Assembly and Repair of Biological Iron-Sulfur Clusters

生物铁硫簇的组装和修复

• 批准号: 9315821
• 负责人: MICHAEL K. JOHNSON
• 金额: $ 32.4万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-01-01 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9315821
• 关键词: Active Sites; Address; Aerobic; Aging-Related Process; Anabolism; Anemia; Archaea; Ataxia; Atherosclerosis; Bacteria; Biochemical; Biogenesis; Biological; Biological Assay; Biophysics; Carbon; Carrier Proteins; Chloroplasts; Circular Dichroism; Complement; Complex; Coupled; Cyanobacterium; Defect; Disease; Electron Spin Resonance Spectroscopy; Electron Transport; Enzymes; Eukaryota; Fumarates; Goals; Health; Homeostasis; Human; Hydrogen; In Situ; In Vitro; Iron; Iron Overload; Iron-Sulfur Proteins; Life; Ligation; Magnetism; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Metabolism; Mitochondria; Molecular; Molecular Biology; Molecular Biology Techniques; Myopathy; Nature; Neurodegenerative Disorders; Nitrates; Nitrogen; Nitrogen Fixation; Organism; Oxidation-Reduction; Oxidative Stress; Oxygen; Plants; Plastids; Process; Property; Prosthesis; Proteins; Reactive Oxygen Species; Regulation; Respiratory Chain; Role; Scaffolding Protein; Signal Transduction; Site; Specificity; Sulfur; System; TXN gene; Techniques; Temperature; Work; absorption; age related; base; biophysical techniques; circular magnetic dichroism; cysteine desulfurase; design; experimental study; genetic regulatory protein; glutaredoxin; human disease; in vivo; iron deficiency; oxidative damage; pathogenic bacteria; public health relevance; repaired; respiratory; stem; trafficking

DESCRIPTION (provided by applicant): Iron-sulfur clusters are present in more than 300 different types of enzymes or proteins and constitute one of the most ancient, ubiquitous and structurally diverse classes of biological prosthetic groups. However, the most common type of cluster, the cubane-type [Fe4S4] cluster, is particularly sensitive to oxidative degradation. Hence, the process of iron-sulfur biosynthesis and repair is essential to almost all aerobic life forms and is remarkably conserved in prokaryotic and eukaryotic organisms. Three distinct types of iron-sulfur cluster assembly machinery have emerged in bacteria, termed the NIF, ISC and SUF systems, and the ISC and SUF systems form the basis of the eukaryotic mitochondrial and plastid iron-sulfur cluster assembly machineries, respectively. In each case the overall mechanism involves cysteine desulfurase-mediated assembly of transient clusters on scaffold proteins and subsequent transfer of preformed clusters or cluster fragments to apo proteins. However, in no case is the assembly, repair or transfer mechanism understood at the molecular level. The long-term goal of this project is a molecular-level understanding of iron-sulfur cluster biosynthesis and repair using the NIF, ISC and SUF systems, and accessory proteins such as monothiol glutaredoxins and thioredoxin-like Nfu proteins. Elucidating the mechanism of iron-sulfur cluster biosynthesis and repair is central to understanding cellular iron homeostasis and thereby human diseases associated with iron-overload, oxidative stress and defects in the mitochondrial respiratory chain. The approach involves using molecular biology techniques to effect large scale expression and/or site-specific changes in the target enzymes and proteins, biochemical and enzymatic assays, and the application of biophysical spectroscopic techniques (electron paramagnetic resonance, UV-visible absorption, circular dichroism, and magnetically-induced circular dichroism, resonance Raman, M�ssbauer, and mass spectrometry) that can probe the nature, ligation and detailed properties of iron or iron-sulfur centers during cluster biosynthesis, degradation, repair or transfer to acceptor proteins. The objectives are to establish the molecular mechanisms of assembly, degradation, repair, and transfer of iron-sulfur clusters, the specificity of cluster transfer with respect to acceptor proteins, and the means by which iron-sulfur proteins regulate cellular iron homeostasis.

描述（由申请人提供）：铁硫簇存在于 300 多种不同类型的酶或蛋白质中，构成最古老、普遍且结构多样的生物辅基类别之一。立方烷型[Fe4S4]簇对氧化降解特别敏感，因此，铁硫生物合成和修复过程对几乎所有物质都至关重要。细菌中出现了三种不同类型的铁硫簇组装机制，称为 NIF、ISC 和 SUF 系统，ISC 和 SUF 系统构成了真核线粒体的基础。在每种情况下，总体机制分别涉及半胱氨酸脱硫酶介导的支架蛋白上瞬时簇的组装以及随后的转移。然而，在任何情况下都无法在分子水平上理解铁硫簇的组装、修复或转移机制。 使用 NIF、ISC 和 SUF 系统以及辅助蛋白（例如单硫醇谷氧还蛋白和硫氧还蛋白样 Nfu 蛋白）进行生物合成和修复。阐明铁硫簇生物合成和修复的机制对于了解细胞铁稳态以及与此相关的人类疾病至关重要。该方法涉及使用分子生物学技术来影响大规模表达和/或线粒体呼吸链中的铁过载、氧化应激和缺陷。目标酶和蛋白质的位点特异性变化、生化和酶测定以及生物物理光谱技术的应用（电子顺磁共振、紫外可见吸收、圆二色性和磁诱导圆二色性、共振拉曼、穆斯堡尔、和质谱），可以在簇生物合成、降解、修复或转移到受体期间探测铁或铁硫中心的性质、连接和详细特性目标是建立蛋白质。 铁硫簇组装、降解、修复和转移的分子机制，簇转移相对于受体蛋白的特异性，以及铁硫蛋白调节细胞铁稳态的方式。