Studies on the optimal expression condition, purification and its characterization of ScFv-2F3.
- Author:
Yuan-Ming LUO
1
;
Ying MU
;
Jing-Yan WEI
;
Gang-Lin YAN
;
Gui-Min LUO
Author Information
1. Key Laboratory of Molecular Enzymology and Engineering of Educational Ministry, Jilin University, Changchun 130023, China.
- Publication Type:Journal Article
- MeSH:
Antibodies, Catalytic;
biosynthesis;
chemistry;
genetics;
isolation & purification;
Bioreactors;
microbiology;
Cloning, Molecular;
Escherichia coli;
Gene Expression;
Immunoglobulin Fragments;
biosynthesis;
chemistry;
genetics;
isolation & purification;
Inclusion Bodies;
metabolism;
Protein Folding;
Protein Renaturation;
Recombinant Proteins;
biosynthesis;
chemistry;
genetics;
isolation & purification;
Selenium;
metabolism
- From:
Chinese Journal of Biotechnology
2002;18(1):74-78
- CountryChina
- Language:Chinese
-
Abstract:
The expression vectors of the gene encoding ScFv-2F3 were transformed into E. coli BL21(DE3). Clones of higher expression were first selected, then were grown in the presence of IPTG at 37 degrees C to induce its expression. The culture conditions were carefully optimized. It was found that optimal conditions were as follows: the induction was started as OD590 reached to 1.0-1.8; the concentration of IPTG was 0.3-0.5 mmol/L and induction time is 7 h. The yield of ScFv-2F3 expressed in the selected clones is about 20% of the total proteins. The optimal culture conditions were successfully applied to fermenter of 50 L. The conditions of washing the inclusion bodies were also optimized. A two-step method was used to renature the inclusion body. The expression product of interest and its biological activities were characterized with Western blotting and ELISA. A novel selenium-containing single-chain abzyme with GPX activity was prepared.
