Oxygen Activation by Mononuclear Copper(I) Active Sites

单核铜 (I) 活性位点的氧活化

• 批准号: 10556191
• 负责人: Wesley John Transue
• 金额: $ 2.91万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-02-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10556191
• 关键词: Active Sites; Address; Amines; Behavior; Binding; Bioinorganic Chemistry; Biological; Biological Process; Catalysis; Cations; Chemistry; Chronic Disease; Clinical; Communities; Copper; Coupled; Dangerousness; Data; Detection; Dioxygen; Discipline; Distant; Dopamine; Electron Spin Resonance Spectroscopy; Electron Transport; Electrons; Environment; Enzymes; Equilibrium; Freezing; Geometry; Goals; Health; Histidine; Human; Hydrogen; Hydrogen Peroxide; Immersion; Industrialization; Invertebrates; Investigation; Ions; Lead; Life; Ligands; Light; Lytic; Measures; Mediating; Methane; Methane hydroxylase; Methanol; Methionine; Methods; Mixed Function Oxygenases; Monitor; Mononuclear; N-terminal; Neurotransmitters; Outcome; Oxidants; Oxidases; Oxidative Stress; Oxides; Oxygen; Particulate; Pattern; Peroxidases; Peroxides; Polysaccharides; Process; Publishing; Raman Spectrum Analysis; Reaction; Regulation; Reporting; Roentgen Rays; Role; Site; Site-Directed Mutagenesis; Spectrum Analysis; Starch; Structure; Suggestion; Sulfur; Techniques; Testing; Thermodynamics; Training; Triplet Multiple Birth; Tyramine; Virulence; Virulence Factors; absorption; circular magnetic dichroism; cold temperature; computer studies; density; electronic structure; enzyme activity; enzyme mechanism; experience; experimental study; greenhouse gases; insight; instrumentation; man; neurochemistry; oxidation; oxidative damage; pathogen; spectroscopic data; theories

Project Summary/Abstract. (From Original Proposal) The interactions of coupled binuclear copper sites and dioxygen have been extensively studied in contexts crucial to life. This contrasts with analogous interactions between mononuclear copper sites and dioxygen, which are less well understood despite their importance in clinical, biological, and industrial settings. As such, furthering our understanding of these enzymes' mechanisms of action would have far reaching implications across a wide set of disciplines. This project focuses on mechanistic elucidation of the oxidative behaviors of three di erent monooxyge- nases featuring single copper active sites: tyramine -monooxygenase (T M), lytic polysaccharide monooxygenase (LPMO), and particulate methane monooxygenase (pMMO). The rst, T M, is closely related to human dopamine -monooxygenase (D M) and participates in invertebrate neurotransmitter regulation. As a noncoupled binuclear copper enzyme, it features two copper centers separated by 11 A, only one of which engages with O2. Out- standing questions on the timing of O2 binding to the reduced enzyme, hydrogen atom abstraction (HAA), and intermetallic electron transfer persist. The second enzyme, LPMO, has large industrial applications in renewable biofuels and has also been implicated as a virulence factor in several pathogens. Its active site comprises a single copper ion coordinated to two histidine residues and the N -terminal amine in a rare \histidine brace" geometry. Computational results inform the planned experiments, suggesting several mechanistic possibilities; though, some recent reports have questioned the role of O2 in favor of H2O2. The last enzyme, pMMO, is important in con- version of methane, a dangerous greenhouse gas, into methanol, a renewable fuel. It has long been thought to possess a coupled binuclear copper active site, but has recently been reappraised to have a mononuclear copper site also engaged in a histidine brace structural motif. This new suggestion means there is little mechanistic insight available, though it draws parallels between pMMO and LPMO. The project ultimately aims to shed light on how these enzymes oxidize their substrates, and to uncover useful and generalizable structure-function relations to be exploited in further clinical and industrial applications. Our speci c aims involve investigation of each class of enzyme using a battery of spectroscopies to uncover informative intermediates, capitalizing on the extensive instrumentation and experience available in the Solomon lab. As many of these transformations involve para- magnetic species, electron paramagnetic resonance (EPR) and magnetic circular dichroism (MCD) experiments will allow direct interrogation of the copper center, particularly in combination with rapid freeze quench (RFQ) techniques. Additionally, stopped ow absorption, resonance Raman (rR), X-ray absorption (XAS), and X-ray emission (XES) will be heavily employed, especially when studying diamagnetic states. All of these studies will be supported by thorough computational investigations using density functional theory (DFT) methods, which will allow for further insight into the electronic structures and reaction energies. The training plan involves immersion in these spectroscopic techniques and in the eld of bioinorganic chemistry, all of which are new to the applicant.

项目摘要/摘要。 （来自原始建议） 耦合双核铜位点和二恶英的相互作用已在上下文中进行了广泛的研究 生命。这与单核铜位点和二氧化物之间的类似相互作用形成对比，这较少 尽管它们在临床，生物学和工业环境中都很重要，但他们的理解很广。因此，进一步 了解这些酶的作用机制将在一系列广泛的含义 学科。该项目的重点是对三个不同单元的氧化行为的机械阐明 - 具有单铜主动部位的Nases：酪胺 - 单氧酶（T M），裂解多糖单加氧酶 （LPMO）和颗粒甲烷单加氧酶（PMMO）。 rST T M与人类多巴胺密切相关 - 糖酶（D M），并参与无脊椎动物神经递质调节。作为非偶联的双核 它具有两个铜中心，分别为11 A，其中只有一个与O2互动。出去- 关于O2时间与减少酶，氢原子抽象（HAA）和 金属间电子传输持续存在。第二种酶LPMO在可再生 生物燃料，也被认为是几种病原体中的毒力因子。它的主动站点包括一个 铜离子在稀有的\组氨酸支撑几何形状中与两个组氨酸残基和N-末端胺配位。 计算结果为计划的实验提供了信息，提出了几种机械可能性。不过，有些 最近的报告质疑O2对H2O2的作用。最后的酶，PMMO，对 甲烷的版本是一种危险的温室气体，可将甲醇（可再生燃料）甲醇。长期以来一直认为 具有耦合的双核铜活性位点，但最近已重新评估以具有单核铜位点 还参与了组氨酸支撑结构基序。这个新建议意味着几乎没有机械洞察力 可用，尽管它在PMMO和LPMO之间取得了相似之处。该项目最终旨在阐明 这些酶如何氧化其底物，并发现有用且可推广的结构功能关系 在进一步的临床和工业应用中被利用。我们的特定目的涉及每个班级的调查 使用一系列光谱镜来揭示信息中间体的酶，并利用了广泛的中间体 所罗门实验室可用的仪器和经验。由于许多这些转变都涉及 磁性物种，电子顺磁共振（EPR）和磁性圆形二色性（MCD）实验 将允许直接询问铜中心，特别是与快速冷冻淬火（RFQ）结合使用 技术。此外，停止的OW吸收，共振拉曼（RR），X射线吸收（XAS）和X射线 发射（XES）将被大量使用，尤其是在研究磁磁状态时。所有这些研究将是 使用密度功能理论（DFT）方法的彻底计算研究支持，这将 可以进一步深入了解电子结构和反应能。培训计划涉及沉浸 在这些光谱技术和生物无机化学的ELD中，所有这些都对申请人来说都是新的。