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: 7923548
• 负责人: Stephen J. Lippard
• 金额: $ 5.74万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7923548
• 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; 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 complex; 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）：这项研究的长期目标是提高我们对多组分单氧酶的理解，这些单氧酶激活二氧化合物的选择性羟基化碳氢化合物的选择性羟基化。这些显着的酶系统通常由羟化酶，还原酶和调节蛋白组成，可消耗四种底物 - 碳氢化合物，O2，电子和质子 - 生产酒精和水。该家族的旗舰成员是可溶性甲烷单加掺杂酶（SMMO），该酶在其羟化酶成分的活性位点具有羧酸桥桥接的非血红素二氨基中心，其中可以实现甲烷对甲醇的选择性氧化。从植物到哺乳动物的生物体中发生的类似单元也激活了O2以执行各种相关功能。 X射线晶体学和NMR光谱法研究了单个成分蛋白及其之间的复合物的结构，即三个细菌系统，SMMO，甲苯/O-二氧酶和苯酚羟化酶的结构。将通过快速混合和冻结方法（包括一种新型的亚MS技术）来研究天然和突变羟基酶的O2激活，结合了光学吸收，共振拉曼，IR，EPR，EPR，EXAFS和Mossbauer光谱。动力学分离的中间体与底物的反应将进行研究，以揭示导致烷烃，烷烃，体育烷和杂原子取代底物的立体特异性羟基化的机制。将确定激活参数和动力学同位素效应，以与理论得出的值进行比较，以测试提出的机械途径。还将研究还原酶或Rieske成分蛋白与羟化酶之间的电子转移和质子转移反应。将准备和表征羟化酶二铁中心的小分子类似物。将研究通过与O2反应（LL）模型复合物产生的中间体的结构和性能，并研究其氧化束缚或外源底物的能力。新开发的制备途径将为羧酸桥桥梁的二元中心的Fe-Fe载体提供N-Donors Syn的仿生配合物。该项目与公共卫生有关，因为细菌单加氧酶可以防止Chu（一种温室气体）到达大气，在地下水中降解氯化碳氢化合物，并被激活以进行油溢油的生物修复。对本研究提供的O2激活和底物羟基化的分子机制的了解将促进环境去污染的新策略以及催化剂的发展以将甲烷转化为甲醇。