Integrated bioprocessing and genetic strategies to enhance soluble expression of anti-HER2 immunotoxin in E. Coli

Abstract

Abstract Immunotoxins are widely applied for cancer therapy. However, bacterial expression of immunotoxins usually leads to the formation of insoluble and non-functional recombinant proteins. This study was aimed to improve soluble expression of a novel anti-HER2 immunotoxin under the regulation of the trc promoter in Escherichia coli by optimization of the cultivation conditions using response surface methodology (RSM). To conduct RSM, four cultivation variables (i.e., inducer concentration, post-induction time, post-induction temperature, and medium recipe), were selected for statistical characterization and optimization using the Box-Behnken design and Design Expert software. Based on the developed model using the Box-Behnken design, the optimal cultivation conditions for soluble expression of anti-HER2 immunotoxin were determined to be 0.1 mM IPTG for induction in the LB medium at 33 °C for 18 h. The expressed immunotoxin was successfully purified using affinity chromatography with more than 90% purity and its bioactivity was confirmed using cell-based ELISA. Technical approach developed in this study can be generally applied to enhance the production yield and quality of recombinant proteins using E. coli as the gene expression system.

Keywords

• Immunotoxin
• Response surface methodology (RSM)
• Cultivation optimization
• Protein solubility
• Escherichia coli