Antibiotic resistance is an increasing global problem, in 2019 4.95 million people died of illnesses where antimicrobial resistance was a factor. The ESKAPE pathogens are of particular concern due
to their high level of resistance to existing antimicrobials, therefore the search for novel drugs and targets is becoming increasingly important. One currently unexplored area for novel drug targets is the TCA cycle, which is essential for cell survival. Bacterial cells conserve carbon in the TCA cycle with the glyoxylate shunt, which is catalysed by isocitrate lyase 1 (ICL1). ICL1 is not present in
human cells thus making it an ideal target for antimicrobials. A novel drug has been shown to inhibit ICL1 from Mycobacterium tuberculosis, this study investigates the wider antimicrobial activity of the novel drug against ICL1 from Burkholderia pseudomallei, Brucella melitensis and Pseudomonas aeruginosa. Recombinant ICL1 proteins from the different bacterial species were expressed and purified using histidine tags for nickel ion affinity chromatography. The activity of the different ICL1 species in the presence of the drug was then studied using a phenylhydrazine reporter assay, which found the drug significantly (p<0.001) reduced the rate of reaction to levels comparable to known inhibitor itaconate for all of the species. The drug was most effective at reducing the rate of reaction in B. pseudomallei (by 58%), followed by B. melitensis (55%) and finally P. aeruginosa (34%). Inhibitor binding was then studied using differential scanning fluorimetry which suggested on average the drug increased the melting temperature by 14.75°C (+-2.22°C), which was higher than itaconate (8.86°C +-6.20°C). The activity of the drug was not correlated to the similarity of the active site but did correlate to the evolutionary distance from M. tuberculosis ICL1, suggesting the drug binds to a region of similarity away from the active site.
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