Synthesis of DL-threo-3-(1-fluoro-1-methylethyl)- and DL-threo-3-(1,1-difluoroethyl)malic acids. Mechanistic studies of 3-isopropylmalate dehydrogenase

Abstract

For both mechanistic studies and the development of novel inhibitors of 3-isopropylmalate dehydrogenase enzyme (IPMDH), which is involved in the rate-determining step in the biosynthetic pathway of the essential amino acid L-leucine, (2R*,3S*)-3-(1-fluoro-1-methylethyl)- and ( 2R*,3S*)-3-(1,1-difluoroethyl)malic acids (F-IPM and F₂-EM) were designed based on the concept of mechanism-based inhibition, and the reaction kinetics with these fluorinated substrates were analysed. The reaction of F-IPM with IPMDH was studied by NMR spectroscopy and product isolation. F-IPM underwent, after the normal enzyme reaction, the expected additional elimination reaction to afford an α,β-unsaturated carbonyl product, which turned out not to participate in any covalent-bond-forming reaction. The conformation of the reaction intermediate during the IPMDH reaction and the functional-group arrangement in the active site of IPMDH are discussed.