Functional Synthetic Models of Cu-dependent Monooxygenases

铜依赖性单加氧酶的功能合成模型

• 批准号: 10229556
• 负责人: Isaac Garcia-Bosch
• 金额: $ 37.1万
• 依托单位: CARNEGIE-MELLON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10229556
• 关键词: Biochemical; Biochemistry; Biological; Complex; Copper; Enzymes; Equation; Generations; Hydrogen Bonding; Hydroxylation; Ions; Kinetics; Ligands; Lytic; Metals; Methane; Methane hydroxylase; Methods; Mixed Function Oxygenases; Modeling; Modification; Mononuclear; Oxidants; Oxidation-Reduction; Oxides; Particulate; Polysaccharides; Property; Protocols documentation; Reaction; Research Project Grants; Research Proposals; Role; Spectrum Analysis; Structure; experimental study; interest; metalloenzyme; oxidation; protonation; scaffold

Project Summary: In this research project, we develop synthetic inorganic copper complexes to understand the fundamental aspects of structure and function in Cu-dependent monooxygenase enzymes. These metalloenzymes contain 1 or 2 Cu ions in their active center and they couple the reduction of O2 with the oxidation of substrates via formation of transient Cun/O2 species. We are particularly interested in studying the reactivity of mononuclear Cu/O2 intermediates since they have been proposed as active oxidants in the hydroxylation of strong C-H bonds in enzymes such as particulate methane monooxygenases (pMMOs) and lytic polysaccharide monooxygenases (LPMOs). Many questions concerning the identity of the active Cu/O2 species remain unanswered, including: i) oxidation state of Cu (CuI vs. CuII vs. CuIII); ii) reduction/protonation state of O2 (O2−,(H)O22−, (H)O2−) and the pKa and redox potentials associated with these Cu/O2 species; iii) mechanism by which the Cu/O2 intermediates carry out C-H hydroxylations (e.g. O-O cleavage mechanism before or after C-H oxidation?; generation of high-valent Cu-oxyl species before substrate hydroxylation?). In this research proposal, we tackle this problem using two different approaches: 1) We utilize ligand scaffolds (L) that contain C-H substrates covalently attached to their structure (substrate- ligands) that permit us to generate and characterize LCu/O2 species and evaluate their reactivity towards intramolecular C-H hydroxylation. Substrate-ligand modifications will permit us to: i) evaluate the ability of the Cu/O2 species to oxidize sp3 C-H bonds and sp2 C-H bonds; ii) control the stereo-electronic properties of the Cu complexes by the use of different ligand donors (i.e. N2, N3, N4) that will lead to the generation of mononuclear and dinuclear LCu/O2 species, and analyze their reactivity towards intramolecular C-H hydroxylation including characterization of reaction intermediates, kinetics and computations; iii) utilize this approach (Cu-directed hydroxylations) to develop synthetic protocols to promote challenging organic transformations such as enantioselective C-H hydroxylations and one-pot synthesis of 1,3-oxazines. 2) We synthesize mononuclear Cu complexes bearing redox-active ligands with tunable H-bonds that stabilize Cu-hydroxo and Cu-oxyl cores. These unusual Cu complexes are able to reach multiple oxidation states via oxidation of the metal and/or ligand scaffold. These high-valent CuO(H) cores will be characterized by various spectroscopic methods and their ability to perform biorelevant intermolecular 2e− C-H hydroxylations will be examined systematically using the Bordwell equation (i.e. species with higher redox potential and higher pKa should be capable of oxidizing stronger C-H bonds), kinetic experiments and analysis of the reactions products derived from hydroxylation (e.g. organic product(s) and oxidation/protonation state of the final Cu complexes). Overall, these studies will contribute to a broader understanding of the biochemical role of Cu ions involved in O2 reduction and biologically relevant oxidations.

项目摘要： 在该研究项目中，我们开发了合成无机铜复合物来了解基本 结构和功能的方面Cu依赖性单加氧酶酶。这些金属酶含有 1或2个铜离子在其活性中心，将O2的还原与底物的氧化相结合 瞬态CUN/O2物种的形成。我们对研究单核的反应性特别感兴趣 Cu/O2中间体由于已被提出为强氧化物中的活性氧化物 酶的键，例如特定的甲烷单加氧酶（PMMOS）和裂解多糖 单加氧酶（LPMO）。有关活性Cu/O2物种的身份的许多问题仍然存在 未得到答复，包括：i）Cu的氧化状态（Cui vs. Cuii vs. Cuiii）； ii）O2的还原/质子化状态 （O2-，（H）O22-，（H）O2-）以及与这些Cu/O2物种相关的PKA和氧化还原电位； iii）机制 CU/O2中间体进行C-H羟基化（例如O-O裂解机构之前或之后 C-H氧化？底物羟基化之前产生高价值的Cu-氧基？）。在这项研究中 提案，我们使用两种不同的方法解决了这个问题： 1）我们利用配体支架（L），其中包含与其结构共同附着的C-H底物（底物 - 配体），使我们能够生成和表征LCU/O2物种并评估它们对 分子内C-H羟基化。底物配体修改将使我们能够：i）评估 Cu/o2物种与氧化物SP3 C-H键和SP2 C-H键； ii）控制该立体电子特性 CU复合物通过使用不同的配体供体（即N2，N3，N4），这将导致产生 单核和双核LCU/O2种，并分析其对分子内C-H的反应性 羟基化包括反应中间体，动力学和计算的表征； iii）利用这一点 方法（CU指导的羟基化）以开发合成方案以促进有机挑战 诸如对映选择性的C-H羟基化和1,3-恶阵的一锅合成之类的转换。 2）我们合成具有可调h键的氧化还原活性配体的单核Cu复合物 Cu-Hydroxo和Cu-oxyl核。这些不寻常的Cu复合物能够通过 金属和/或配体支架的氧化。这些高价值CuO（H）核将以各种特征 光谱法及其执行生物含量分子间2E-C-H羟基的能力将是 使用Bordwell方程进行系统检查（即具有较高氧化还原电位和较高PKA的物种 应该能够氧化强C-H键），动力学实验和反应产物的分析 源自羟基化（例如有机产物（S）和最终Cu络合物的氧化/质子化状态）。 总体而言，这些研究将有助于更广泛地了解参与参与的Cu离子的生化作用 O2还原和生物学相关的氧化。