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

作　　者：王郅媛[1,2] 李赤霞 张萌 王友升[1,4] WANG Zhiyuan;LI Chixia;ZHANG Meng;WANG Yousheng(Beijing Advanced Innovation Center for Food Nutrition and Human Health/School of Light Industry, Beijing Technology & Business University, Beijing 100048, China;Beijing Scientific Instruments and Equipment Cooperation Service Center, Beijing 100048, China;Shandong KEEPFIT Biotech Co, Rizhao 276800, China;Rizhao HUAWEI Institute of Comprehensive Health Industries, Rizhao 276800, China)

机构地区：[1]北京工商大学北京食品营养与人类健康高精尖创新中心/轻工科学技术学院,北京100048 [2]北京科学仪器装备协作服务中心,北京100048 [3]山东凯普菲特生物科技有限公司,山东日照276800 [4]日照华伟大健康产业研究院,山东日照276800

出　　处：《食品科学技术学报》2020年第5期85-90,共6页Journal of Food Science and Technology

基　　金：国家自然科学基金面上项目(31972127);北京市教委科技发展重点项目(KZ201910011013)。

摘　　要：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.

关 键 词：PDE9A 外源表达 分离纯化 HPLC 酶活特性分析 

分 类 号：TS201.2[轻工技术与工程—食品科学] Q556[轻工技术与工程—食品科学与工程]