title: Design and expression of fusion protein consists of HBsAg and Polyepitope of HCV as an HCV potential vaccine.
abstract: BACKGROUND
  Hepatitis C virus (HCV) infection is a serious public health threat worldwide. Cellular immune responses, especially cytotoxic T-lymphocytes (CTLs), play a critical role in immune response toward the HCV clearance. Since polytope vaccines have the ability to stimulate the cellular immunity, a recombinant fusion protein was developed in this study.
  
  
  METHODS
  The designed fusion protein is composed of hepatitis B surface antigen (HBsAg), as an immunocarrier, fused to an HCV polytope sequence. The polytope containing five immunogenic epitopes of HCV was designed to induce specific CTL responses. The construct was cloned into the pET-28a, and its expression was investigated in BL21 (DE3), BL21 pLysS, BL21 pLysE, and BL21 AI Escherichia coli strains using 12% gel sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Finally, the identity of expressed fusion protein was confirmed by Western blotting using anti-His monoclonal antibody and affinity chromatography was applied to purify the expressed protein.
  
  
  RESULTS
  The accuracy of the construct was confirmed by restriction map analysis and sequencing. The transformation of the construct into the BL21 (DE3), pLysS, and pLysE E. coli strains did not lead to any expression. The fusion protein was found to be toxic for E. coli DE3. By applying two steps inhibition, the fusion protein was successfully expressed in BL21 (AI) E. coli strain.
  
  
  CONCLUSIONS
  The HBsAg-polytope fusion protein expressed in this study can be further evaluated for its immunogenicity in animal models.
authors.affiliation: Department of Biology, Islamic Azad University, Tonekabon Branch, Tonekabon, Iran.
authors.forename: Monireh
authors.initials: M
authors.lastname: Gholizadeh
authors.affiliation: Department of Genetics, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran.
authors.forename: Hossein
authors.initials: H
authors.lastname: Khanahmad
authors.affiliation: Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Arash
authors.initials: A
authors.lastname: Memarnejadian
authors.affiliation: Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Aghasadeghi
authors.affiliation: Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Farzin
authors.initials: F
authors.lastname: Roohvand
authors.affiliation: Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Seyed Mehdi
authors.initials: SM
authors.lastname: Sadat
authors.affiliation: Department Of Rabies, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Reza Ahangari
authors.initials: RA
authors.lastname: Cohan
authors.affiliation: Department of Biology, Islamic Azad University, Tonekabon Branch, Tonekabon, Iran.
authors.forename: Ali
authors.initials: A
authors.lastname: Nazemi
authors.affiliation: Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Motevalli
authors.affiliation: Department Of Rabies, Pasteur Institute of Iran, Tehran, Iran ; Department of Immunology, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.
authors.forename: Vahid
authors.initials: V
authors.lastname: Asgary
authors.affiliation: Department of Molecular Medicine, Pasteur Institute of Iran, Tehran, Iran.
authors.forename: Roghaye
authors.initials: R
authors.lastname: Arezumand
keywords: Expression; hepatitis C virus; polytope vaccine