Mechanistic Insights into the Antimicrobial Effect of Benzodioxane-Benzamides Against Escherichia coli

Background/Objectives: The bacterial cell division machinery is emerging as an attractive target for antimicrobial compounds. FtsZ, a highly conserved essential division protein, is the target for a number of small molecules such as benzamides. Recent studies show that benzodioxane-benzamides (BDOBs) are among the most potent inhibitors of FtsZ function in Gram-positive bacteria, although their ability to inhibit Gram-negative FtsZ, in particular Escherichia coli FtsZ, has been more controversial. Methods: Here, we use genetic and cytological methods to demonstrate that FtsZ of efflux pump-disabled E. coli can be efficiently targeted by BDOBs. Results: We show that engineered mutants and spontaneous variants map in or near the interdomain cleft (IDC) of FtsZ that confers resistance to BDOBs, similar to previous results with Gram-positive FtsZs. We also uncover spontaneous extragenic mutants that can confer high levels of resistance to at least one potent BDOB, including a mutant that encodes a novel hyperfission variant of the essential cell division protein FtsW. Conclusions: Our evidence indicates that as with Gram-positive bacteria, the IDC of Gram-negative bacterial FtsZ is directly targeted by BDOBs, provided efflux pumps are disabled. We also conclude that FtsZ-independent factors can influence the effect of BDOBs on E. coli cell division, including activation of division septum synthesis.