Combating Fosfomycin Resistance in Methicillin-Resistant Staphylococcus aureus

项目总结

PROJECT SUMMARY The treatment of bacterial infections is compromised by the spread of multidrug-resistant (MDR) and extensively drug-resistant (XDR) pathogens. New strategies to combat resistant as well as emerging bacteria are urgently needed to limit further development of antimicrobial resistance. Combining currently approved antibiotics with resistance neutralizing agents is a promising direction. The overall goal of our research is to develop effective front-line antimicrobial strategies from an approved, safe, and broad-spectrum antibiotic so newer antimicrobials, as well as final-option drugs, can be held in reserve to minimize emerging resistance. Fosfomycin is effective against both Gram-negative and Gram-positive pathogens and represents a promising candidate for developing a front-line agent. Primary resistance to fosfomycin arises from fosfomycin-modifying enzymes of the Vicinal Oxygen Chelate (VOC) superfamily. The primary role of VOC enzymes is to detoxify endogenous and xenobiotic compounds. Genes encoding VOC fosfomycin resistance enzymes have been identified in almost all of the most drug-resistant Gram-negative and Gram-positive pathogens and in Mycobacteria. FosB is the principal fosfomycin-modifying enzyme of methicillin-resistant Staphylococcus aureus (MRSA). It covalently attaches bacillithiol (BSH) to fosfomycin, inactivating the antibiotic. FosB knockout strains of MRSA demonstrate significantly increased susceptibility to fosfomycin, identifying suppression of the enzyme as a potential therapeutic strategy. We hypothesize that combining X-ray structural data of BSH-bound FosB with structure-based virtual inhibitor screening will identify small molecules that can serve as FosB inhibitors and lower the MIC of fosfomycin in MRSA, thereby making fosfomycin an effective treatment against MRSA. In Aim 1, we will use structure-based virtual screening to identify new small molecule scaffolds that inhibit FosB and evaluate them with respect to kinetics and synergistic effectiveness when combined with fosfomycin. In addition, we will determine crystal structures of FosB with the new compounds to guide future medicinal chemistry approaches. In Aim 2, we will determine a novel structure of FosB complexed with BSH. To date, none of the VOC fosfomycin resistance enzymes have been structurally characterized with respect to their native thiol, and a structure of FosB with BSH bound will be transformative to our understanding of the mechanism of VOC- catalyzed fosfomycin resistance. The BSH-bound structure will then be used as a starting model for additional structure-based virtual screening. This research will involve undergraduate students in meaningful projects that expose them to a wide range of techniques and help develop their critical thinking, research skills, and interest in biomedical careers.