Identification of a novel epitope in the C terminus of hepatitis C virus-E2 protein that induces potent and cross-reactive neutralizing antibodies

Abstract

Hepatitis C virus (HCV) is a leading cause of chronic viral hepatitis, but an effective vaccine is still not available to prevent infection. Use of neutralizing antibodies could be a potential therapeutic option. In this study, the presence of anti-HCV antibodies in HCV-infected patients was assessed from 50 patients and the presence of neutralizing antibodies was examined using 'hepatitis C virus-like particles'. Antibodies from two samples exhibited significant inhibitory activity, suggesting that these may neutralize viral infection. Antigenic determinants generating the neutralizing antibodies from these two samples were delineated by epitope mapping using the core, E1 and E2 regions and a stretch of 45 amino acid peptide (E2C45) derived from the C-terminal region of HCV-E2 protein (aa 634-679) was designed. Results suggest that this hitherto uncharacterized region has the potential to generate neutralizing antibodies against HCV and thus be effective in preventing virus entry into liver cells. Computational analysis of the structure of the modelled peptide (E2C45) suggested high conformational entropy for this region. Furthermore, E2C45 peptide-generated antibodies could block virus entry and monoclonal antibodies generated against this peptide could also significantly reduce virus replication in a cell culture system. It is possible that the inhibition could be partly due to a conformational alteration of the CD81-binding region, preventing virus attachment to liver cells. In conclusion, this work focused on the discovery of a novel epitope at the C terminus of E2 that induces potent neutralizing antibodies in HCV-infected patients.

A plant‐derived dehydrorotenoid: a new inhibitor of hepatitis C virus entry

“…Seventy-two hours post infection, cells were fixed with paraformaldehyde, permeabilised with 0.1% Triton-X-100 and blocked with 3% BSA in PBS. Next cells were incubated with combinations of the labgenerated anti-E2 protein mAbs A8A11 and C10E8 [32], for 90 min, washed with PBS and then incubated with Alexa-488 conjugated anti-mouse-IgG antibody (Invitrogen, ThermoFisher Scientific). The nuclei were stained with DAPI (Sigma-Aldrich).…”

Characterization of linear epitope specificity of antibodies potentially contributing to spontaneous clearance of hepatitis C virus

“…Plate was washed and incubated for 1 hour with goat anti-human alkaline phosphatase conjugated IgG (Santa Cruz, sc-2454) for human serum samples and goat antimouse alkaline phosphatase conjugated IgG (Abcam, ab97020) as secondary antibodies at 1000-fold dilution. Absorbance was measured at 405 nm on Synergy HTX reader after adding p-nitrophenyl phosphate substrate [28,29]. Data was plotted using the Graphpad prism software.…”

Single Chain Fragment Variable (scFv) corresponding to a novel epitope within HCV envelope protein restricts virus entry into hepatocytes

“…The scFv can be produced in bacterial expression systems for largescale production [9][10][11]. Several scFvs have been reported to control virus infection, including scFvs against chicken infectious bursal disease virus, scFvs targeting human influenza virus H5N1, and scFvs against the phosphoprotein of Newcastle disease virus [12][13][14].…”