Abstract
Since the soluble form of IFNβ dramatically inhibits the growth of bacterial hosts, the recombinant pharmaceutical IFNβ was produced as an inclusion body in vitro to overcome this problem. In this study, for the production of IFNβ, pTXB1 was used as the expression vector, which was infused with intein (INT) tag and chitin binding domain (CBD) for increasing the expression of the soluble form of IFNβ. For this, the IFNβ encoding gene was amplified using specific primers and then cloned into the NdeI and SapI sites of the pTXB1 vector. Next, the new construct (pTX-IFNβ) was transformed into ER2566 competent cells and the expression of the fusion protein was assessed by SDS-PAGE and western blotting techniques. After sonication of the pellets in an appropriate lysis buffer, supernatant and pellets were analyzed to determine the solubility of IFNβ. The expressed IFNβ was mostly observed as inclusion bodies, and the solubilization did not increase by decreasing the induction temperature. The expression of IFNβ fused with intein tag was confirmed by western blot using anti-CBD. In addition, the soluble form of IFNβ fusion protein was determined to be about 35%, using the SDS-PAGE technique. In conclusion, the expression of IFNβ fused with INT and CBD can be a suitable strategy for the soluble expression of this pharmaceutical protein. In addition, the auto-cleavage characteristic of INT induced by thiol agent, temperature or pH can be used to increase the protein solubilization in the process of producing IFNβ protein.
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We would like to thank the research council of Malek-Ashtar University of Technology for the financial support of this investigation.
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Seidmoradei, R., Zeinoddini, M., Saeedinia, A.R. et al. Intein-Mediated Fusion Soluble Expression of Recombinant Human Interferon Beta (rhIFN-β). Iran J Sci Technol Trans Sci 44, 371–377 (2020). https://doi.org/10.1007/s40995-020-00844-0
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DOI: https://doi.org/10.1007/s40995-020-00844-0