文献类型：期刊文献

中文题名：重组人内皮抑素层析复性的相关研究

英文题名：On-Column Refolding and Purification of Recombinant Human Endostatin from Inclusion Bodies Expressed in Escherichia coli

作者：乔路[1];夏杰[1];陆兵[1];徐殿胜[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2008

卷号：34

期号：2

起止页码：203

中文期刊名：华东理工大学学报（自然科学版）

外文期刊名：Journal of East China University of Science and Technology

收录：CSTPCD;;EI(收录号：20081911245036);Scopus;北大核心:【北大核心2004】；CSCD:【CSCD2011_2012】；

语种：中文

中文关键词：重组人内皮抑素;包涵体;柱复性;层析;纯化

外文关键词：recombinant human endostatin; inclusion body; on-column refolding; chromatography; purification

摘要：研究了由大肠杆菌表达的重组人内皮抑素(Recombinant human endostatin,rhES)的复性、纯化过程。比较先层析复性后纯化(工艺1)和先纯化后层析复性(工艺2)两种工艺的结果表明:工艺1所得rhES的收率和活性回收率分别为64.44%和213.19%,与工艺2相比分别提高8.70%和33.04%。工艺1中,研究了3种凝胶过滤柱复性及复性pH和上样蛋白浓度对复性的影响,并优化相关工艺。结果表明:在pH9.0,rhES浓度3.91 mg/mL的条件下,采用双梯度凝胶过滤Sephacryl S-100 HR进行层析复性,Sephadex G-25脱盐后,用SP Sepharose FF阳离子交换吸附纯化,获得活性回收率为213.19%,HPLC纯度为97.07%的rhES,其IC50为1.58μg/mL。
In this paper, the procedures for on column refolding after and prior to purification of recombinant human endostatin（rhES） expressed in Escherichia coli were described. In the procedure of purification after on-column refolding,the yield and activity recovery of rhES were 64.44% and 213.19%, compared with those in the procedure of on-column refolding after purification increased by 8. 70% and 33.04%, respectively. Three gel exclusion chromatography refolding methods were compared. The effects of pH of refolding buffer and protein loading concentration were investigated and purification method was also optimized. A dual gradient Sephacryl S-100 HR SEC refolding method at pH9.0 with 3.91 mg/mL protein loading was established. After desalting with Sephadex G-25, the refolded rhES was purified by SP Sepharose FF. The purity of rhES was 97.07% analyzed by HPLC, and the activity recovery approached to 213.19%. The concentration at half-maximum inhibition （IC50） for rhES was approximately 1.58 μg/ mL.