与芦丁、槲皮素相比，异槲皮素在多个方面具有更好的生物活性和生物利用度。α-L-鼠李糖苷酶是转化芦丁制备异槲皮素的关键酶。课题首次以极端嗜热菌T. petrophila 的α-L-鼠李糖苷酶基因为研究对象，通过结晶条件优化，制备蛋白晶体及酶蛋白质与底物共结晶的复合物；通过结构解析，阐明结构与功能关系，阐明酶催化特异性及不能有效水解芦丁等含鼠李糖苷的黄酮化合物的分子机理；利用基因工程技术定点突变酶基因，并以该基因和黑曲霉α-L-究鼠李糖苷酶基因为亲本，重组催化结构域，构建同时具有嗜热和高效催化芦丁的鼠李糖苷酶。研究将填补极端嗜热鼠李糖苷酶晶体结构的空白，将首次建立该酶的催化特性、结构与功能的关系等理论，有望能获得新型嗜热α-L-鼠李糖苷酶突变基因及其基因工程菌。结果将为进一步改造该酶的催化性能和挖掘酶的新用途提供理论和方法的指导，将为异槲皮素的生物制备及其在食品、医药等领域中的应用奠定基础。

Compared with rutin and quercetin, isoquercitrin showed better bioactivity and bioavailability. Alpha-L-rhamnosidase is the key enzyme for the biotransformation from rutin to isoquercitrin.. Through the preparation of protein crystal and analysis of the structure, α-L-rhamnosidase from Thermotoga petrophila was firstly used as the research object in this project to clarify the relationship between the structure and function of the enzyme, the catalytic specificity of the thermophilic α-L-rhamnosidase and the molecular mechanism for the inefficiency in the catalysis of flavonoids containing rhamnoside (rutin et. al). Based on experimental data, the α-L-rhamnosidase from T. petrophila is site-directed modified by using genetic engineering technology. To construct a novel α-L-rhamnosidase with thermostability and high rutin-hydrolytic efficiency, α-L-rhamnosidase from T. petrophila and Aspergillus niger were used as the parent gene, and the catalytic domain will be recombined. . This study filled the blank in the crystal structure of thermophilic α-L-rhamnosidase. The theory about the catalytic properties and structure-activity relationship of the enzyme will be build. It’s expected that novel α-L-rhamnosidase mutants and related recombinant stains will be obtained. The results will provide theoretical basis and guidance for the deeply research, modification and application of α-L-rhamnosidases. In addition, the results will lay the foundation for bio-preparation of isoquercitrin and accelerate its application in food and medicine.