"双酶一体"连续催化体系的建立及其研究

两种酶共同作用以催化两步反应过程。双酶催化的优点是第一个酶催化后的产物可被第二个酶作为底物催化，这样可更高效率地催化反应物。本项目拟建立并研究"双酶一体"催化体系。拟研究的模式双酶体系为D-氨基酸氧化酶/过氧化氢酶，过氧化氢酶/葡萄糖氧化酶，淀粉酶/糖化酶。在细胞外通过内含子自我催化，以短链肽把两种酶紧密连接为一体，这是本项目的创新之处。拟研究的酶连接过程不需要其它酶和化学催化剂催化。由于是在细胞外通过自我剪接的方法把两种酶连接起来，因而本项目有别于通过基因融合在细胞内表达融合蛋白的方法。本项目将比较研究"双酶一体"体系与两个酶混合使用的催化效率。研究酶连接效率及其影响因素、"双酶一体"与底物的作用机理。期望本项目研究的酶连接方法能够应用于"多酶一体"体系的构建。

Two enzymes work together for the catalysis in two-step reactions. In these bioreactions, the product of one enzyme is the substrate of the other. This project will establish and study the two-enzyme systems. The model two-enzyme systems include D-Amino acid oxidase/catalase,glucose oxidase/catalase, and alpha-amylase/glucoamylase. Enzyme splicing will be carried out though the self catalysis of intein, which is carried out outside the cells. This is the novelty of the project. The methodology of establishing two-enzyme systems is different from that by gene fusion for the expression of hybrid proteins. This project will study the enzyme splicing efficiency, the interaction of the enzymes of the two-enzyme with the substrates. It is expected that the developed methodology of establishing two-enzyme systems can be extended to the establishment of multi-enzyme systems.