Study on Purification and Renaturation of His6-?TNF?Fusion Protein (IB)
- VernacularTitle:人His6-IFN?/TNF?(His6?-TNFN?)融合蛋白(包涵体)的分离、纯化与复性研究
- Author:
Jianfa YU
;
Qing ZHOU
;
Zhizhang MA
- Publication Type:Journal Article
- Keywords:
h?TNF?Human IFN?/TNF?;
fusion protein;
purification and renaturation
- From:
Chinese Journal of Cancer Biotherapy
1996;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
The E. coli BL21(DE3) strain bearing the plasmid(T_7lac/His6-tag) with the h?TNF?recombinant gene was grown in LB medium containing SO?g/ml kanamycin, at 37 ℃ , up to the late logarithmic phase(OD590 ~ 0.5)then induced with IPTG (final concentration lmmol/L)for 5 hours. SDS-PAGE analysis revealed that expression level of the product(His6-?TNF?was up to 45% of the total bacterial proteins and was mainly as insoluble inclusion bodies ( IBs) . After cell lysis, the IBs was separated by centrifugation, dissolved in 7mol/L urea, then purified by Ni column ( Ni~(2+) -Sepharose 6B) . And the purity of more than 95% and the recovery of about 90% were obtained. The purified product was refolded with the renaturation buffer under low temperafure(
