中文摘要：

在蛋白晶体结构难以获得的情况下,通过设计突变体来获取6-羟基-3-琥珀酰吡啶单加氧酶Hsp B的结构信息。首先获取Hsp B蛋白的同源序列并进行比对,之后对Hsp B蛋白进行同源建模和从头建模,并与底物2,5-二羟基吡啶（HSP）进行对接模拟;通过分子模拟、序列比对和参考同源蛋白晶体三种方式,设计并构建Hsp B酶的25个突变体;通过突变体的表达纯化和酶动力学常数测定来研究Hsp B的结构性质。根据实验结果,推测FAD的正确结合在稳定Hsp B蛋白结构中具有重要的作用,同时推测底物HSP和辅酶NADH处于同一活性中心并与不同位点相互作用。吡啶衍生物是极具工业价值的化合物,生物催化法是合成吡啶衍生物的有效途径,而吡啶衍生物的生物催化研究较少,通过考察突变体的性质,推测了Hsp B的部分结构信息,为此类吡啶单加氧酶的工业改造和应用奠定了基础。

英文摘要：

Pyridine derivatives are the important value-added chemicals,and biocatalysis is a potential technology for the industrial synthesis of pyridine derivatives. The structure information of 6-hydroxy-3-succinoylpyridine monooxygenase（ Hsp B） by mutant construction was investigated. The space structure of Hsp B has been built through computer modeling and docked with its substrate HSP（ 2,5-dihydroxy-pyridine）. Then 25 mutations of Hsp B were constructed and studied,according to molecular simulation,sequence alignment,and homologous crystal references. All the mutants have been expressed and purified,and the kinetic parameters of soluble mutants have been measured. According to the properties of mutants,it can infer that the correct binding of FAD possesses important roles in protein stability,and moreover,substrate HSP,and co-enzyme NADH live in the same activity center in HSP but interact with different amino acids. The structure information will help us for the industrial application of pyridine monooxygenase.