DESCRIPTION (provided by applicant): The ability of microbes to acquire iron is a key aspect of microbial invasiveness and pathogenicity. TonB-dependent transporters in the Gram-negative bacterial outer membrane catalyze the uptake of iron complexes called ferric siderophores, and the assimilation of these metal chelates is a determinant of the pathogenesis of many species, including Escherichia coli, Salmonella typhi, Neisseria meningitidis, Vibrio cholerae and others. Unfortunately, the full molecular understanding of these transport reactions is lacking, which impairs our ability to conceive of potential inhibitors. Nevertheless, it's known that the E. coli outer membrane protein FepA accomplishes active iron transport by a process that involves extensive conformational motion. The proposed research capitalizes on this knowledge with a fluorescence spectroscopic assay that monitors uptake of the siderophore ferric enterobactin. The experiments will encompass high-throughput screening of validation and primary libraries of compounds at the University of Kansas High-Throughput Screening facility. The three libraries include standards and test chemicals that total ~17,500 unique molecules. These tests, performed in 384-well microtiter plates, will identify compounds that inhibit TonB-dependent iron transport by Gram- negative bacteria, creating a panel of candidates for further testing as antibiotics against bacterial pathogenesis.

PUBLIC HEALTH RELEVANCE: Bacteria need iron to survive in the wild and in animal hosts. The proposed research will use a novel fluorescence assay to screen large libraries of compounds for chemicals that inhibit Gram-negative bacterial iron acquisition, thereby thwart bacterial growth, and lead to the development of new antibiotics against bacterial pathogens.