环核苷酸磷酸二酯酶PDE9A的体外表达、纯化与酶活特性分析

Expression,Purification and Enzyme Activity Analysis of Cyclic Nucleotide Phosphodiesterase PDE9A

环核苷酸磷酸二酯酶PDE9A作为研究具有调节血糖等功效的天然活性物质的新型靶点,日益受到关注。通过异丙基硫代半乳糖苷诱导大肠杆菌BL21感受态细胞表达获得PDE9A,并通过Ni-NAT琼脂糖树脂亲和柱、Q-Sepharose离子交换纯化系统和Sephacryl S300分子筛纯化系统进行逐级纯化,最终获得高纯度的PDE9A蛋白,并应用高效液相色谱进行酶活特性检测,证明制备得到的蛋白质具有较高的水解cGMP能力,能够为新型降血糖功能性食品的开发提供依据。

As a novel target of blood glucose regulation,natural active substance cyclic nucleotide phosphodiesterase PDE9A was gradually attracted the attention.The target protein was obtained from E.coli BL21 cells via IPTG-induced expression.Then the protein was gradually purified by nickel column affinity purification system,Q-Sepharose ion exchange purification system and Sephacryl S300 molecular sieve purification system.Finally,the acquired target PDE9A protein with high purity was proved to have high affinity for substrate cGMP digestion,which provided a preliminary basis for the development of novel functional food for reducing blood glucose in the future.