Function of SecA2 in Mycobacterium tuberculosis protein export

描述（由申请方提供）：结核分枝杆菌是结核病（TB）的病原体。结核病每年造成约200万人死亡，世界上约三分之一的人口感染结核病。结核病每年新感染世界人口的近1%。结核病是艾滋病毒/艾滋病患者的主要死因。虽然结核病通常可以用抗生素治疗，但在全世界抗击结核病的斗争中，一个严重的问题是结核病的多重耐药菌株的出现。为了开发新药的合理靶点，必须更好地了解分枝杆菌的生理学。病原菌如M.结核杆菌分泌蛋白质以逃避宿主防御机制并在宿主细胞中存活，使得蛋白质输出成为合乎逻辑的药物靶点。这项工作的长期目标是表征M的Sec依赖性蛋白输出。结核涉及二聚体SecA和SecYEG的Sec-dependent易位途径用于许多蛋白质的输出。SecA是一种在所有细菌中发现的必需蛋白质，是一种ATP酶，并通过SecYEG膜易位机制提供蛋白质输出所需的能量。近年来，包括M.已发现结核病携带两种SecA蛋白，SecA 1和SecA 2。SecA 1是一般蛋白输出所必需的，而SecA 2是某些毒力蛋白分泌的特异性蛋白。然而，为什么这些微生物需要两种SecA蛋白尚不清楚。由于SecA蛋白通常对特定微生物具有特异性，因此SecA 1和SecA 2为药物开发提供了良好的靶点。因此，我们建议表征这些SecA蛋白中的每一个在从M.结核我们将使用生物化学和生物物理学技术，包括ATP酶结合和水解测定，分析性超电泳监测亚基协会，SecA蛋白的特点，并开发体外膜结合和易位测定M。结核蛋白此R 03补助金的具体问题是：1）。SecA 1和SecA 2是否具有相似的结合和水解ATP的能力？2)SecA 1和SecA 2相互作用吗？和3.）SecA 1和SecA 2如何与膜转运机制相互作用？设计这些实验以开始深入表征两种SecA蛋白在M.结核结核分枝杆菌（Mycobacterium tuberculosis）是结核病（TB）的病原体。结核病每年造成约200万人死亡，世界上约三分之一的人口感染结核病。结核病每年新感染世界人口的近1%。结核病是艾滋病毒/艾滋病患者的主要死因。虽然结核病通常可以用抗生素治疗，但世界范围内防治结核病的一个严重问题是结核病的多重耐药菌株的出现。为了开发新药的合理靶点，必须更好地了解分枝杆菌的生理学。

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis is the causative agent for the disease tuberculosis (TB). TB kills about 2 million people each year and around a third of the world's population is infected with TB. TB newly infects nearly 1% of the world's population each year. TB is a leading cause of death among people with HIV/AIDS. Though TB can normally be treated with antibiotics, a serious problem in the worldwide fight against TB is the emergence of multi-drug resistant strains of TB. In order to develop logical targets for new drugs, the physiology of Mycobacteria must be better understood. Pathogenic bacteria such as M. tuberculosis secrete proteins in order to evade host defense mechanisms and survive in host cells, making protein export a logical drug target. The long-term goal of this work is to characterize the Sec-dependent protein export of M. tuberculosis. The Sec-dependent translocation pathway that involves dimeric SecA and SecYEG is used for export of many proteins. SecA, an essential protein found in all bacteria, is an ATPase and provides the energy used in the export of proteins through the SecYEG membrane translocation machinery. Recently, several pathogenic microorganisms, including M. tuberculosis, have been discovered to carry two SecA proteins, SecA1 and SecA2. SecA1 is essential for general protein export while SecA2 is specific for secretion of some virulence proteins. However, why these microorganisms require two SecA proteins is not understood. Because SecA proteins are generally specific to a particular microorganism, SecA1 and SecA2 provide good targets for drug development. Therefore, we propose to characterize the function of each of these SecA proteins in protein export from M. tuberculosis. We will characterize the SecA proteins using biochemical and biophysical techniques including ATPase binding and hydrolysis assays, analytical ultracentrifugation to monitor subunit associations, and develop in vitro membrane binding and translocation assays for M. tuberculosis proteins. The specific questions for this R03 grant are: 1). Do SecA1 and SecA2 possess similar abilities to bind and hydrolyze ATP?; 2) Do SecA1 and SecA2 interact with each other?; and 3.) How do SecA1 and SecA2 interact with membrane translocation machinery? These experiments are designed to initiate an in depth characterization of the function of both SecA proteins in M. tuberculosis. Mycobacterium tuberculosis is the causative agent for the disease tuberculosis (TB). TB kills about 2 million people each year and around a third of the world's population is infected with TB. TB newly infects nearly 1% of the world's population each year. TB is a leading cause of death among people with HIV/AIDS. Though TB can normally be treated with antibiotics, a serious problem in the world-wide fight against TB is the emergence of multi-drug resistant strains of TB. In order to develop logical targets for new drugs, the physiology of Mycobacteria must be better understood.