A Novel and Efficient Platform for Discovering Noncanonical Quorum-Quenching Proteins

Quorum sensing (QS) is a well-known chemical signaling system responsible for intercellular communication that is widespread in bacteria. Acyl-homoserine lactone (AHL) is the most-studied QS signal. Previously, bacterially encoded AHL-degrading enzymes were considered to be canonical quorum-quenching proteins that have been widely used to control pathogenic infections. Here, we report a novel platform that enabled the efficient discovery of noncanonical AHL quorum-quenching proteins. This platform initially asked bacteriologists to carry out comparative genomic analyses between phylogenetically related AHL-producing and non-AHL-producing members to identify genes that are conservatively shared by non-AHL-producing members but absent in AHL-producing species. These candidate genes were then introduced into recombinant AHL-producing E. coli to screen for target proteins with the ability to block AHL production. Via this platform, we found that non-AHL-producing Lysobacter containing numerous environmentally ubiquitous members encoded a conserved glycosyltransferase-like protein Le4759, which was experimentally shown to be a noncanonical AHL-quenching protein. Le4759 could not directly degrade exogenous AHL but rather recognized and altered the activities of multiple AHL synthases through protein-protein interactions. This versatile capability enabled Le4759 to block specific AHL synthase such as CarI from Pectobacterium carotovorum to reduce its protein abundance to suppress AHL synthesis, thereby impairing bacterial infection. Thus, this study provided bacteriologists with a unique platform to discover noncanonical quorum-quenching proteins that could be developed as promising next-generation drug candidates to overcome emerging bacterial antibiotic resistance. IMPORTANCE Targeting and blocking bacterial quorum sensing (QS), the process known as quorum quenching (QQ) is an effective mean to control bacterial infection and overcome the emerging antibiotic resistance. Previously, diverse QS signal-degradation enzymes are identified as canonical QQ proteins. Here, we provided a novel and universal platform that enabled to discover previously unidentified noncanonical QQ proteins that were unable to degrade acyl-homoserine lactone (AHL) but could block AHL generation by recognizing multiple AHL synthases via direct protein-protein interactions. Our findings are believed to trigger broad interest for bacteriologists to identify potentially widely distributed noncanonical QQ proteins that have great potential for developing next-generation anti-infectious drugs.