-double bonds in phospholipid side chains which is a mechanistically intriguing reaction, and it may be a possible target for the antibiotic treatment of drug-resistant bacterial infections. Overall, our efforts will require a multi-faceted approach including the expression and purification of the desired enzymes, the synthesis of substrate analogs, inhibitors, and cofactors, the characterization of products isolated from enzymatic incubations with these compounds, and the use of physical and spectroscopic methods for assessing the course and kinetics of the reactions. These mechanistic studies will not only provide valuable information for designing strategies to control and/or mimic the catalytic roles of the target enzymes, but will also enable us to refine our assessment concerning the potential use of the cyclopropyl group as a mechanistic probe. Thus, our anticipated results should make a significant contribution to the broad field of mechanistic enzymology.