双酶体系合成右旋糖酐过程的聚合机理与调控研究

The aim of the project is to develop dual-enzyme system in which Dextransucrase and Dextranase are used as polymerization catalyst and molecular-weight regulator for clinical dextran production. By analyzing changes of substrate concentration and by-products, the molecular weight of dextran and polymerization degree using high performance gel permeation chromatography (HPGPC), polymerization kinetics including the length of repeating units and copolymerization equation can be obtained. With the use of Dextranase, dextran molecular weight could be effectively controlled during the bio-catalytic process. The effect of Dextranase for producing dextran with a specific molecular weight and distribution is also investigated. Analytical techniques i.e. Fourier transfer infrared (FTIR), Raman and nuclear magnetic resonance (NMR) are used to determine the structure of dextran products. From molecular point of view, the polymerization mechanism of Dextransucrase and the chain scission mechanism of Dextranase can be revealed. In summary, this project aims to investigate the catalytic behaviour of Dual-enzyme bio-synthesis system for directly producing clinical dextran, the studies in exploring enzymatic reactions, dextran polymerization kinetics and chain scission mechanism have a significant potential in providing technical and theoretical support for the production of bio-macromolecule products and carbohydrate-based drugs .

针对目前多糖聚合动力学研究难以深入、酶催化调控多糖聚合机制尚不明确的实际问题，以Dextransucrase合成酶为右旋糖酐聚合催化剂、Dextranase分解酶为聚合度调控剂构建的双酶体系，在生物体外由蔗糖定向合成药用右旋糖酐。通过HPGPC对合成过程底物浓度、副产物浓度及合成产物分子量及聚合度的跟踪检测，获得聚合过程重复单元长度、动力学链长及共聚方程和聚合度公式，开展右旋糖酐生物合成动力学研究；利用Dextranase分解酶调控右旋糖酐聚合，跟踪检测调控过程重复单元长度、动力学链长及聚合方式的变化，研究定向合成过程Dextranase的调控作用及量效关系；通过FTIR、Raman及NMR技术对双酶体系目标产物进行分析，研究定向合成过程Dextransucrase的催化机制及Dextranase的断键规律，探索药用右旋糖酐生物合成的聚合机理，为糖类药物的制备提供技术和理论上的支持。

针对目前多糖聚合动力学研究难以深入、酶催化调控多糖聚合机制尚不明确的实际问题，以Dextransucrase合成酶为右旋糖酐聚合催化剂、Dextranase分解酶为聚合度调控剂构建的双酶体系，在生物体外由蔗糖定向合成药用右旋糖酐。通过对双酶合成过程底物浓度、副产物浓度及合成产物右旋糖酐分子量及聚合度的跟踪检测，探明聚合过程重复单元长度、动力学链长及共聚方程和聚合度变化的规律，阐明右旋糖酐生物合成动力学机制；利用Dextranase分解酶调控右旋糖酐聚合过程，跟踪检测调控过程重复单元长度、动力学链长及聚合度的变化，探明定向合成过程Dextranase分解酶的调控作用及量效关系。基于双酶体系成功构建了药用右旋糖酐定向合成的新途径，实现具有特定分子量分布和高均一度药用右旋糖酐产物的高产率，为糖类聚合物定向制备、糖类药物的开发提供技术和理论上的支持。