芳环断裂双加氧酶反应机理及结构－功能关系的理论研究

The aromatic ring cleavage is a key process both in the biological degradation of aromatic compounds and in the amino acid metabolism of mammalians. This process is catalyzed by Aromatic Ring-Cleavage Dioxygenase (ARCD) in nature. This project proposes exploring the catalytic mechanism and structure-function relationship of the ARCD by employing the hybrid quantum mechanics/molecular mechanics (QM/MM) approach. The research objective of this proposal includes two enzymes, i.e. homoprotocatechuate 2,3-dioxygenase (HPCD) and protocatechuate 3,4-dioxygenase (3,4-PCD), which have different regioselectivity of ring-cleavage and belong to extradiol and intradiol dioxygenase respectively. The proposed research mainly concerns the mechanism of enzymatic catalysis at the molecular level and structure-function relationship, as well as the modeling of the various spectroscopics found in experiments. The aim of this proposal is to uncover the fundamental factor of the different regioselectivity of the aromatic ring-fission, which can eventually provide insight into the mechanism of these enzymes. These researches will generate a wealth of data on geometries, electronic structures, spin states, and thermochemical properties, which can help to improve the catalytic efficiency of genetically mutated enzymes and their bio-inspired or biomimetic synthetic analogous for the experimental community.

芳环断裂过程是芳香族化合物微生物降解过程中的重要步骤，在动物的氨基酸代谢中也起到重要的作用。自然界中这一过程主要是通过芳环断裂双加氧酶(ARCD酶)来得以实现。本项目拟应用量子力学与分子力学相结合的(QM/MM)方法研究芳环断裂双加氧酶的反应机理及其结构－功能关系。研究对象为同原儿茶酚2,3-双加氧酶(HPCD)和原儿茶酚3，4－双加氧酶(3,4-PCD)，这两种酶在芳环断裂过程中的开环区域选择性不同，分别属于双羟外双加氧酶和双羟内双加氧酶。研究内容主要涉及分子水平的酶催化过程微观机理及其结构－功能关系，还包括活性物种的光谱模拟。研究目标是从微观层次上搞清和理解芳环开环过程中产生不同区域选择性的根本原因，从而对芳环断裂双加氧酶催化的反应机理有新的认识。这些研究将产生大量几何结构、电子结构、自旋态、动力学和热力学数据,及各种光谱数据，能够对该领域中酶和仿酶的改良起积极的推动作用。

芳环断裂双加氧酶催化的芳环断裂反应是芳香族化合物微生物降解过程的关键步骤之一，对维持全球碳素循环起着非常重要的作用。揭示芳环断裂双加氧酶的结构和机理能够为设计和发展仿生催化剂提供帮助。本项目采用混合量子力学/分子力学方法研究了三种芳环断裂双加氧酶，同原儿茶酚2，3-双加氧酶（HPCD）、对氨基苯酚双加氧酶（APD）、尿黑酸1，2-双加氧酶（HGDO）的氧分子活化和催化反应机理。对金属-氧分子加合物的本质、反应的活性氧物种以及催化反应机理进行了详细的讨论。我们的计算研究不仅重复了已有的实验结果（动力学、中间体结构、中间体出现的顺序、反应速率决定步骤、以及光谱性质），还预测了实验未捕捉到的中间体的性质，为进一步的实验工作提供依据。此外，我们还从中得到一个重要的启示：第二配位层和底物在形成高效的酶催化剂中起着至关重要的作用。这将为新型仿生催化剂的设计提供理论依据。