Project Summary Quorum sensing is a bacterial cell-cell communication system that functions through thesynthesis, secretion, and detection of signaling molecules called autoinducers. Quorum sensing enablesbacteria to assess their cell density and coordinate group behaviors that are advantageous at high celldensity. Many bacteria that cause acute and chronic infections in humans use quorum sensing tocontrol expression of virulence factors and formation of biofilms, so inhibitors of quorum sensing holdsignificant promise as novel antibacterial agents. By targeting bacterial virulence as opposed to growthand survival, quorum sensing inhibitors avoid the selective pressure for drug resistance that is inherentto traditional antibiotics. The main quorum sensing system in Gram-negative bacteria is based on the synthesis anddetection of acyl homoserine lactone (HSL) autoinducers. Efforts to inhibit HSL-mediated quorumsensing have focused on developing synthetic HSL analogs that act as antagonists of HSL receptors.These compounds have been shown to block virulence controlled by quorum sensing, validating quorumsensing as an antibacterial drug target. The goal of the proposed project is to develop inhibitors of HSL synthase enzymes, analternative quorum sensing drug target that remains underexplored. Aim 1 involves the design andsynthesis of small-molecule transition state analogs of the HSL synthase enzymatic reaction, an inhibitordesign approach that has produced potent inhibitors of mechanistically similar enzymes. Aim 2 involvesassembling robust assay platforms that will evaluate the synthetic transition state analogs as HSLsynthase enzyme inhibitors and measure their ability to suppress bacterial behaviors controlled by HSLquorum sensing. The proposed project may result in potent, broad-spectrum inhibitors of HSL quorumsensing, validating HSL synthase enzymes as a novel antimicrobial drug target.