Identification of a human epitope in hepatitis C virus (HCV) core protein using a molecularly cloned antibody repertoire from a non-symptomatic, anti-HCV-positive patient

J Gen Virol. 2000 Feb;81(Pt 2):461-9. doi: 10.1099/0022-1317-81-2-461.

Abstract

Healthy carriers of hepatitis C virus (HCV) infection exhibit a specific antibody response against all HCV antigens, which could play a role in disease control. Generation of panels of human antibodies may permit a thorough characterization of this response and further identify particular antibodies with potential clinical value. To this effect, we have established a human phage-display antibody library from a patient exhibiting a high antibody response against HCV antigens and no clinical symptoms of disease. This library was screened against a recombinant core antigen [amino acids (aa) 1-119] produced in E. coli. Two recombinant Fab-carrying phages (rFabCs) were isolated and characterized. Both rFabC3 and rFabC14 recognize aa 1-48 on core antigen, but rFabC14 is competed out by a synthetic peptide, C(2-20) (aa 1-20), at much lower concentrations than rFabC3. In order to identify more precisely the recognition sites of these antibodies, we produced soluble forms of the rFabs (sFabs), and used them to pan a random phage-display peptide library. A single peptide sequence, QLITKPL, was identified with sFabC3, while two equally represented sequences, HAFPHLH and SAPSSKN, were isolated using sFabC14. The QLITKPL sequence was partially localized between aa 8 and 14 of core protein, but no clear homology was found for the two sFabC14 peptides. However, we confirmed the specificity of these peptides by competition experiments with sFabC14.

MeSH terms

  • Adult
  • Amino Acid Sequence
  • Antigens, Viral* / genetics
  • Base Sequence
  • Binding, Competitive
  • Carrier State / immunology
  • Cloning, Molecular
  • DNA Primers / genetics
  • Epitope Mapping
  • Epitopes / genetics
  • Escherichia coli / genetics
  • Female
  • Hepacivirus / genetics
  • Hepacivirus / immunology*
  • Hepatitis C / genetics
  • Hepatitis C / immunology
  • Hepatitis C Antibodies / blood
  • Hepatitis C Antibodies / genetics*
  • Humans
  • Immunoglobulin Fab Fragments / genetics
  • Immunoglobulin Fab Fragments / metabolism
  • In Vitro Techniques
  • Molecular Sequence Data
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Viral Core Proteins / genetics
  • Viral Core Proteins / immunology

Substances

  • Antigens, Viral
  • DNA Primers
  • Epitopes
  • Hepatitis C Antibodies
  • Immunoglobulin Fab Fragments
  • Recombinant Proteins
  • Viral Core Proteins

Associated data

  • GENBANK/AJ251251
  • GENBANK/AJ251252
  • GENBANK/AJ251253
  • GENBANK/AJ251291