From in-silico immunogenicity verification to in vitro expression of recombinant Core-NS3 fusion protein of HCV

• Published in: Bratislavské lékarské listy Vol. 118; no. 4; pp. 189 - 195
• Main Authors: Hekmat, S, Siadat, S D, Aghasadeghi, M R, Sadat, S M, Bahramali, G, Aslani, M M, Mahdavi, M, Shahbazi, S
• Format: Journal Article
• Language: English
• Published: Slovakia 2017
• Subjects: Hepacivirus - immunology; Hepatitis C - immunology; Hepatitis C - prevention & control; Hepatitis C - therapy; Humans; Immunotherapy - methods; Viral Hepatitis Vaccines - immunology; Viral Nonstructural Proteins - immunology; Core NS3; HCV vaccine; fusion protein
• ISSN: 0006-9248
• DOI: 10.4149/BLL_2017_038

Hepatitis C virus (HCV) is a serious global health burden. There is no effective vaccine against HCV and new direct acting antivirals (DAAs) are so expensive and virtually unavailable to the public. Therefore, seeking for therapeutic or prophylactic vaccines is exigent and reliever. The secondary and tertiary structures of the recombinant Core-NS3 (rC-N) fusion protein of HCV and its B and T-cells epitopes were evaluated with bioinformatics software. Cloning and in vitro expression of rC-N were performed by pET24a(+) and E.coli BL21-DE3 expression host, respectively. The recombinant protein purification was done by affinity chromatography method and then identified by Western blotting using anti-His monoclonal antibody. The sequences of rC-N protein consist of 1-118 amino acid parts of Core and 1095-1384 amino acids of NS3 were connected by a flexible linker (AAY) with proteasome cleavable site. The expressed and purified 46.7292 kDa rC-N protein had antigenic value up to threshold and conservancy found in this chimeric protein. Ramchandran Plot analysis represented that most residues were fallen in favourable regions. It also interacted with both type I and II major histocompatibility complex (MHC I, II) molecules. The rC-N had antigenic behaviour to create T cell responses. The results indicated that conserved rC-N protein had the ability to induce T-cell-mediated immune responses and it could be utilized as a therapeutic vaccine candidate against HCV (Tab. 3, Fig. 4, Ref. 40).