Nonheme Diiron Centers and the Biological Oxidation of Hydrocarbons

非血红素二铁中心和碳氢化合物的生物氧化

• 批准号: 7280453
• 负责人: Stephen J. Lippard
• 金额: $ 47.51万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-01-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7280453
• 关键词: Active Sites; Address; Alcohols; Alkanes; Alkenes; Amines; Amino Acids; Area; Binding; Biological; Biomimetics; Bioremediations; Chemistry; Chlorinated Hydrocarbons; Complex; Coupling; Decontamination; Development; Dioxygen; Electron Transport; Electrons; Enzymes; Family member; Freezing; Future; Genetic Structures; Goals; Heme; Homologous Protein; Human Resources; Hydrocarbons; Hydrogen Peroxide; Hydroxylation; Individual; Investigation; Isotopes; Kinetics; Knowledge; Learning; Mammals; Measures; Methane; Methane hydroxylase; Methanol; Methodology; Methods; Mixed Function Oxygenases; Modeling; Molecular; Mono-S; Mossbauer Spectroscopy; Movement; NADH; NMR Spectroscopy; Nature; Oils; Optics; Organism; Oxidation-Reduction; Oxidoreductase; Oxygenases; Pathway interactions; Peroxides; Phenol 2-monooxygenase; Plants; Preparation; Process; Property; Proteins; Protons; Public Health; Quantum Mechanics; Range; Reaction; Research; Research Activity; Research Personnel; Rest; Roentgen Rays; Role; Route; Shunt Device; Site; Structure; Sulfides; System; Techniques; Testing; Water; Work; X ray diffraction analysis; X-Ray Crystallography; X-Ray Diffraction; absorption; analog; carboxylate; catalyst; chemical property; design; genetic regulatory protein; greenhouse gases; ground water; insight; macromolecule; molecular mechanics; mutant; novel; novel strategies; oxidation; planetary Atmosphere; prevent; programs; protein structure; small molecule; stoichiometry; toluene 2-xylene monooxygenase; vector

DESCRIPTION (provided by applicant): The long-term goal of this research is to advance our understanding of multicomponent mono-oxygenases that activate dioxygen for the selective hydroxylation of hydrocarbons. These remarkable enzyme systems, which typically consist of hydroxylase, reductase, and regulatory proteins, consume four substrates - a hydrocarbon, O2, electrons, and protons - to produce alcohol and water. The flagship member of the family is soluble methane monooxygenase (sMMO), which has a carboxylate-bridged non-heme diiron center at the active site of its hydroxylase component where selective oxidation of methane to methanol is achieved. Similar units that occur in organisms ranging from plants to mammals activate O2 to perform a variety of related functions. Structures of the individual component proteins, and of complexes between them, from three bacterial systems, sMMO, toluene/o-xylene monooxygenase, and phenol hydroxylase, will be investigated by X-ray crystallography and NMR spectroscopy. O2 activation in native and mutant hydroxylases will be studied by rapid-mixing and freeze-quench methodologies, including a novel sub-ms technique, combined with optical absorption, resonance Raman, IR, EPR, EXAFS, and Mossbauer spectroscopy. Reactions of kinetically isolated intermediates with substrates will be investigated to reveal mechanisms responsible for stereospecific hydroxylation of alkanes, alkenes, arenes, and heteroatom-substituted substrates. Activation parameters and kinetic isotope effects will be determined for comparison with theoretically derived values to test proposed mechanistic pathways. Electron-transfer and proton-translocation reactions between the reductase or Rieske component proteins and the hydroxylases will be investigated. Small molecule analogs of the hydroxylase diiron centers will be prepared and characterized. The structures and properties of intermediates generated by reacting diiron(ll) model complexes with O2 and their ability to oxidize tethered or exogenous substrates will be studied. A newly developed preparative route will afford biomimetic complexes with N-donors syn to the Fe-Fe vector of the carboxylate-bridged dimetallic center. This project is relevant to public health because bacterial monooxygenases prevent ChU, a greenhouse gas, from reaching the atmosphere, degrade chlorinated hydrocarbons in ground water, and are activated for bioremediation of oil spills. Knowledge of the molecular mechanisms of O2 activation and substrate hydroxylation provided by this research will advance novel strategies for environmental decontamination and the development of catalysts to convert methane to methanol.

描述（由申请人提供）：本研究的长期目标是促进我们对多组分单加氧酶的理解，该酶可激活双氧以进行烃的选择性羟基化。这些非凡的酶系统通常由羟化酶、还原酶和调节蛋白组成，消耗四种底物--碳氢化合物、氧气、电子和质子--产生酒精和水。该家族的旗舰成员是可溶性甲烷单加氧酶（sMMO），其在其羟化酶组分的活性位点处具有羧酸根桥接的非血红素二铁中心，其中实现甲烷选择性氧化为甲醇。从植物到哺乳动物的生物体中存在的类似单位激活O2以执行各种相关功能。单个组分蛋白质的结构，以及它们之间的复合物，从三个细菌系统，sMMO，甲苯/邻二甲苯单加氧酶，苯酚羟化酶，将通过X-射线晶体学和NMR光谱研究。天然和突变的羟化酶中的O2活化将通过快速混合和冷冻淬灭方法进行研究，包括一种新的亚ms技术，结合光学吸收，共振拉曼，IR，EPR，EXAFS和穆斯堡尔光谱。将研究动力学分离的中间体与底物的反应，以揭示负责烷烃，烯烃，芳烃和杂原子取代的底物的立体定向羟基化的机制。将确定活化参数和动力学同位素效应，以便与理论推导值进行比较，以测试拟议的机制途径。还原酶或Rieske组分蛋白质和羟化酶之间的电子转移和质子易位反应将被研究。羟化酶二铁中心的小分子类似物将被制备和表征。将研究通过使二铁（II）模型络合物与O2反应产生的中间体的结构和性质以及它们氧化拴系或外源底物的能力。一种新开发的制备路线将提供仿生配合物与N-供体syn的Fe-Fe载体的羧酸根桥接的双金属中心。该项目与公共卫生有关，因为细菌单加氧酶可防止ChU（一种温室气体）进入大气，降解地下水中的氯化烃，并被激活用于石油泄漏的生物修复。本研究提供的氧气活化和底物羟基化的分子机制的知识将推进环境净化和催化剂的发展，以将甲烷转化为甲醇的新策略。