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Updated: Jun 24, 2026
A Convenient and General Expression Platform for the Production of Secreted Proteins from Human Cells
Published on: July 31, 2012
[Expression, purification and refolding of streptavidin-tagged human tumor necrosis factor-alpha fusion protein]
Cui-Xiang Xu1, Zhi-Ming Hu, Jing-Long Li
1Institute of Biological Treatment, School of Biotechnology, Southern Medical University, Guangzhou, China. xucuixiang1129@163.com
Objective:
To study the purification, refolding and bioactivity of streptavidin-tagged human tumor necrosis factor-alpha (SA-TNF-alpha) bi-functional fusion protein.
Methods:
SA-TNF-alpha fusion protein was expressed in BL21(DE3) host bacteria, purified using Ni-NTA affinity chromatography and refolded by dilution and dialysis followed by identification using Western blotting. The effect of SA-TNF-alpha fusion protein against L929 cells was evaluated by MTT assay. Flow cytometry was used to analyze the surface modification of biotinylated MB49 tumor cells by SA-TNF-alpha fusion protein.
Results:
Recombinant SA- TNF-alpha fusion protein was expressed in BL21(DE3) at about 30% of total bacterial protein, with a purity of about 95% after purification. The SA-TNF-alpha fusion protein existed as dimmers, tetramers and higher order structures after refolding. The fusion protein exhibited a bi-functionality by inhibiting L929 cells and SA-mediated high-affinity binding to biotinylated cell surfaces, with an anchor modification rate of above 90%.
Conclusion:
The dimmers, tetramers and higher order structures of the obtained SA-TNF-alpha fusion protein all exhibit a bi-functionality, and may serve as a potential candidate therapeutic agent for tumors.
