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

• Zixuan Liu ^[1] ; Jizhou Zhang ^[1] ; Hongqiong Fan ^[2] ; Ruofeng Yin ^[3] ; Zhong Zheng ^[1] ; Qian Xu ^[1] ; Qing Liu ^[1] ; Haiting He ^[1] ; Xiaofan Peng ^[1] ; XinXin Wang ^[1] ; Xiaokun Li ^[1] ; Yechen Xiao ^[1]
  1. [1] Jilin University

     Jilin University

     China

     
  2. [2] First Hospital of Jilin University

     First Hospital of Jilin University

     China

     
  3. [3] China-Japan Union Hospital of Jilin University Department of Orthopaedic Surgery

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• Localización: Electronic Journal of Biotechnology, ISSN-e 0717-3458, Vol. 18, Nº. 4, 2015, págs. 302-306
• Idioma: inglés
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  • Background 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 bacteria were induced by 0.5 mM Isopropyl-ß-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.