Novel Inhibitors to DHPS to Probe Catalytic Mechanism & Therapeutic Potential - r

描述(申请人提供)：叶酸生物合成是高等动物所没有的一种重要的细菌途径，70多年来一直是抗菌药物的靶标。磺胺类药物通过作为对氨基苯甲酸(PABA)的非生产底物类似物来抑制该途径中的二氢蝶呤酸合酶(DHPS)。然而，灵活的PABA结合位点在结构上对耐药性突变敏感，磺胺类药物正在迅速变得无效。相比之下，第二个DHPS底物的结合部位，即蝶呤焦磷酸，被埋在一个保守的口袋里，不太可能容忍突变。我们建议产生新的有效的DHPS抑制剂，专门作用于这一口袋。这些抑制剂有可能被开发成新的治疗剂，仍然以叶酸合成为靶点，但避免了耐药性问题。要产生任何酶的有效抑制剂，了解其活性部位的结构和机制是至关重要的。对于DHPs来说，这种信息基本上是缺乏的，了解DHPs如何在分子水平上进行催化将是该提案的中心目标。另一个目标是了解磺胺耐药性的分子和功能基础，以避免在我们未来的药物设计中出现这个问题。这一全面的方法将需要结构生物学、药物化学、生物化学、计算生物学和微生物学方面的两位主要研究人员的共同专业知识。该项目还将纳入基于片段的药物发现这一相对较新的技术，以确定易于通过化学合成来阐述的新型小型抑制性化学支架。申请中描述的令人振奋的新数据为该项目能够实现其最终目标提供了强有力的支持，即快速开发新的抗菌药，将耐药性的出现降至最低。虽然我们的目标是开发广谱抗感染药物，但我们的微生物筛查将重点放在金黄色葡萄球菌和新城疫杆菌上，这些细菌近年来在美国出现了特别有问题的传染病病原体。 与公众健康相关：本提案旨在从结构上了解和开发磺胺类抗菌剂二氢蝶呤合成酶的酶靶标；研究酶的作用机制、耐药性和抑制剂设计。预计这些研究将导致新的抗菌剂和对靶向磺胺类耐药性的新理解。

DESCRIPTION (provided by applicant): Folate biosynthesis is an essential bacterial pathway that is absent in higher animals, and it has been a target of antibacterial agents for over 70 years. Sulfa drugs inhibit the enzyme dihydropteroate synthase (DHPS) in the pathway by acting as non-productive substrate analogs of p-aminobenzoic acid (pABA). However, the flexible pABA binding site is structurally susceptible to resistance mutations, and the sulfa drugs are rapidly becoming therapeutically ineffective. In contrast, the binding site of the second DHPS substrate, pterin-pyrophosphate, is buried in a conserved pocket that is less likely to tolerate mutations. We propose to generate new classes of potent DHPS inhibitors that specifically engage this pocket. These inhibitors can potentially be developed into novel therapeutic agents that still target folate synthesis but which avoid the problems of resistance. To generate effective inhibitors of any enzyme, it is crucial to understand the structure and mechanism of its active site. This information is largely absent for DHPS, and understanding how DHPS performs catalysis at the molecular level will be a central goal of the proposal. Another goal will be to understand the molecular and functional basis of sulfa drug resistance to avoid this problem in our future drug designs. This comprehensive approach will require the joint expertise of the two principal investigators in structural biology, medicinal chemistry, biochemistry, computational biology and microbiology. The project will also incorporate the relatively new technique of fragment based drug discovery to identify novel small inhibitory chemical scaffolds that can readily be elaborated by chemical synthesis. Exciting new data described in the application provide strong support that the project can realize its ultimate goal which is to rapidly develop new antimicrobials that minimize the emergence of drug resistance. Although we aim to develop broad-spectrum anti-infective agents, our microbiological screening will focus on S. aureus and P. jirovecii that have emerged as particularly problematical infectious disease agents in the U.S. in recent years. PUBLIC HEALTH RELEVANCE: This proposal is aimed at structurally understanding and exploiting the enzyme target of the sulfonamide antibacterial agents, dihydropteroate synthase; to study enzyme mechanism, drug resistance and inhibitor design. It is expected that these studies will lead to new antimicrobial agents and a new understanding of target based sulfonamide resistance.