Project 2: Targeting Gram-positive Cell Envelope Assembly

Project 2: Targeting Gram-positive Cell Envelope Assembly Invasive methicillin-resistant Staphylococcus aureus (MRSA) and drug-resistant Streptococcus pneumoniae infections together are responsible for almost 50% of all deaths due to antibiotic-resistant infections in the United States. Their prevalence, conservatively estimated at a combined 900,000 infections each year, means that they will continue to be a threat for the foreseeable future. It is crucial to identify new vulnerabilities in these organisms and to maintain a pipeline of targets and compounds that can be used to treat the infections they cause. In this proposal, we will pursue cell envelope targets in S. aureus and S. pneumoniae, emphasizing those that are critical for barrier integrity because targeting them could allow continued use of antibiotics to which these organisms have developed widespread resistance, such as beta-lactams. Aim 1 will take a compound-driven approach to new target discovery in S. aureus that exploits an innovative platform that combines pathway- directed chemical screening with a transposon-based pipeline to rapidly identify targets for compounds that inhibit cell envelope processes. Aim 2 will take a target-driven approach that advances our understanding of the mechanism and structure of LtaS, a polytopic membrane protein that synthesizes lipoteichoic acids on the Gram- positive cell surface. Based on discoveries made with a tool compound, we will also identify S. aureus LtaS inhibitors using a pathway-directed screening approach. Aim 3 will focus on the regulation of teichoic acid polymers in S. pneumoniae. The balance between lipoteichoic acids and wall teichoic acids (WTAs) on the surface of this pathogen have been found to spatially control cell wall hydrolases to guide expansion of the PG meshwork during growth. To prevent bacteriolysis, WTA levels must be carefully controlled in S. pneumoniae, a vulnerability exploited by beta-lactams, which promote a dramatic increase in the WTA content of the wall. We have identified an essential WTA hydrolase (WhyD) that when inactivated mimics beta-lactam treatment to increase WTA levels and induce cell lysis. We will characterize the function and regulation of this enzyme and investigate its potential as a novel autolysis-inducing therapeutic target.