| 摘要: |
| [目的]建立聚乙二醇修饰蛋白的快速分离纯化方法.[方法]以4种聚乙二醇化药用蛋白为研究对象,采用阳离子交换树脂进行纯化.纯化条件:Hiprep 16/10 CM Sepharose FF(1 mL)预装柱;流动相:A液为20 mmol/L pH4.0的醋酸缓冲液;B液为A液中加入1 mol/L NaCl;B液以0～100%离子强度线性梯度洗脱100 min,流速0.2 mL/min,波长280 nm检测,收集各洗脱峰.[结果]一步纯化可将单点修饰产物与聚乙二醇、多点修饰产物以及未修饰蛋白分离开来,单点修饰产物经SDS-PAGE检测呈单一条带,经质谱检测其相对分子质量与预期相符.[结论]所建立的纯化方法可快速分离纯化聚乙二醇修饰蛋白. |
| 关键词: 聚乙二醇 药用蛋白 分离与纯化 |
| DOI： |
| 分类号: |
| 基金项目:上海市科委重点科技攻关项目，上海-SK研究与发展基金 |
|
| Rapid isolation and purification of pegylated proteins |
|
|
| Abstract: |
| 【Objective】 The study established a rapid isolation and purification method for pegylated proteins. 【Method】 To purify 4 kinds of pegylated pharmaceutical proteins,cation ion exchange chromatography was used.The purification was carried out by using a Hiprep 16/10 CM Sepharose FF (1 mL) column with the mixture of (A) 20 mmol/L pH4.0 sodium acetate buffer and (B) 20 mmol/L pH4.0 sodium acetate buffer containing 1 mol/L NaCl as mobile phase in gradient mode.B buffer was eluted from 0-100% in 100 min,at a flow rate of 0.2 mL/min and the detection wavelength was 280 nm.The eluates were collected and assayed by SDS PAGE.【Result】 The purified monopegylated proteins were achieved from the modified mixture on cation exchange chromatography by one step purification procedure.The monopegylated proteins showed single band on SDS PAGE gel.The molecular weights of 4 kinds of monopegylated proteins were determined by MALDI-TOF mass spectrometry and the results were just as expected. 【Conclusion】 This developed method was suitable for rapid purification of pegylated proteins. |
| Key words: poly(ethylene glycol) pharmaceutical protein isolation and purification |
