Cloning, Expression, and Refolding of PPE17 Protein of Mycobacterium Tuberculosis as a Promising Vaccine Candidate
Journal Title: Iranian Journal of Medical Sciences - Year 2019, Vol 44, Issue 1
Abstract
Background: Tuberculosis as a global health problem requires special attention in the contexts of prevention and control. Subunit vaccines are promising strategies to protect burdens of tuberculosis via increasing the BCG protection. The present study aimed to design a vaccine study by means of high-throughput expression and correct refolding of recombinant protein PPE17. Methods: We aimed to clone, express, and refold PPE17 protein of Mycobacterium tuberculosis in bacterial systems as a promising vaccine candidate. The PPE17 (Rv1168c) gene was PCR amplified and inserted into pET-21b(+) vector, cloned in E. coli TOP10, and finally expressed in E. coli BL21(DE3). Results: The expressed recombinant protein was entirely found in insoluble form (inclusion bodies). The insoluble protein was denatured in 6M guanidine-HCl and refolded by descending denaturant concentration dialysis. Moreover, the recombinant protein was purified by Ni–NTA column chromatography. The changing temperature had no effects on solubilizing protein and the maximum expression was achieved at 0.5 mM concentration of isopropyl-D-thiogalactopyranoside (IPTG) induction. Conclusion: Bacterial expression system is a timesaving tool and refolding by descending denaturant concentration dialysis is a rapid and reliable method.
Authors and Affiliations
Adel Najafi, Mohsen Tafaghodi, Mojtaba Sankian, Yousef Amini, Kiarash Ghazvini
Keywords
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