Expression and purification of soluble single-chain Fv against human fibroblast growth factor receptor 3 fused with Sumo tag in Escherichia coli

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Abstract

Background: Overexpression or mutated activation of Fibroblast growth factor receptor 3 (FGFR3) is involved in the pathogenesis ofmany tumors.More and more studies focus on the potential usage of therapeutic antibodies against FGFR3. Results: In this study, a novel single-chain Fv (ScFv) against FGFR3 was prepared and characterized. To achieve the soluble expression, ScFv was fused with Sumo (Small ubiquitin-related modifier) by polymerase chain reaction (PCR), and cloned into pET-20b. The recombinant bacteria were induced by 0.5 mM Isopropyl-β-D-thiogalactopyranoside (IPTG) for 16 h at 20°C, and the supernatant liquid of Sumo-ScFv was harvested and purified by Ni-NTA chromatography. After being cleaved by the Sumo protease, the recombinant ScFv was released from the fusion protein, and further purified by Ni-NTA chromatography. The purity of ScFv was shown to be higher than 95% and their yield reached 4 mg per liter of bacterial culture. In vitro data showed that ScFv can significantly attenuate FGF9-induced phosphorylation of FGFR3. Conclusion: We provide a novel method to produce soluble expression and bioactive functions of ScFv in Escherichia coli.

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Liu, Z., Zhang, J., Fan, H., Yin, R., Zheng, Z., Xu, Q., … Xiao, Y. (2015). Expression and purification of soluble single-chain Fv against human fibroblast growth factor receptor 3 fused with Sumo tag in Escherichia coli. Electronic Journal of Biotechnology, 18(4), 302–306. https://doi.org/10.1016/j.ejbt.2015.05.006

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