Expression of full length of HCV-E2 gene in yeast Pichia pastoris
Subject Areas : VirologyRaheleh Solat 1 , Pooneh Rahimi 2 , Gholam Reza Bakhshi Khaniki 3 , Mohammad Reza Aghasadeghi 4 , Rouhallah Vahabpour 5 , Mehdi Shokri 6
1 - M.Sc., Department of Biology, Payameh Nour University, Tehran, Iran.
2 - Assistant Professor, Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
3 - Professor, Department of Biology, Payameh Nour University, Tehran, Iran
4 - Associate Professor, Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
5 - Ph.D. student, Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
6 - Ph.D. student, Department of Immunology, Pasteur Institute of Iran,Tehran, Iran.
Keywords: Glycoprotein E2, Hepatitis C virus JFH1 strain, pPICZAa vector, Pichia Pastoris,
Abstract :
Background & Objectives: Hepatitis C virus infection initiates through binding of E2 glycoprotein to its specific human liver cell receptor. Therefore, this glycoprotein is considered as one of the important targets in drug and vaccine researches against this infection. This project was accomplished for the first time to transfer the full length of E2 gene by using recombinant pPICZAa-E2 (HCV-2a, JFH1) vector into Pichia pastoris yeast KM71H strain, and to evaluate the possibility of its expression in this expression system. Materials & Methods: The E2 gene was amplified by using primers containing the EcoRI and XbaI restriction sites and was inserted into the (T-PTG 19) and (pPICZAa) vectors to be transferred into the E. coli. The recombinant pPICZAa-E2 vector was transferred into the yeast through electroporation, and it was evaluated by digestion and sequencing. The yeast was grown in YNB medium contained methanol for five days. Gene expression was studied by western blot. Results: Construction of a recombinant vector pPICZAa-(JFH1)E2, transforming the yeast and its expression in KM71H strain were successfully done. Conclusion: In this study, the whole length of E2 Ag of HCV JFH1, as the protype strain of this virus, was successfully expressed. The result of this study can be used for further analysis of the structure and function of this protein.