Combating Fosfomycin Resistance in Methicillin-Resistant Staphylococcus aureus

项目摘要 细菌感染的治疗受到多药耐药（MDR）和广泛耐药的传播的影响。 耐药（XDR）病原体。迫切需要新的策略来对抗耐药细菌和新出现的细菌。 需要限制抗生素耐药性的进一步发展。将目前批准的抗生素与 抗性中和剂是一个很有前途的方向。我们研究的总体目标是开发有效的 一线抗微生物策略，从一个批准的，安全的，广谱抗生素，所以更新的抗微生物剂， 以及最终选择药物，可以作为储备，以尽量减少新出现的耐药性。 磷霉素对革兰氏阴性和革兰氏阳性病原体都有效，并且代表了一种有前途的抗生素。 发展一线特工的候选人对磷霉素的原发性耐药来自磷霉素修饰的 邻位氧螯合物（VOC）超家族的酶。VOC酶的主要作用是解毒 内源性和外源性化合物。编码VOC磷霉素抗性酶的基因已被发现。 在几乎所有最耐药的革兰氏阴性和革兰氏阳性病原体中， 分枝杆菌。FosB是耐甲氧西林金黄色葡萄球菌的主要磷霉素修饰酶 （MRSA）。它共价连接bacillithiol（BSH）磷霉素，灭活抗生素。FosB基因敲除菌株 的MRSA表现出对磷霉素的敏感性显著增加， 作为一种潜在的治疗策略。我们假设结合BSH结合的FosB的X射线结构数据 基于结构的虚拟抑制剂筛选将鉴定可用作FosB抑制剂的小分子， 降低磷霉素在MRSA中的MIC，从而使磷霉素成为针对MRSA的有效治疗。在Aim中 1，我们将使用基于结构的虚拟筛选来鉴定抑制FosB的新的小分子支架， 评价它们与磷霉素联合时的动力学和协同作用。此外，本发明还提供了一种方法， 我们将用新化合物确定FosB的晶体结构，以指导未来的药物化学。 接近。在目标2中，我们将确定一个新的结构的FosB与BSH配合物。迄今为止， VOC磷霉素抗性酶的结构特征在于其天然巯基， 结合BSH的FosB结构将改变我们对VOC机制的理解- 催化磷霉素抗性。BSH结合的结构然后将被用作附加的初始模型。 基于结构的虚拟筛选。这项研究将使本科生参与有意义的项目， 让他们接触到广泛的技术，并帮助发展他们的批判性思维，研究技能和兴趣 在生物医学职业中。

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.