Optimization of Fermentation Conditions for Reteplase Expression by Escherichia Coli Using Response Surface Methodology



Zare H¹ ; Sadeghi HMM¹ ; Akbari V¹
Source: Avicenna Journal of Medical Biotechnology Published:2019

Abstract

Background: Expression of heterologous proteins at large scale is often a challenging job due to plasmid instability, accumulation of acetate and oxidative damage in bio-reactors. Therefore, it is necessary to optimize parameters influencing cell growth and expression of recombinant protein. Methods: In the present study, the optimal culture conditions for expression of retep-lase by Escherichia coli (E. coli) BL21 (DE3) in a bench-top bioreactor was determined. Response Surface Methodology (RSM) based on Box-Behnken design was used to evaluate the effect of three variables (i.e., temperature, shaking speed and pH) and their interactions with cellular growth and protein production. The obtained data were analyzed by Design Expert software. Results: Based on results of 15 experiments, a response surface quadratic model was developed which was used to explain the relation between production of reteplase and three investigated variables. The high value of R-Squared (0.9894) and F-value of 51.99 confirmed the accuracy of this model. According to the developed model, the optimum fermentation conditions for reteplase expression were temperature of 32°C, shaking speed of 210 rpm, and pH of 8.4. This predicted condition was applied for the production of reteplase in the bioreactor leading to a protein yield of 188 mg/l. Conclusion: Our results indicate the significant role of culture conditions (e.g., pH, temperature and oxygen supply) in protein expression at large scale and confirm the need for optimization. The proposed strategy here can also be applied to experimental set-up of optimization for fermentation of other proteins. © 2019, Avicenna Journal of Medical Biotechnology. All rights reserved.