Chemical Control of Quorum Sensing in <i>E. coli</i>: Identification of Small Molecule Modulators of SdiA and Mechanistic Characterization of a Covalent Inhibitor

Abstract

Enterohemorrhagic <i>Escherichia coli</i> (EHEC) is the causative agent of severe diarrheal disease in humans. Cattle are the natural reservoir of EHEC, and approximately 75% of EHEC infections in humans stem from bovine products. Many common bacterial pathogens, including EHEC, rely on chemical communication systems, such as quorum sensing (QS), to regulate virulence and facilitate host colonization. EHEC uses SdiA from <i>E. coli</i> (SdiA_EC), an orphan LuxR-type receptor, to sense <i>N</i>-acyl l-homoserine lactone (AHL) QS signals produced by other members of the bovine enteric microbiome. SdiA_EC regulates two phenotypes critical for colonizing cattle: acid resistance and the formation of attaching and effacing lesions. Despite the importance of SdiA_EC, there is very little known about its selectivity for different AHL signals, and no chemical inhibitors that act specifically on SdiA_EC have been reported. Such compounds would represent valuable tools to study the roles of QS in EHEC virulence. To identify chemical modulators of SdiA_EC and delineate the structure-activity relationships (SARs) for AHL activity in this receptor, we report herein the screening of a focused library composed largely of AHLs and AHL analogues in an SdiA_EC reporter assay. We describe the identity and SARs of potent modulators of SdiA_EC activity, examine the promiscuity of SdiA_EC, characterize the mechanism of a covalent inhibitor, and provide phenotypic assay data to support that these compounds can control SdiA_EC-dependent acid resistance in <i>E. coli</i>. These SdiA_EC modulators could be used to advance the study of LuxR-type receptor/ligand interactions, the biological roles of orphan LuxR-type receptors, and potential QS-based therapeutic approaches.