SYNTHESIS OF DL-THREO-3-(1-FLUORO-1-METHYLETHYL)- AND DL-THREO-3-(1,1-DIFLUOROETHYL)MALIC ACIDS - MECHANISTIC STUDIES OF 3-ISOPROPYLMALATE DEHYDROGENASE

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 F-2-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 alpha,beta-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.