Characterization of novel MurA inhibitors

描述（由申请人提供）：青霉素和其他现代抗生素控制感染性疾病的能力已被多种抗生素耐药菌株的出现所侵蚀。CDC估计，每年有9万美国人死于细菌感染，其中70%的感染涉及耐药性菌株。细菌的存活严格依赖于胞质酶MurA的功能，MurA催化细菌细胞合成中的第一个关键步骤。它是天然存在的广谱抗生素磷霉素的靶点。磷霉素是一种环氧化物，共价攻击MurA的关键半胱氨酸残基（Cys 115）。这种残基导向的作用模式使磷霉素成为一种相对较差的药物，因为在病原菌如结核分枝杆菌中，Cys 115被天冬氨酸取代。此外，由于磷霉素向细胞中的转运减少，以及磷霉素抗性蛋白（FosA）的失活，越来越多的病原菌已对磷霉素产生抗性。因此，迫切需要开发通过不同分子作用模式靶向MurA的新型药物。 我们最近确定了五个新的MurA抑制剂独特的支架和IC 50值范围从2 μ M到8 μ M的高通量筛选（HTS）使用50，000化合物库。本提案的中心目标是彻底评估这些新的先导结构对MurA的作用方式。具体的目标，它集成了酶动力学和蛋白质晶体学，是执行抑制动力学和确定的MurA与这些抑制剂结合的晶体结构，以解决原子细节的结构-活性关系。该提案的长期目标是为合理设计特异性靶向MurA的新型强效广谱抗菌药物提供基础。

DESCRIPTION (provided by applicant): The ability of penicillin and other modern antibiotics to control infectious diseases has been steadily eroded by the emergence of multiple-antibiotic-resistant strains of bacteria. The CDC estimates that 90,000 Americans die from bacterial infections annually, with 70% of these infections involving antibiotic-resistant strains. Bacterial survival strictly depends on the functionality of the cytosolic enzyme MurA, which catalyzes the first committed step in the synthesis of the bacterial cell. It is the target of the naturally occurring, broad spectrum antibiotic fosfomycin. Fosfomycin, an epoxide, covalently attacks a critical cysteine-residue of MurA (Cys115). This residue-directed mode of action renders fosfomycin a relatively poor drug, because in pathogenic bacteria, such as Mycobacterium tuberculosis, Cys115 is replaced by an aspartate. In addition, an ever-increasing number of pathogenic bacteria have developed resistance to fosfomycin due to decreased transport of fosfomycin into the cell, and inactivation by a fosfomycin resistance protein (FosA). Thus, there is a critical need for the development of novel drugs targeting MurA by a different molecular mode of action. We have recently identified five new MurA inhibitors with unique scaffolds and IC50 values ranging from 2 mu M to 8 mu M by high-throughput screening (HTS) using a 50,000 compound library. The central goal of this proposal is to thoroughly evaluate the mode of action of these new lead structures on MurA. The specific aim, which integrates enzyme kinetics and protein crystallography, is to perform inhibition kinetics and determine the crystal structure of MurA bound with these inhibitors in order to resolve the structure-activity relationships in atomic detail. The long-term goal of this proposal is to provide a basis for the rational design of novel potent broad-spectrum antibacterial drugs that specifically target MurA.