Deciphering the Enzymatic Mechanism of Superoxide Dismutase

破译超氧化物歧化酶的酶促机制

• 批准号: 10418479
• 负责人: Gloria Borgstahl
• 金额: $ 43.14万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10418479
• 关键词: 3-Dimensional; Active Sites; Amino Acids; Antioxidants; Binding; Biological Process; Biology; Bypass; Cardiovascular system; Cells; Charge; Complex; Coupled; Coupling; Crystallization; Crystallography; Data; Data Collection; Dioxygen; Disease; Electron Transport; Electrons; Electrostatics; Environment; Enzymatic Biochemistry; Enzymes; Functional disorder; Future; Goals; Homeostasis; Human; Hydrogen; Hydrogen Peroxide; Investigation; Laboratories; Life; Ligand Binding; Ligands; Malignant Neoplasms; Manganese; Maps; Measures; Metabolic; Metals; Methods; Mitochondria; Mitochondrial Matrix; Modeling; Molecular; Nature; Neurodegenerative Disorders; Neutron Diffraction; Neutrons; Organism; Oxidation-Reduction; Oxidative Stress; Oxides; Oxidoreductase; Oxygen; Pathology; Periodicity; Positioning Attribute; Proteins; Protocols documentation; Protons; Reaction; Reactive Oxygen Species; Research; Research Project Grants; Rest; Role; SOD2 gene; Signal Transduction; Solvents; Spectrum Analysis; Structure; Superoxide Dismutase; Superoxides; Surface; System; Testing; Therapeutic; Therapeutic Intervention; Time; Variant; Water; X-Ray Crystallography; base; computational chemistry; design; experimental study; improved; interest; ionization; manganese oxide; metalloenzyme; novel strategies; prevent; protonation; reaction rate; structural biology; tool; working group

Abstract Superoxide dismutases (SODs) are the major regulators of oxidative stress and therefore the first line of defense to protect organisms against metabolic- and environmentally-induced reactive oxygen species (ROS). Human mitochondrial manganese SOD (MnSOD) expression is modulated to prevent ROS-based damage, promote redox homeostasis, and maintain proper cell signaling. Our research goal is to understand the molecular basis of how MnSOD uses coupled proton-electron transfers to dismute superoxide. For this, the 3D arrangement of all atoms is needed, most importantly the position of protons. Our recent technical advancements with neutron crystallography at Oak Ridge National Laboratory have overcome the limitations of X- ray crystallography – revealing proton positions with high detail while also allowing control of the metal electronic state. In this research project, MnSOD neutron maps will reveal the proton relays to the active site metal and the protonation states of metal- bound ligands. The scientific hypothesis for this study is that MnSOD transfers protons from a small group of water molecules via partially solvent-exposed amino acids to the nearly completely buried manganese for the dismutation of superoxide to hydrogen peroxide and molecular oxygen via cyclic metal redox reactions. The specific aims are to characterize the electron-coupled proton relays of MnSOD by investigating the proton environment of (1) the resting states of the reduced and oxidized manganese active sites, (2) the product inhibited Mn-peroxo complex, and (3) the superoxide bound enzyme. Spectroscopy on crystals will be performed to help design/understand crystallographic experiments, and computational chemistry studies on neutron derived all-atom structures will help tie the results together and test our interpretations about the enzymatic activity. The resulting protocols, methods, and structures will be of specific interest to those in the fields of structural biology, antioxidants, and metallo-enzymology and of interest to biologists in general.

摘要 超氧化物歧化酶（SOD）是氧化应激的主要调节剂， 因此，保护生物体免受代谢和 环境诱导的活性氧（ROS）。人线粒体 锰SOD（MnSOD）表达被调节以防止基于ROS的损伤， 促进氧化还原稳态，并维持适当的细胞信号传导。我们的研究目标是 了解MnSOD如何利用耦合质子电子转移的分子基础 分解超氧化物。为此，需要所有原子的3D排列， 重要的是质子的位置。我们最近在中子方面的技术进步 橡树岭国家实验室的晶体学已经克服了X-射线衍射的局限性， 射线晶体学-揭示质子的位置与高细节，同时也允许 控制金属的电子状态。在这个研究项目中，MnSOD中子图将 揭示了质子传递到活性中心金属和金属的质子化状态- 结合配体。这项研究的科学假设是MnSOD转移质子 从一小群水分子通过部分溶剂暴露的氨基酸到 几乎完全掩埋了锰，使超氧化物歧化为氢 过氧化物和分子氧通过循环金属氧化还原反应。具体目标是 通过研究MnSOD的质子传递特性， 环境（1）还原和氧化的锰活性物质的静止状态 （2）产物抑制Mn-过氧络合物，（3）超氧阴离子结合 酵素将对晶体进行光谱分析，以帮助设计/理解 晶体学实验和中子衍生的计算化学研究 全原子结构将有助于将结果联系在一起，并测试我们对原子结构的解释。 酶活性由此产生的协议、方法和结构将具有特定的 结构生物学、抗氧化剂和金属酶学领域的人感兴趣 也是生物学家普遍感兴趣的。