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

Zixuan Liu, Jizhou Zhang, Hongqiong Fan, Ruofeng Yin, Zhong Zheng, Qian Xu, Qing Liu, Haiting He, Xiaofan Peng, XinXin Wang, Xiaokun Li, Yechen Xiao
2015 Electronic Journal of Biotechnology  
Overexpression or mutated activation of Fibroblast growth factor receptor 3 (FGFR3) is involved in the pathogenesis of many 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
more » ... 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.
doi:10.1016/j.ejbt.2015.05.006 fatcat:n4btluxbzzbodox4rtgrbl7fsy