New inhibitors of the Pseudomonas aeruginosa enzyme, PqsE, and methods assessing their potential to induce a conformational change via active site binding

Abstract

Pseudomonas aeruginosa is an opportunistic pathogen known for its ability to produce virulence factors and form biofilms. These, among many other traits, enable P. aeruginosa to cause infections, evade host immune responses, and resist treatment with antimicrobial agents. Both the ability to form biofilms and produce virulence factors are regulated via the bacterial cell-cell communication process called quorum sensing. A key molecular event that enables quorum sensing in P. aeruginosa is the physical interaction between an enzyme, PqsE and a master quorum-sensing receptor/transcription factor RhlR, which regulates the expression of a wide variety of virulence-associated genes. Previous work identified active site mutations in PqsE that induce a conformational change weakening the interaction with RhlR. These active site mutations weakened the PqsE-RhlR interaction to the extent that several virulence genes were not activated, and the mutant strains of P. aeruginosa failed to colonize the lungs of a mouse. In this study, we designed a series of molecules to probe binding in the active site of PqsE as a strategy for inhibiting the PqsE-RhlR interaction and thus, the activation of RhlR-controlled genes in P. aeruginosa. HJ1 and HJ5 are new molecules that both bind in the active site of PqsE, and while HJ5 appears to bind in an alternate mode compared to HJ1, it does not induce a conformational change to weaken the PqsE-RhlR interaction. Here, we introduce multiple experimental approaches to assess the way in which these new molecules engage in the PqsE active site. HJ5 can serve as a promising starting point for the development of molecules that target the PqsE active site and allosterically inhibit the interaction with RhlR, thus decreasing virulence in P. aeruginosa.