The pace of antibiotic development and approval is woefully insufficient to counter antibacterial resistance, creating a dire health care situation tht is likely to worsen given the recent global emergence of bacteria species, such as Acinetobacter baumannii, that are completely resistant to all current antibiotics. In response, agencies, such as the Infectious Diseases Society of America, have called for renewed investment in antimicrobial development strategies and approaches. To that end, we have recently shown that components of the Gram-positive bacterial RNA degradation apparatus are attractive antibiotic targets. But, small molecule inhibitors of this machinery do not affect RNA turnover in Gram-negative bacteria, which produce and rely on an entirely different set of mRNA degradation enzymes. The current application is designed to evaluate the promise of the key mediator of Gram-negative RNA degradation, ribonuclease E (RNase E), as a target for antimicrobial development. Our rationale for doing so is that the enzyme is essential, well conserved across Gram-negative pathogens of immediate health care concern, and it does not display similarity to any predicted mammalian protein. Accordingly, we have developed and validated high throughput screening and secondary assays to measure Escherichia coli, A. baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa RNase E activity. These assays will be used to launch a conventional enzymatic HTS of small molecule compound libraries as a means to identify RNase E inhibitors (Aim 1). In parallel, we will also perform a conventional fragment based screening approach that exploits the solved E. coli crystal structure, to improve our probability of identifying quality RNase E inhibitors (Aim 2). Compounds from either/both approaches will progress through a common series of cheminformatics, selectivity, antimicrobial, cytotoxicity, and cellular mechanism of action studies to ultimately determine whether quality small molecule RNase E inhibitors that display on-target antimicrobial activity can be readily identified.
Current antibiotics are losing their foothold as treatment options for bacterial infections. Consequently, there is an undeniable need for new therapeutic strategies for the intervention of these organisms. This project will test the promise of the centrl component of the Gram-negative RNA turnover machinery, ribonuclease E (RNase E), as a novel target for antimicrobial development.
