Kinetic characterization of inosine monophosphate dehydrogenase of Leishmania donovani

Trypanosomatid protozoan pathogens are purine auxotrophs that are highly dependent on the enzyme inosine monophosphate dehydrogenase (IMPDH) for the synthesis of guanylate nucleotides. Enzymatic characterization of the Leishmania donovani IMPDH (LdIMPDH) overexpressed in E. coli revealed that this enzyme was highly specific for the substrates IMP and NAD+ with Kmapp values of 33 and 390 μM, respectively. In contrast to other IMPDHs, LdIMPDH exhibits no substrate inhibition in high concentrations of NAD+. Kinetic studies revealed that XMP and GMP were inhibitors with Ki values of ∼26 and 210 μM, respectively, suggesting that these nucleotides may regulate LdIMPDH activity. Mycophenolic acid was also a potent inhibitor of L. donovani IMPDH with a Ki value of ∼25 nM. Confocal immunofluorescence microscopy and subcellular fractionation localized LdIMPDH to the glycosome. Protein-protein interaction assays revealed that LdIMPDH associated tightly with glycosomal protein sorting receptor LdPEX5.