Identification of novel diphenyl urea inhibitors of Mt-GuaB2 active against Mycobacterium tuberculosis

Abstract

In contrast with most bacteria, which harbour a single inosine monophosphate dehydrogenase (IMPDH) gene, the genomic sequence ofMycobacterium tuberculosisH37Rv predicts three genes encoding IMPDH:guaB1,guaB2andguaB3. These three genes were cloned and expressed inEscherichia colito evaluate functional IMPDH activity. Purified recombinant Mt-GuaB2, which uses inosine monophosphate as a substrate, was identified as the only active GuaB orthologue inM. tuberculosisand showed optimal activity at pH 8.5 and 37 °C. Mt-GuaB2 was inhibited significantlyin vitroby a panel of diphenyl urea-based derivatives, which were also potent anti-mycobacterial agents againstM. tuberculosisandMycobacterium smegmatis, with MICs in the range of 0.2–0.5 μg ml−1. When Mt-GuaB2 was overexpressed on a plasmidin transinM. smegmatis, a diphenyl urea analogue showed a 16-fold increase in MIC. Interestingly, when Mt-GuaB orthologues (Mt-GuaB1 and 3) were also overexpressed on a plasmidin transinM. smegmatis, they also conferred resistance, suggesting that although these Mt-GuaB orthologues were inactivein vitro, they presumably titrate the effect of the inhibitory properties of the active compoundsin vivo.