畜牧兽医学报2018，Vol.49Issue(1)：75-82,8.DOI:10.11843/j.issn.0366-6964.2018.01.009
铁蛋白Ferritin原核表达和纯化及纳米颗粒胞外自组装
Prokaryotic Expression and Purification of Ferritin and Nano-particles Self-assembling in vitro
摘要
Abstract
This study aimed to express Helicobacter Pylori nonheme iron-containing Ferritin protein in Escherichia coli,and obtain the Ferritin nano-particles with natural nanostructure,for the application in disease and antigen presentation study by surface modification.To improve the expression of Ferritin,Ferritin gene was synthesized according to the Escherichia coli biased codon and cloned to the pET-32a vector.The recombinant protein was purified by affinity chromatography using His-trap Ni2+ column,and refold by gradient dialysis.The refolding protein was detected by transmission electron microscope to identify the nano-structure of the self-assembling protein.The results showed that the prokaryotic expression vector pET-32a-Ferritin was successfully constructed,and the recombinant His-Ferritin protein was expressed after induced by IPTG.The recombinant Ferritin protein was mainly expressed in the form of inclusion body.By use of Histrap Ni2+ affinity chromatography column,we obtained the recombinant Ferritin protein with 96％ purity.The purified protein was in gradient dialysis to 2 mol · L-1 urea buffer with a desalting column to obtain the refold protein.The refold protein had a nanostructure with the diameter of 12-20 nm under the transmission electron microscope,which was similar to the natural nanoparticles.In this study,the prokaryotic expression system of Ferritin was successfully established and the recombinant protein with nanostructure was prepared by induction expression,affinity chromatography purification and refolding.This study lays the foundation for the application of Ferritin in biomedical field.关键词
铁蛋白/原核表达/亲和层析/胞外自组装Key words
Ferritin/prokaryotic expression/affinity chromatography/self-assembling in vitro分类
生物科学引用本文复制引用
李志鹏,刘福航,崔奎青,石德顺,刘庆友..铁蛋白Ferritin原核表达和纯化及纳米颗粒胞外自组装[J].畜牧兽医学报,2018,49(1):75-82,8.基金项目
国家863重点项目(2013AA102504) （2013AA102504）
广西科技开发项目(桂科攻1598013-2) （桂科攻1598013-2）
广西研究生教育创新计划项目(YCBZ2017004) （YCBZ2017004）
