Characterization of the Suf Fe-S Cluster Biosynthesis Pathway Under Stress

胁迫下 Suf Fe-S 簇生物合成途径的表征

• 批准号: 7614249
• 负责人: Franklin Wayne Outten
• 金额: $ 21.72万
• 依托单位: UNIVERSITY OF SOUTH CAROLINA AT COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-02 至 2012-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7614249
• 关键词: ATP phosphohydrolase; ATP-Binding Cassette Transporters; Age; Anabolism; Animal Model; Antibiotics; Bacillary Dysentery; Bacteria; Bacterial Genome; Bioavailable; Biochemical; Biogenesis; Bioinorganic Chemistry; Cessation of life; Child; Complex; Dependence; Destinations; Enzymes; Escherichia; Escherichia coli; Exposure to; Generations; Genes; Genetic; Genus Mycobacterium; Goals; Growth; Heme Iron; Homeostasis; Homologous Gene; Host Defense; Human; In Vitro; Infection; Iron; Mammalian Cell; Metalloproteins; Methods; Microbial Genetics; Molecular; Molecular Biology; Molecular Chaperones; Mycobacterium tuberculosis; Nutrient; Operon; Oxidative Stress; Pathogenesis; Pathway interactions; Positioning Attribute; Process; Protein Chemistry; Proteins; Research Personnel; Resistance; Role; Route; Scaffolding Protein; Shigella; Site; Staging; Starvation; Stress; Sulfur; System; Testing; Tuberculosis; Work; combat; cysteine desulfurase; design; experience; in vivo; inhibitor/antagonist; novel; pathogen; pathogenic bacteria; phosphogluconate dehydratase; programs; reconstitution; scaffold

DESCRIPTION (provided by applicant): Iron is critical for growth of bacterial pathogens due to the need for iron in heme and iron-sulfur (Fe-S) clusters. To combat infection, the human host uses two general strategies that exploit this iron dependence: iron sequestration to minimize bioavailable iron for the pathogen and attack of the pathogen's iron metalloproteins via generation of oxidative stress. Our long-term goal is to characterize the genetic and biochemical systems utilized by bacterial pathogens to preserve intracellular iron homeostasis during stress. The objective of this proposal is to determine the biochemical mechanisms used by the Suf pathway to build Fe-S clusters during iron starvation and oxidative stress. The sufABCDSE operon is activated in bacteria to build essential Fe-S clusters during exposure to oxidative stress and iron starvation. The Suf pathway is conserved in many bacterial pathogens such as Shigella and Mycobacterium tuberculosis. Shigella is responsible for the deaths of 11 million people each year due to bacillary dysentery, the majority of which are children under the age of five. The suf operon may be important for Shigella pathogenesis since it is transcribed as Shigella enters the intracellular stage of its pathogenic lifecycle. M. tuberculosis is the causative agent of tuberculosis and directly causes 2 million deaths each year. In M. tuberculosis, the suf genes are essential since deletion of the suf genes is lethal in M. tuberculosis and related Mycobacteria. Despite its importance, the molecular details of in vivo Suf function are still unclear. The SufS enzyme is a cysteine desulfurase that provides sulfur for Fe-S cluster assembly, while the functions of SufA, SufB, SufC, SufD, and SufE are not fully known. Our Aims are to (1) characterize the step-by-step path of sulfur transfer from SufS to its ultimate destination for Fe-S cluster assembly; (2) identify the site(s) of Fe-S cluster assembly in the Suf operon; and (3) determine the function of SufC ATPase activity in the multi-protein SufBCD complex during Fe-S cluster assembly. We will use methods in protein chemistry, bioinorganic chemistry, molecular biology, and microbial genetics to accomplish our Aims in the facile model organism Escherichia coli. Lay Statement: Disruption of bacterial iron homeostasis is a key mechanism used by mammalian cells to limit bacterial growth during infection because iron is an essential nutrient for many pathogenic bacteria. We believe that targeting the Suf Fe-S cluster biogenesis pathway with novel antibiotics could be a strategy for assisting the host defenses in disrupting bacterial iron homeostasis. Our proposal is designed to characterize the Suf pathway so that we might design specific inhibitors against the Suf proteins that will act as a new class of antibiotics

描述(申请人提供)：铁是细菌病原体生长的关键，因为需要在血红素和铁-硫磺簇(铁-S)中铁。为了对抗感染，人类宿主使用两种常见的策略来利用这种铁依赖：铁隔离以最大限度地减少病原体的生物可用铁，以及通过产生氧化应激攻击病原体的铁金属蛋白。我们的长期目标是确定细菌病原体在应激过程中利用的遗传和生化系统，以保持细胞内铁的稳态。这项建议的目的是确定在铁饥饿和氧化应激过程中，SuF途径建立Fe-S簇所使用的生化机制。SufABCDSE操纵子在细菌中被激活，在暴露于氧化应激和铁饥饿期间建立必要的铁-S簇。Suf途径在许多细菌病原体中是保守的，如志贺氏菌和结核分枝杆菌。志贺氏菌每年导致1100万人死于细菌性痢疾，其中大部分是5岁以下的儿童。Suf操纵子可能在志贺氏菌致病过程中起重要作用，因为它是随着志贺氏菌进入其致病生命周期的细胞内阶段而转录的。结核分枝杆菌是结核病的病原体，每年直接导致200万人死亡。在结核分枝杆菌中，suf基因是必不可少的，因为suf基因的缺失在结核分枝杆菌和相关分枝杆菌中是致命的。尽管它很重要，但体内Suf功能的分子细节仍不清楚。SufS酶是一种半胱氨酸脱硫酶，为Fe-S簇组装提供硫，而SufA、SufB、SuFc、SufD和Sufe的功能尚不完全清楚。我们的目标是(1)表征硫从SuFS到最终目的地的逐步转移路径，以实现Fe-S团簇组装；(2)确定Fe-S团簇组装在SuF操纵子中的位置(S)；以及(3)确定SUFC ATPase活性在Fe-S团簇组装过程中的作用。我们将使用蛋白质化学、生物无机化学、分子生物学和微生物遗传学的方法来实现我们在简易模式生物大肠杆菌中的目标。Lay声明：破坏细菌铁稳态是哺乳动物细胞在感染期间限制细菌生长的关键机制，因为铁是许多致病细菌的基本营养物质。我们认为，用新型抗生素靶向Suf Fe-S簇生物发生途径可能是帮助宿主防御破坏细菌铁稳态的一种策略。我们的建议旨在描述Suf途径的特征，以便我们可以针对Suf蛋白设计特定的抑制剂，作为一种新的抗生素类别。