Human IMP dehydrogenase catalyzes the dehalogenation of 2-fluoro- and 2-chloroinosine 5'-monophosphate in the absence of NAD.

The ability of human type II inosine monophosphate dehydrogenase (IMPDH, EC 1.1.1.205) to catalyze the formation of xanthosine 5'-monophosphate (XMP) from C2 halogen-substituted analogs of IMP has been investigated. Adenosine deaminase was used to enzymatically synthesize 2-fluoroinosine and 2-chloroinosine from the 2-fluoro- and 2-chloroadenosine nucleoside analogs. Chemical phosphorylation yielded the corresponding 5'-nucleoside monophosphate derivatives. IMPDH catalyzes the conversion of both 2-fluoro- and 2-chloroinosine 5'-monophosphate (2-F- and 2-Cl-IMP) to XMP. The dehalogenation reactions proceed without nicotinamide adenine dinucleotide (NAD), the hydride acceptor required for the oxidation of IMP, the normal substrate of the enzyme. Formation of XMP from the 2-halo-IMPs was verified by UV absorption spectroscopy and by HPLC. Formation of XMP from 2-F-IMP yielded stoichiometric amounts of fluoride anion. IMP and XMP were competitive inhibitors toward 2-Cl-IMP in the dehalogenation reaction. Neither 2-F-IMP nor 2-Cl-IMP irreversibly inactivate IMPDH. Kinetic constants for the dehalogenation reactions have been determined and compared to the dehydrogenation reaction at 25 degrees C. (For 2-F-IMP: kcat = 0.058 s-1, Km = 62 microM. For 2-Cl-IMP: kcat = 0.049 s-1, Km = 48 microM. For the IMP dehydrogenation reaction: kcat = 0.25 s-1, Km [IMP] = 4.1 microM, Km [NAD] = 29 microM). Hydrolytic dehalogenation of 2-halo-IMPs without a requirement for NAD demonstrates the formation of a tetrahedral intermediate in the catalytic mechanism of IMP dehydrogenase.