Membrane Protein Structures and Interactions in the M. tuberculosis Divisome

 描述(申请人提供)：对结核分枝杆菌(结核分枝杆菌)的细胞分裂机制或分裂体知之甚少，结核分枝杆菌是结核病的病原体。自从20世纪70年代引入了为期6个月的多药疗程以来，就没有开发出新的结核病药物类别。在过去十年中，耐多药和广泛耐药结核病的发病率迅速增加，因此迫切需要新的治疗方法。该项目的目标是促进对作为潜在药物靶点的Mtb分裂组的了解，并为此实现对分裂体蛋白质如何相互作用的理解，从而了解它们如何被招募到分裂组中。四种跨膜蛋白作为结构表征的目标，既有单独的，也有作为复合体的。具体目标是：(1)表征蛋白质靶标中固有无序区域的新生和结合诱导的结构；(2)确定这些蛋白质的全长结构I脂双层；(3)确定这些膜蛋白之间以及与其他分裂体蛋白和肽聚糖前体或片段形成的复合体的结构。将微生物学和结构生物学的专业知识与膜蛋白和计算生物物理学相结合，这项研究将带来关于分裂组中心层的活动和相互作用的关键知识。将使用一系列工具，包括细菌双杂交分析、固体和溶液核磁共振以及限制分子动力学。为了应对在类天然脂质双层环境中表征蛋白质和蛋白质复合体的挑战，将开发新的结构方法，包括一种来自固体核磁共振的新型结构约束，以及一种用于表征膜蛋白固有无序区域的新型固体核磁共振。这个跨学科的团队，拥有重要和独特的个人经验，并通过先前的联合出版物，处于完成拟议研究的理想位置。结核分枝杆菌分裂组的这些结构特征及其功能含义将为结核病创造新的治疗机会。这里开发的技术将代表膜蛋白结构生物学的前沿。1

 DESCRIPTION (provided by applicant): Relatively little is known about the cell division machinery, or divisome, of Mycobacteria tuberculosis (Mtb), the causative agent of tuberculosis (TB). There have been no new TB drug classes developed since a multidrug 6-month treatment course was introduced in the 1970s. New therapies are desperately needed for TB as the occurrence of multi-drug and extensive drug-resistant TB has increased rapidly in the past decade. The goal of this project is to advance knowledge of the Mtb divisome as potential drug targets and towards this end achieve an understanding of how the divisome proteins interact with each other and hence, how they are recruited to the divisome. Four transmembrane proteins are targeted for structural characterizations, both individually and as complexes. The Specific Aims are: (1) characterize nascent and binding-induced structures of intrinsically disordered regions in the protein targets; (2) determine full-length structures of these proteins i lipid bilayers; (3) determine structures of complexes formed between these membrane proteins and with other divisome proteins and peptidoglycan precursors or fragments. Combining expertise in microbiology and structural biology with membrane protein and computational biophysics, the research will lead to critical knowledge on the activities and interactions of central layers in the divisome. An array of tools will be used, including bacterial two-hybrid assays, solid state and solution NMR, and restrained molecular dynamics. To meet the challenge of characterizing the proteins and protein complexes in a native-like, lipid bilayer environment, new structural methodologies, including a novel type of structural restraints from solid state NMR and a novel use of solid state NMR for conformational characterization of intrinsically disordered regions in membrane proteins, will be developed. This interdisciplinary team, with significant and unique experiences of the individuals and through prior joint publications, is ideally positioned to accomplish the proposed studies. These structural characterizations of the Mtb divisome and their functional implications will generate novel therapeutic opportunities for TB. The technologies developed here will represent the frontier of membrane protein structure biology. 1