Targeting the FtsH protease unmasks a universal vulnerability to antimicrobial peptoids

The rapid rise of multidrug-resistant (MDR) bacteria highlights the need for new antimicrobial agents beyond traditional mechanisms. Synthetic peptoids are promising therapeutics. Mechanistic studies reveal peptoids kill bacteria not by damaging membranes but by causing widespread macromolecular aggregation of intracellular proteins and nucleic acids. This proteotoxic stress challenges the bacterial ATP-dependent protease system, though precise defense pathways remain elusive. Understanding how bacteria counter this stress is essential for developing effective combination treatments. We showed that FtsH is the key protective mechanism, as only ΔftsH drives a 16-fold sensitization, identifying it as the bottleneck for survival under peptoid stress. Its protective role fundamentally depends on bacterial energy metabolism. Disabling FtsH increases susceptibility in Pseudomonas aeruginosa (P. aeruginosa), suggesting that FtsH represents a shared vulnerability across tested peptoid scaffolds. This uncovers a common vulnerability within the proteostasis network of MDR pathogens. FtsH is intrinsically less active under low-energy conditions typical of antibiotic-tolerant persister cells. This metabolic repression renders persisters uniquely susceptible to peptoid-induced proteotoxic stress. Combining the peptoid (TM5) with an FtsH inhibitor (carbonyl cyanide m-chlorophenyl hydrazone) or an energy-depleting compound is highly effective, providing a mechanism-driven strategy to overcome persister cell's drug tolerance.