[1]张亭亭,李招发,许瑞安.重组大鼠细胞球蛋白的制备、纯化与活性[J].华侨大学学报(自然科学版),2009,30(6):668-672.[doi:10.11830/ISSN.1000-5013.2009.06.0668]
　ZHANG Ting-ting,LI Zhao-fa,XU Rui-an.Preparation,Purification and Activity of Recombinant Rat Cytoglobin[J].Journal of Huaqiao University(Natural Science),2009,30(6):668-672.[doi:10.11830/ISSN.1000-5013.2009.06.0668]
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重组大鼠细胞球蛋白的制备、纯化与活性()
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《华侨大学学报（自然科学版）》[ISSN:1000-5013/CN:35-1079/N]

卷:
第30卷
期数:
2009年第6期
页码:
668-672
栏目:
出版日期:
2009-11-20

文章信息/Info

Title:
Preparation,Purification and Activity of Recombinant Rat Cytoglobin
文章编号:
1000-5013(2009)06-0668-05
作者:
张亭亭; 李招发; 许瑞安
华侨大学分子药物学研究所
Author(s):
ZHANG Ting-ting;  LI Zhao-fa;  XU Rui-an
Institute of Molecular Medicine, Huaqiao University, Quanzhou 362021, China
关键词:
重组细胞球蛋白;  高效可溶表达;  分离纯化;  鉴定;  活性检测
Keywords:
recombinant CYGB;  soluble high-expression;  isolation and purification;  identification;  activity analysis
分类号:
Q78
DOI:
10.11830/ISSN.1000-5013.2009.06.0668
文献标志码:
A
摘要:
通过聚合酶链式反应(PCR)克隆鼠源细胞球蛋白(Cytoglobin,CYGB)基因,构建原核表达载体pET22b-Cygb,转入E.coliBL21(DE3)经乳糖诱导表达CYGB,发现CYGB主要以可溶形式存在,其表达量占可溶性总蛋白的35%以上.通过DEAE阴离子交换柱层析和Sephacryl S-100凝胶过滤层析,CYGB纯度可超过95%.重组表达的可溶性CYGB不但具有过氧化物酶活性,其活性为(3.23±0.12)mkat.(g.min)-1,还能保护胎鼠原代皮肤成纤维细胞减轻氧化应激损伤(H2O2模型).
Abstract:
Rat Cygb cDNA was cloned through PCR with rAAV-Cygb as template and inserted into a prokaryotic expression vector to construct a recombinant plasmid,pET22b-Cygb.Then the recombinant plasmid pET22b-Cygb was transformed into E.coli BL21(DE3) and expressed by lactose induction.The CYGB protein accounted for more than 35% of total soluble protein.The purified CYGB protein was achieved through DEAE anion-exchange chromatography and Sephacryl S-100 gel filtration chromatography,respectively.Eventually,the purity of CYGB protein was over 95%.After purification,its antioxidation activity was measured through its protection effect to primary skin fibroblast of fetal rat resulted from hydrogen peroxide damage,and its peroxidase activity unit was(3.23±0.12) mkat·(g·min)-1.

参考文献/References:

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备注/Memo

备注/Memo:
国家高新技术研究发展(863)计划项目(2005216060,2008AA022135); 福建省自然科学基金资助项目(2007J0105); 福建省科技厅重点项目(2008Y0084)
更新日期/Last Update: 2014-03-23