Functional Synthetic Models of Cu-dependent Monooxygenases

项目摘要： 在这个研究项目中，我们开发合成无机铜配合物，以了解基本的 铜依赖性单加氧酶的结构和功能方面。这些金属酶含有 1或2个Cu离子在其活性中心，它们耦合O2的还原与底物的氧化， 瞬时Cun/O2物种的形成。我们特别感兴趣的是研究单核细胞的反应性， Cu/O2中间体，因为它们已被提议作为强C-H羟基化的活性氧化剂 酶如颗粒甲烷单加氧酶（pMMO）和溶解性多糖中的键 单加氧酶（LPMO）。关于活性Cu/O2物种的身份仍然存在许多问题 未回答，包括：i）Cu的氧化态（CuI vs. CuII vs. CuIII）; ii）O2的还原/质子化态 (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中间体通过其进行C-H羟基化（例如，O-O裂解机理在C-H羟基化之前或之后 C-H氧化？;在底物羟基化之前产生高价Cu-氧基物质？）。本研究 根据我们的建议，我们采用两种不同的方法来解决这个问题： 1)我们利用配体支架（L），所述配体支架含有共价连接至其结构的C-H底物（底物- 配体），使我们能够产生和表征LCu/O2物种，并评估其对 分子内C-H羟基化。底物-配体修饰将允许我们：i）评估底物-配体修饰的能力。 Cu/O2物种氧化sp3 C-H键和sp2 C-H键; ii）控制化合物的立体电子性质。 通过使用不同的配体供体（即N2、N3、N4），这将导致生成 单核和双核LCu/O2物种，并分析其对分子内C-H的反应性 羟基化，包括反应中间体的表征、动力学和计算; iii）利用该羟基化， 方法（Cu定向羟基化），以开发合成方案，促进具有挑战性的有机 这些方法包括对映体选择性C-H羟基化和1，3-恶嗪的一锅法合成。 2)我们合成单核铜配合物轴承氧化还原活性配体与可调的H-键， Cu-羟基和Cu-氧基核。这些不寻常的Cu络合物能够通过以下方式达到多个氧化态： 金属和/或配体支架的氧化。这些高价CuO（H）核的特征在于各种不同的结构。 光谱方法及其进行生物相关的分子间2 e − C-H羟基化的能力将是 使用Bordwell方程（即具有较高氧化还原电位和较高pKa的物质）系统地检查 应该能够氧化更强的C-H键），动力学实验和反应产物的分析 衍生自羟基化（例如有机产物和最终Cu络合物的氧化/质子化状态）。 总的来说，这些研究将有助于更广泛地了解铜离子的生物化学作用， O2还原和生物相关氧化。

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.