Expression of HCV genotype-4 core antigen in prokaryotic E. coli system for diagnosis of HCV infection in Egypt

Theodor Bilharz Research Institute

Bibliographic Information

Authors: Saleh E.M.; Gouda A.E.; Medhat A.M.; Ahmed H.O.; Shemis M.A.

Journal: Protein Expression and Purification

Publisher: Academic Press Inc.

Publication Date: December 2021

Volume / Issue: Volume 188

Article No.: 105965

ISSN: 10465928

DOI: 10.1016/j.pep.2021.105965

Scopus: View on Scopus

PubMed: 34461217

Document Type: Article

Authors and Affiliations

Saleh E.M., Biochemistry Department, Faculty of Science, Ain Shams University, Cairo, Egypt; Gouda A.E., Biochemistry and Molecular Biology Department, Theodor Bilharz Research Institute, Giza, Egypt; Medhat A.M., Biochemistry Department, Faculty of Science, Ain Shams University, Cairo, Egypt; Ahmed H.O., Biochemistry and Molecular Biology Department, Theodor Bilharz Research Institute, Giza, Egypt; Shemis M.A., Biochemistry and Molecular Biology Department, Theodor Bilharz Research Institute, Giza, Egypt

Abstract

Background: Egypt has a high prevalence of hepatitis C virus (HCV) infection with 92.5% of genotype-4. Aim: This study aimed to clone and express the core gene of HCV genotype-4 for using it to develop a highly sensitive, specific, and cost-effective diagnostic assay for detecting HCV infection. Methods: Using synthetic HCV genotype-4 core gene, pET15b as E. coli expression vector, and 1 mM lactose as inducer, the HCV core protein (MW 17 kDa) was expressed in the form of inclusion bodies (IBs) that was purified and solubilized using 8 M guanidinium HCl. The recombinant core protein was in vitro refolded by a rapid dilution method for further purification using weak cation exchange liquid chromatography. The immunogenicity of the purified protein was tested by ELISA using 129 serum samples. Results: The recombinant core protein was successfully expressed and purified. The results also showed that the in-house anti-HCV core assay is accurate, specific (~96.6%), and highly sensitive (~100%) in accordance with the commercial ELISA kit. Conclusion: The sensitivity, specificity, and reproducibility of the developed assay were high and promising to be used as a screening assay for detecting HCV infection. © 2021 Elsevier Inc.

Keywords

Core protein; ELISA; HCV; Inclusion bodies; Refolding; Antibodies, Viral; Antigens, Viral; Chromatography, Ion Exchange; Cloning, Molecular; Egypt; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Gene Expression; Genetic Vectors; Genotype; Guanidine; Hepacivirus; Hepatitis C; Humans; Immune Sera; Prevalence; Protein Refolding; Recombinant Proteins; Viral Core Proteins; antiserum; nucleocapsid protein, Hepatitis C virus; recombinant protein; virus antibody; virus antigen; biosynthesis; blood; cell inclusion; chemistry; classification; enzyme linked immunosorbent assay; gene vector; genetics; human; immunology; ion exchange chromatography; isolation and purification; metabolism; molecular cloning; procedures; virology

Citation Information

Scopus Citations: 6

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