Role of FHL-mediated formate metabolism in Mycobacterium tuberculosis persistence

FHL介导的甲酸盐代谢在结核分枝杆菌持久性中的作用

基本信息

批准号：
8353015
负责人：
THOMAS C. ZAHRT
金额：
$ 22.62万
依托单位：
MEDICAL COLLEGE OF WISCONSIN
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-07-01 至 2014-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8353015
关键词：
Acid Fast Bacillae Staining Method Address Antibiotics Bacteria Biological Assay Biology Carbon Dioxide Cell Respiration Cessation of life Complement Complex Development Energy Metabolism Enterobacteriaceae Environment Enzymes Escherichia coli Formate dehydrogenase Formates Future Gases Gene Expression Genetic Granuloma Growth Human Hypoxia Immunosuppression In Vitro Individual Infection Knowledge Lead Link Lung diseases Maintenance Mediating Metabolism Morbidity - disease rate Multienzyme Complexes Mycobacterium tuberculosis Nitric Oxide Operon Oxidants Physiological Physiology Play Population Production Proteomics Respiration Role Shapes Signal Transduction Source Staging Testing Therapeutic Intervention Time Tuberculosis base formate hydrogenlyase in vivo insight latent infection mortality mutant novel oxidation pathogen research study response retinal rods success

项目摘要

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis is a rod-shaped, acid-fast, human-specific pathogen and is the causative agent of the respiratory disease tuberculosis (TB). The bacterium is a significant source of morbidity and mortality worldwide, and is currently predicted to infect approximately 2 billion individuals. The success of M. tuberculosis as a pathogen is closely linked with its ability to establish latent infections in susceptible individuas and reactivate at later timed during periods of immunosuppression. Determinants important for the establishment, maintenance, and reactivation of M. tuberculosis from long-term, persistent infection within the host are poorly understood. It is thought that latency initiates following encasement of M. tuberculosis within granulomatous lesions, and the recognition of specific signals present within this environment that inhibit aerobic respiration and promote the transition of M. tuberculosis into an altered physiological state of nonreplicating persistence (NRP). Recently, a locus (Rv0081-Rv0088) predicted to encode determinants comprising a formate hydrogenlyase (FHL) were found to be upregulated following exposure of M. tuberculosis to conditions (hypoxia and nitric oxide) promoting NRP in vitro. In addition, this locus was found to be directly regulated by two response regulators (DosR/DevR and MprA) known to contribute to persistence by M. tuberculosis in vitro and in vivo. FHL plays an important role in energy metabolism within enteric bacteria during periods of microaerophilic/anaerobic growth by mediating the oxidation of formate to CO2 and H2 in the absence of an external electron acceptor. This proposal seeks to fill a current gap in our knowledge by investigating aspects of M. tuberculosis metabolism under physiologically relevant conditions. We hypothesize that M. tuberculosis synthesizes a functional FHL enzyme complex that is required for energy metabolism and survival during periods of NRP. To address this hypothesis, two specific aims have been proposed. First, the ability of M. tuberculosis FHL to mediate the oxidation of formate to CO2 and H2 will be examined using spectrophotometric- and gas chromatographic-based approaches. Second, determinants comprising the predicted FHL will be examined to determine if they interact to form FHL and are required for survival of M. tuberculosis during NRP. Collectively, these studies are expected to provide novel insights into M. tuberculosis physiology under conditions associated with NRP and latency. Delineation of the basis for formate metabolism may also identify new enzymes that can be targeted for therapeutic intervention of M. tuberculosis during periods of NRP when the bacterium is otherwise recalcitrant to currently available antibiotics. PUBLIC HEALTH RELEVANCE: Experiments described in this proposal will investigate the importance of FHL-mediate formate metabolism in the physiology of Mycobacterium tuberculosis during non-replicating persistence. Examination of determinants comprising this enzyme complex is expected to fill a critical gap in our knowledge of M. tuberculosis physiology by furthering our understanding of M. tuberculosis metabolism under conditions associated with persistent infection. Completion of this proposal may lead to identification of new enzymes that can be targeted for therapeutic intervention of M. tuberculosis during latency when the bacterium is otherwise recalcitrant to currently available antibiotics.

描述（由申请人提供）：结核分枝杆菌是一种棒状，酸性，人类特异性病原体，是呼吸道疾病结核病（TB）的病因。该细菌是全球发病率和死亡率的重要来源，目前预计将感染约20亿个人。结核分枝杆菌作为病原体的成功与其在免疫抑制期间在后期的时间内重新激活并在后期重新激活其潜在感染的能力紧密相关。对宿主内部持续感染的结核分枝杆菌的建立，维持和重新激活至关重要的决定因素对宿主内的持续感染很少。人们认为，在肉芽肿病变内封装结核分枝杆菌后，潜伏期会启动，以及对这种环境中存在的特定信号的识别，可抑制有氧呼吸并促进过渡结核分枝杆菌变为非复制持久性生理状态（NRP）。最近，发现包含甲酸氢化酶（FHL）的决定因素（RV0081-RV0088）被发现在暴露于结核分枝杆菌（H.此外，发现该基因座是由两个响应调节剂（DOSR/DEVR和MPRA）直接调节的，已知可以在体外和体内受到结核分枝杆菌的持久性。在没有外部电子受体的情况下，FHL在微粒细胞/厌氧生长期间在肠细菌中的能量代谢中起重要作用。该提案试图通过研究生理上相关条件下的结核分枝杆菌代谢的各个方面来填补我们所知的当前空白。我们假设结核分枝杆菌合成了一种功能性FHL酶复合物，该酶复合物是NRP期间能量代谢和生存所必需的。为了解决这一假设，已经提出了两个具体目标。首先，将使用基于分光光度计和气体色谱法的方法来检查结核分枝杆菌FHL介导甲酸盐氧化为CO2和H2的能力。其次，将检查包含预测FHL的决定因素，以确定它们是否相互作用以形成FHL，并且在NRP期间需要结核分枝杆菌的存活所必需。总的来说，这些研究预计将在与NRP和潜伏期相关的条件下对结核分枝杆菌生理的新颖见解。在NRP期间，当细菌拒绝当前可用的抗生素时，在NRP期间，甲酸代谢基础的描述还可以鉴定出可用于结核分枝杆菌的治疗干预的新酶。公共卫生相关性：本提案中描述的实验将调查FHL甲酸盐代谢在非复制持久性过程中结核分枝杆菌生理学中的重要性。预计包含该酶复合物的决定因素的检查将在与持续感染有关的条件下进一步了解结核分枝杆菌代谢，从而填补我们对结核分枝杆菌生理学的了解的关键空白。该提案的完成可能会导致对新酶的鉴定，这些酶可以针对潜伏期期间结核分枝杆菌的治疗干预，而细菌否则可以抑制当前可用的抗生素。