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:: Volume 7, Issue 1 (6-2020) ::
vacres 2020, 7(1): 44-49 Back to browse issues page
Purification of Functional HEV-ORF2 Protein from Inclusion Bodies for Vaccine and Diagnostic Applications
Fatemeh Motevalli , Mohammad Reza Amiran , Mazyar Etemadzadeh, Golnaz Bahramali , Soroush Sardari , Seyede Zahra Moravej, Pooneh Rahimi , Abolfazl Fateh , Seyed Alireza Seyed Siamdoust , Mohammad Ali Zaheri Birgani , Mojtaba Hamidi-Fard *
Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran; Viral Vaccine Research Center, Pasteur Institute of Iran, Tehran, Iran
Abstract:   (771 Views)
Introduction: Hepatitis E virus (HEV) causes emerging diseases in poor regions of the world. The ORF2 is the only protein encoded by the virus to make the viral capsid. The aggregation of proteins into inclusion bodies (IBs) while expressing ORF2 is a major challenge in bioengineering. Methods: The ORF2 conserved sequence was expressed in Escherichia coli BL21 and assessed by a modified SDS-PAGE containing a weak denaturing environment to solubilize the recombinant ORF2 protein and Western blotting. The protein of interest was evaluated by secondary structure prediction using SOPMA, homology modeling by I-TASSER and Circular Dichroism analyses. The function of the recombinant protein was investigated by an in-house ELISA using serum specimens of HEV infected patients. Results: The solubilized form of ORF2 protein was successfully expressed in E. coli BL21 and was confirmed by SDS-PAGE and Western blotting. Secondary structure prediction, homology modeling and CD analysis of the protein of interest demonstrated that the native structure of ORF2 was almost intact. The specific anti-HEV antibody was detected using this recombinant protein and an in-house ELISA test. Conclusion: We achieved new combinations of chemical agents, consisted of low concentrations of urea and detergents to overcome the aggregation of ORF2 protein in IBs inside E. coli BL21.
Keywords: Hepatitis E virus, ORF2, Inclusion bodies, Protein solubilization
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Type of Study: Original article | Subject: Synthetic peptides and novel immunogens
Received: 2020/08/31 | Accepted: 2020/06/11 | Published: 2020/09/24
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Motevalli F, Amiran M R, Etemadzadeh M, Bahramali G, Sardari S, Moravej S Z, et al . Purification of Functional HEV-ORF2 Protein from Inclusion Bodies for Vaccine and Diagnostic Applications. vacres. 2020; 7 (1) :44-49
URL: http://vacres.pasteur.ac.ir/article-1-207-en.html

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