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（由申请人提供）：铁对于细菌病原体的生长至关重要，这是由于血红素和铁硫（FE-S）簇中的铁所需的。为了对抗感染，人类宿主使用两种利用这种铁依赖性的一般策略：铁固存在，以最大程度地减少生物利用铁，从而通过产生氧化应激，以使病原体的铁金属蛋白的病原体和攻击。我们的长期目标是表征细菌病原体使用的遗传和生化系统，以在压力期间保留细胞内铁稳态。该提案的目的是确定在铁饥饿和氧化应激期间，SUF途径使用的生化机制。 Sufabcdse操纵子在细菌中被激活，以在暴露于氧化应激和铁饥饿时构建必需的Fe-S簇。 SUF途径在许多细菌病原体（例如志贺氏菌和结核分枝杆菌）中是保守的。 Shigella由于细菌痢疾而导致每年1100万人死亡，其中大多数是五岁以下的儿童。 SUF操纵子对志贺氏菌发病机理可能很重要，因为它被志贺氏菌进入其致病生命周期的细胞内阶段。结核分枝杆菌是结核病的病因，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的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，SUFC，SUFD和SUFE的功能尚不完全清楚。我们的目的是（1）表征硫从Sufs转移到Fe-S群集组装的最终目的地的分步路径； （2）确定SUF操纵子中Fe-S群集组件的位置； （3）确定在Fe-S群集组装过程中，SUFC ATPase活性在多蛋白SUFBCD复合物中的功能。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方面的方法来实现我们在便利模型生物体大肠杆菌中的目标。外行陈述：细菌铁稳态的破坏是哺乳动物细胞在感染过程中限制细菌生长的关键机制，因为铁是许多致病细菌的必不可少的营养素。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措施破坏细菌铁稳态的策略。我们的建议旨在表征SUF途径，以便我们可以针对SUF蛋白设计特定的抑制剂，该蛋白将充当新的抗生素