Conformational Switch to a β-Turn in a Staphylococcal Quorum Sensing Signal Peptide Causes a Dramatic Increase in Potency.

We report the solution-phase structures of native signal peptides and related analogs capable of either strongly agonizing or antagonizing the AgrC quorum sensing (QS) receptor in the emerging pathogen Staphylococcus epidermidis. Chronic S. epidermidis infections are often recalcitrant to traditional therapies due to antibiotic resistance and formation of robust biofilms. The accessory gene regulator (agr) QS system plays an important role in biofilm formation in this opportunistic pathogen, and the binding of an autoinducing peptide (AIP) signal to its cognate transmembrane receptor (AgrC) is responsible for controlling agr. Small molecules or peptides capable of modulating this binding event are of significant interest as probes to investigate both the agr system and QS as a potential antivirulence target. We used NMR spectroscopy to characterize the structures of the three native S. epidermidis AIP signals and five non-native analogs with distinct activity profiles in the AgrC-I receptor from S. epidermidis. These studies revealed a suite of structural motifs critical for ligand activity. Interestingly, a unique β-turn was present in the macrocycles of the two most potent AgrC-I modulators, in both an agonist and an antagonist, which was distinct from the macrocycle conformation in the less-potent AgrC-I modulators and in the native AIP-I itself. This previously unknown β-turn provides a structural rationale for these ligands' respective biological activity profiles. Development of analogs to reinforce the β-turn resulted in our first antagonist with subnanomolar potency in AgrC-I, while analogs designed to contain a disrupted β-turn were dramatically less potent relative to their parent compounds. Collectively, these studies provide new insights into the AIP:AgrC interactions crucial for QS activation in S. epidermidis and advance the understanding of QS at the molecular level.