Expression, purification and preliminary activity study of recombinant hepatocyte growth factor protein in E.coli
10.3760/cma.j.issn.0254-5101.2012.11.011
- VernacularTitle:肝细胞生长因子的表达、纯化和基本活性研究
- Author:
Xiaojiao ZHENG
;
Zhou GAO
;
Rongrong SHEN
;
Hang ZHAO
;
Dong CEN
;
Jianping LUO
;
Jianxin Lü
;
Renzhi PEI
;
Shixuan HUA
- Publication Type:Journal Article
- Keywords:
Hepatocyte growth factor;
Protein;
Recombinant;
Purification;
Activity
- From:
Chinese Journal of Microbiology and Immunology
2012;(11):967-971
- CountryChina
- Language:Chinese
-
Abstract:
Objective To prepare hepatocyte growth factor(HGF) recombinant protein and confirm its activity preliminarily according to building HGF gene prokaryotic expression vector and transforming into E.coli.Methods Clone HGF inserted into the vector pET-26b(+) to construct prokaryotic expression vector pET-26b(+)-HGF and transform into E.coli Rosseta(DE3).The transformed bacteria induced by IPTG was purified through Ni-NTA resin affinity chromatography frozen-drying after renaturation.Results HGF gene recombinant prokaryotic expression vector pET-26b(+)-HGF was constructed successfully.E.coli Rosseta(DE3) which was transformed into pET-26b(+)-HGF expresses the target protein as the form of inclusion bodies,accounting for 38％ of the total bacterial proteins,and confirmed by Western blot.HGF protein which was purified by Ni-NTA resin affinity chromatography,has a purity of about 95％,and can promote proliferation,migration,and inhibition of apoptosis for human non-small cell lung cancer cell line A549 cells after interaction.Conclusion HGF gene recombinant prokaryotic expression vector pET-26b (+)-HGF was constructed and expressed in transformed E.coli Rosseta(DE3) successfully.They resumed their recombinant HGF protein structure after purification and renaturation,and had biological activity confirmed by in vitro studies.
