糖氧甲基转移酶MycE催化麦新米星VI甲基转移机理的理论研究

O-linked methylation of sugar substituents is a common modification in the biosynthesis of macrolide antibiotics and other natural products, and is catalyzed by sugar O-methyltransferases. Mycinamicins, a potent macrolide antibiotics, and the first methylation is catalyzed by mycinamicin VI 2’’-O-methyltransferase (MycE). This project intends to employ the quantum chemistry and the hybrid quantum mechanical/molecular mechanical (QM/MM) approach to study the methyl transfer mechanism of mycinamicin VI catalyzed by sugar O-methyltransferase MycE, by building a reasonable model of mycinamicin VI in complex with MycE, which will be investigated in detail to illustrate the substrate binding, the methylation reaction mechanism under part of the enzyme residues and under the whole enzyme environment. There are two major tasks in this project. (i) To determine the reactant complex, transition state structure, determine the proton transfer and methyl transfer proceed through a concerted mechanism or stepwise in the enzymatic methylation, and obtain a complete methylation mechanism; (ii) To clarify interplay of amino acid residues and metal ions in the catalytic mechanism. This project may provide theoretical basis for designing and modifying the existing antibiotics biosynthetic pathways, designing biomimetic catalysts and studying the reaction mechanism catalyzed by the sugar O-methyltransferase family.

取代糖的氧甲基化是大环内酯类抗生素和其它天然产物生物合成途径的常见修饰，且由糖O-甲基转移酶催化。麦新米星（mycinamicins）是一种大环内酯类抗生素，它的第一步甲基化由麦新米星VI 2’’-O-甲基转移酶（MycE）催化完成。本项目拟利用量子化学和混合量子力学/分子力学(QM/MM)方法，构建合理的麦新米星VI与MycE的复合物模型，详细研究底物的结合模式、部份酶残基及整个酶环境下麦新米星VI甲基化过程，探索糖氧甲基转移酶MycE催化麦星米星VI甲基转移的机理。本项目拟解决两大难题：一是确定酶催化甲基化过程的反应复合物、过渡态结构以及质子转移和甲基转移是协同还是分步过程，获取完整的甲基化机理；二是阐明反应中心氨基酸残基和金属离子对催化机理的影响。本项目的研究将为设计并修改麦新米星及其它类似抗生素的现有生物合成途径、设计仿生催化剂和研究糖O-甲基转移酶家族的催化反应机制提供理论基础。

取代糖的氧甲基化是大环内酯类抗生素和其它天然产物生物合成途径的常见修饰。本项目利用DFT理论和QM/MM方法探讨了糖氧甲基转移酶MycE催化麦星米星VI甲基转移的过程。通过建立MycE的简化活性中心模型，用B3LYP方法明确了甲基转移和质子转移不能是分步过程，只能是协同过程，金属Mg2+的作用是定位甲基受体、加速质子转移和稳定氧负离子过渡态。通过建立MycE-SAM-Mg2+-mycinamicin VI 四元复合物结构，用QM/MM (SCC-DFTB/CHARMM)分子动力学方法模拟了底物与酶的结合情况，证实了His278可作为广义碱夺取底物中羟基的H原子，最低能量途径（MEP）及平均力势（PMF）明确了甲基转移和质子转移是协同过程，在近过渡态时质子转移已先于甲基转移完成。通过将Mg2+替换成Ca2+ 和Zn2+后的DFT研究，明确了不同金属离子对MycE酶活性的影响规律。本项目的研究将为设计并修改麦新米星及其它类似抗生素的现有生物合成途径、设计仿生催化剂和研究糖O-甲基转移酶家族的催化反应机制提供理论基础。