Role of NadD in Mycobacterium tuberculosis proteostasis

NadD 在结核分枝杆菌蛋白质稳态中的作用

• 批准号: 10341221
• 负责人: Benjamin S Gold
• 金额: $ 21.19万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-04 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10341221
• 关键词: Acidity; Active Sites; Address; Adenine Nucleotides; Anabolism; Anti-Infective Agents; Antibiotic Therapy; Antibiotics; Antimycobacterial Agents; Antioxidants; Biochemical; Biochemical Genetics; Biochemistry; Biological; Biology; Cause of Death; Chemicals; Chemistry; Collection; Complex; Cyclic Peptides; Development; Drug Targeting; Drug resistance; Enzymes; Etiology; Face; Generations; Genetic; Genetic Transcription; Goals; Green Fluorescent Proteins; Heat Stress Disorders; Homologous Gene; Human; Hypoxia; Immune; Immunity; Impairment; In Vitro; Infection; Infectious Agent; Intoxication; Lead; Luciferases; Measures; Mediating; Mediator of activation protein; Metabolic; Metals; Methods; Molecular Chaperones; Monitor; Mycobacterium tuberculosis; NADP; Niacinamide; Nitrogen; Nucleotide Biosynthesis; Oxygen; Pathogenesis; Pathway interactions; Patients; Peptide Hydrolases; Peptides; Pharmaceutical Preparations; Phenotype; Play; Proteins; Proteolysis; Proteome; Publishing; Reaction; Reactive Nitrogen Species; Reactive Oxygen Species; Recombinants; Reporting; Research; Resistance; Ribosomes; Role; Starvation; Stress; System; Testing; Tuberculosis; Virulence; antioxidant enzyme; chemical genetics; cofactor; drug discovery; drug-sensitive; genetic approach; inhibitor; interest; macrophage; member; mouse model; multicatalytic endopeptidase complex; nicotinate; non-genetic; novel; oxidation; prevent; programs; protein aggregation; proteostasis; proteotoxicity; pyridine nucleotide; repaired; small molecule; tool; tuberculosis treatment

Summary The goal of this research program is to characterize the biology of NadD (nicotinate adenylyltransferase) from Mycbobacterium tuberculosis (Mtb), the etiological agent of tuberculosis (TB). During infection, Mtb faces a diverse set of host microenvironments and host chemistries that can lead to proteotoxic stress, including hypoxia, mild acidity, metal starvation or intoxication, and reactive oxygen and nitrogen species. Antibiotic treatment of TB patients can damage Mtb proteins by stalling nascent proteins on the ribosome or via the formation of reactive oxygen or nitrogen intermediates. Many of these host stresses are associated with preventing or slowing of Mtb replication and the development of non-genetic, phenotypic drug resistance. Mtb maintains protein integrity with the proteostasis network – a collection of over one hundred proteins, including chaperones and their cofactors (ClpB, DnaK, DnaJ1, DnaJ2, GrpE, GroEL/ES) and barrel-shaped proteases (ClpP1P2 and Mtb20S). The transcription of many members of the proteostasis network is induced by proteotoxic stresses such as heat. Genetic and chemical disruption of components of the proteostasis network is associated with decreased virulence in macrophage and murine models of TB. We sought to identify compounds targeting components of the proteostasis network by using a chemical screen for compounds whose activity was potentiated by a proteotoxic stress generated by heat. Our characterization of one heat-stress potentiated compound unexpectedly revealed NadD as the target. NadD is a high priority drug target that lies at a critical intersection of de novo and salvage pathways for NAD(H) and NADP(H) biosynthesis. Chemical inhibition of NadD in wild-type Mtb led to an increase in intrabacterial protein aggregation. We hypothesize that loss of NadD disrupts the function of NADPH-dependent antioxidant defense systems and/or NadD plays a role as an unconventional protein chaperone. This research takes two complementary directions: in Aim 1, we will use genetics and chemical-biology to characterize NadD’s role in the proteostasis network, in particular by monitoring generation of intrabacterial reactive oxygen and nitrogen species, protein carbonylation, and protein aggregation; and in Aim 2, we will define the mechanism by which NadD contributes to Mtb proteostasis. These studies will lay the groundwork to understand how NadD contributes as a noncanonical member of the proteostasis network during infection and antibiotic treatment.

概括 该研究项目的目标是表征 NadD（烟酸腺苷酸转移酶）的生物学特性 结核分枝杆菌 (Mtb) 是结核病 (TB) 的病原体。在感染过程中，Mtb 面临着以下问题。 不同的宿主微环境和宿主化学物质可能导致蛋白毒性应激，包括缺氧， 弱酸性、金属饥饿或中毒以及活性氧和氮的抗生素治疗。 结核病患者可以通过阻止核糖体上的新生蛋白或通过形成反应性抗原来破坏结核分枝杆菌蛋白。 许多宿主应激与预防或减缓结核分枝杆菌有关。 Mtb 的复制和非遗传性、表型耐药性的发展维持了蛋白质的完整性。 蛋白质稳态网络——一百多种蛋白质的集合，包括伴侣及其辅助因子 （ClpB、DnaK、DnaJ1、DnaJ2、GrpE、GroEL/ES）和桶形蛋白酶（ClpP1P2 和 Mtb20S）。 蛋白质稳态网络许多成员的转录是由蛋白质毒性应激（例如热）诱导的。 蛋白质稳态网络成分的遗传和化学破坏与蛋白质稳态下降有关 我们试图鉴定针对结核病成分的化合物。 通过使用化学筛选化合物来建立蛋白质稳态网络，其活性通过 热产生的蛋白毒性应激。我们对一种热应激增强化合物的表征。 出人意料地发现 NadD 是一个高度优先的药物靶点，位于关键的交叉点。 野生型中 NAD(H) 和 NADP(H) 生物合成的从头途径和挽救途径。 Mtb 导致细菌内蛋白质聚集增加。 NADPH 依赖性抗氧化防御系统和/或 NadD 的功能发挥着非常规作用 这项研究有两个互补的方向：在目标 1 中，我们将使用遗传学和 化学生物学来表征 NadD 在蛋白质稳态网络中的作用，特别是通过监测生成 细菌内活性氧和氮物种、蛋白质羰基化和蛋白质聚集； 目标 2，我们将确定 NadD 促进 Mtb 蛋白质稳态的机制。这些研究将奠定基础。 为理解 NadD 作为蛋白质稳态网络的非规范成员如何做出贡献奠定了基础 感染和抗生素治疗。

项目成果

Editorial: Drug-resistant Mycobacterium tuberculosis.

社论：耐药结核分枝杆菌。

• 发表时间: 2023
• 作者: Boshoff, Helena I M;Warner, Digby F;Gold, Ben
• 通讯作者: Gold, Ben