Targeting protein-protein interactions as drug targets

7.项目摘要。该提案描述了一项确定结核病药物靶点的计划， 通过计算和实验相结合的方法来阐明大肠杆菌感染 分枝杆菌和细菌脂肪酸合成中的靶点。在大肠杆菌中，该途径涉及27 AcpP与其伴侣蛋白之间的离散相互作用。在结核病中，FAS-II途径涉及 产生分枝菌酸，其在结核分枝杆菌（Mtb）的毒力中起关键作用。作为 这两种途径都需要一种酰基载体蛋白（ACP）来传递生长的脂肪酰基底物和中间体 对于相关的伴侣蛋白（PP），AcpP.PP复合物内的蛋白质-蛋白质相互作用是必需的。 以获得有效的底物持续合成能力。这些组件的结构和相互作用的解析 负责它们形成的蛋白质不仅会进一步加深我们对这些蛋白质形成机制的了解， 功能和调节，但也将使基于结构的药物发现工作。为了阐明这些 结构，我们将利用我们的知识，大肠杆菌AcpP.PP接口，以确定一个 计算对接协议，产生新的Mtb AcpP.PP结构。我们的计算方法 将通过评估其概括E.已知结构的有效性来优化。大肠杆菌AcPP.PP 通过NMR和X-射线晶体学研究解析复合物。这个计算协议，一旦测试，将 可用于预测AcpP.PP和AcpM.PP复合物。然后，所得复合物的结构将 使用NMR滴定和诱变实验进行验证。生成的模型将使用 分子动力学模拟，它产生了一个动态的理解蛋白质结构，将告知 计算机辅助药物发现的努力。

7. Project Summary. This proposal describes a program to identify drug targets for tuberculosis and Escherichia coli infections through the combination of computational and experimental methods to elucidate targets within mycobacterial and bacterial fatty acid synthesis. In Escherichia coli, the pathway involves 27 discrete interactions between AcpP and its partner proteins. In tuberculosis, the FAS-II pathway is involved the production of mycolic acids, which serve a pivotal role in the virulence of Mycobacterium tuberculosis (Mtb). As both pathways require an acyl carrier protein (ACP) to deliver growing fatty acyl substrates and intermediates to associated partner proteins (PPs), protein-protein interactions within the AcpP.PP complex are an essential for effective substrate processivity. Resolution of the structures of these assemblies and the interactions responsible for their formation will not only further our knowledge of the mechanisms by which these proteins function and are regulated but will also enable structure-based drug discovery work. In order to elucidate these structures, we will leverage our knowledge of the Escherichia coli AcpP.PP interfaces to identify a computational docking protocol that produces novel Mtb AcpP.PP structures. Our computational methodology will be optimized by evaluating its effectiveness in recapitulating known structures of E. coli AcPP.PP complexes resolved via NMR and X-ray crystallography studies. This computational protocol, once tested, will be used to predict both AcpP.PP and AcpM.PP complexes. The structures of the resulting complexes will then be validated using NMR titration and mutagenesis experiments. The resulting models will be refined using molecular dynamics simulations, which yield a dynamical understanding protein structure that will inform computer-aided drug discovery efforts.