title: 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 of Mycobacterium tuberculosis H37Rv predicts three genes encoding IMPDH: guaB1, guaB2 and guaB3. These three genes were cloned and expressed in Escherichia coli to evaluate functional IMPDH activity. Purified recombinant Mt-GuaB2, which uses inosine monophosphate as a substrate, was identified as the only active GuaB orthologue in M. tuberculosis and showed optimal activity at pH 8.5 and 37 °C. Mt-GuaB2 was inhibited significantly in vitro by a panel of diphenyl urea-based derivatives, which were also potent anti-mycobacterial agents against M. tuberculosis and Mycobacterium smegmatis, with MICs in the range of 0.2-0.5 μg ml(-1). When Mt-GuaB2 was overexpressed on a plasmid in trans in M. 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 plasmid in trans in M. smegmatis, they also conferred resistance, suggesting that although these Mt-GuaB orthologues were inactive in vitro, they presumably titrate the effect of the inhibitory properties of the active compounds in vivo.
authors.affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
authors.forename: Veeraraghavan
authors.initials: V
authors.lastname: Usha
authors.affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
authors.forename: Sudagar S
authors.initials: SS
authors.lastname: Gurcha
authors.affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
authors.forename: Andrew L
authors.initials: AL
authors.lastname: Lovering
authors.affiliation: Department of Biological Sciences, University of Warwick, Coventry, UK.
authors.forename: Adrian J
authors.initials: AJ
authors.lastname: Lloyd
authors.affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
authors.forename: Athina
authors.initials: A
authors.lastname: Papaemmanouil
authors.affiliation: Drug Discovery Division, Southern Research Institute, Birmingham, AL 35255, USA.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Reynolds
authors.affiliation: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
authors.forename: Gurdyal S
authors.initials: GS
authors.lastname: Besra
mesh_terms: D000595:Amino Acid Sequence; D000995:Antitubercular Agents / Q000737:chemistry / Q000378:metabolism*; D002224:Carbanilides / Q000737:chemistry / Q000378:metabolism*; D003001:Cloning, Molecular; D024881:Drug Resistance, Bacterial; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007291:Inosine Monophosphate / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D020102:Mycobacterium smegmatis / Q000187:drug effects / Q000201:enzymology / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D000995:Antitubercular Agents; D002224:Carbanilides; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D007291:Inosine Monophosphate; D007168:IMP Dehydrogenase