苦瓜籽MAP30的聚乙二醇化学修饰条件优化及纯化研究  

Modification conditions optimization and purification of PEGylated MAP30 from Semen Momordicae Charantiae

作　　者：张永明[1] 刘盛邦[1] 黄凯[1] 聂宇[1] 孟延发[1] 

机构地区：[1]四川大学生命科学学院生物资源与生态环境教育部重点实验室,四川成都610064

出　　处：《中草药》2008年第11期1640-1643,共4页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金资助项目(C01020404)

摘　　要：目的探讨聚乙二醇修饰苦瓜籽核糖体失活蛋白(MAP30)的反应条件以及修饰产物的纯化方法。方法在不同条件下,将聚乙二醇活性酯(mPEG)2-NHS与MAP30反应,用SDS-PAGE分析不同反应条件对产物成分的影响;采用Sephacryl S-300HR分子筛柱色谱法对修饰产物进行分离纯化,TNBS法测定目标产物PEG-MAP30的氨基修饰度。结果聚乙二醇修饰MAP30的最优反应条件为4℃,pH8.5硼酸硼砂缓冲液,MAP30质量浓度1mg/mL,MAP30与(mPEG)2-NHS质量比1∶10为最佳修饰条件,5min内反应完全;反应产物进一步纯化,经SDS-PAGE检测目标产物PEG-MAP30中不含未修饰的MAP30;TNBS法测定MAP30的氨基修饰度为44.9%。结论初步确定了聚乙二醇修饰MAP30的反应条件和修饰产物的纯化方法。Objective To explore conditions for the modification of MAP30 from Semen Momordica Charantiae with polyethylene glycol （PEG） and the method for the purification of PEG-MAP30. Methods To analyze the influence of various conditions on tile components of PEG-MAP30 by SDS-PAGE, purify PEG MAP30 by Sephacryl S-300 HR molecular sieve column chromatography, and detect the modification rate of amino by TNBS under the optimal reaction condition with the reaction of （mPEG）2-NHS and MAP30. Results Temperature 4℃, boric acid borax buffer with pH 8.5, concentration of MAP30 1 mg/mL, the mass ratio of MAP30 to （mPEG）2-NHS （1 ： 10）, and, 5 rain for reaction time were selected as the optimal reaction condition. Molecular sieve column showed a good purification effect. The modification rate of amino was 44.9% tested by TNBS under the optimal reaction condition. Conclusion The factors influencing the modification of MAP30 with PEG are determined, and a purification method for PEG-MAP30 is developed.

关 键 词：苦瓜籽 核糖体失活蛋白(MAP30) 聚乙二醇 化学修饰 

分 类 号：R284.2[医药卫生—中药学] R286.02[医药卫生—中医学]