title: Presentation of native epitopes in the V1/V2 and V3 regions of human immunodeficiency virus type 1 gp120 by fusion glycoproteins containing isolated gp120 domains.
abstract: The immune response to viral glycoproteins is often directed against conformation- and/or glycosylation-dependent structures; synthetic peptides and bacterially expressed proteins are inadequate probes for the mapping of such epitopes. This report describes a retroviral vector system that presents such native epitopes on chimeric glycoproteins in which protein fragments of interest are fused to the C terminus of the N-terminal domain of the murine leukemia virus surface protein, gp70. The system was used to express two disulfide-bonded domains from gp120, the surface protein of human immunodeficiency virus type 1 (HIV-1), that include potent neutralization epitopes. The resulting fusion glycoproteins were synthesized at high levels and were efficiently transported and secreted. A fusion protein containing the HXB2 V1/V2 domain was recognized by an HIVIIIB-infected patient serum as well as by 17 of 36 HIV-1 seropositive hemophiliac, homosexual male and intravenous drug user patient sera. Many of these HIV+ human sera reacted with V1/V2 domains from several HIV-1 clones expressed in fusion glycoproteins, indicating the presence of cross-reactive antibodies against epitopes in the V1/V2 domain. Recognition of gp(1-263):V1/V2HXB2 by the HIVIIIB-infected human patient serum was largely blocked by synthetic peptides matching V1 but not V2 sequences, while recognition of this construct by a broadly cross-reactive hemophiliac patient serum was not blocked by individual V1 or V2 peptides or by mixtures of these peptides. A construct containing the V3 domain of the IIIB strain of HIV-1, gp(1-263):V3HXB2, was recognized by sera from a human and a chimpanzee that had been infected by HIVIIIB but not by sera from hemophiliac patients who had been infected with HIV-1 of MN-like V3 serotype. The reactive sera had significantly higher titers when assayed against gp(1-263):V3HXB2 than when assayed against matching V3 peptides. Immunoprecipitation of this fusion glycoprotein by the human serum was only partially blocked by V3 peptide, indicating that this infected individual produced antibodies against epitopes in V3 that were expressed on the fusion glycoprotein but not by synthetic peptides. These data demonstrated that the chimeric glycoproteins described here effectively present native epitopes present in the V1/V2 and V3 domains of gp120 and provide efficient methods for detection of antibodies directed against native epitopes in these regions and for characterization of such epitopes.
authors.affiliation: Laboratory of Retroviral Biology, Public Health Research Institute, New York, New York 10016.
authors.forename: S C
authors.initials: SC
authors.lastname: Kayman
authors.forename: Z
authors.initials: Z
authors.lastname: Wu
authors.forename: K
authors.initials: K
authors.lastname: Revesz
authors.forename: H
authors.initials: H
authors.lastname: Chen
authors.forename: R
authors.initials: R
authors.lastname: Kopelman
authors.forename: A
authors.initials: A
authors.lastname: Pinter
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D017951:Antigen Presentation; D002460:Cell Line; D003429:Cross Reactions; D000939:Epitopes / Q000276:immunology*; D005822:Genetic Vectors; D015483:HIV Antibodies / Q000097:blood; D015699:HIV Envelope Protein gp120 / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D015658:HIV Infections / Q000276:immunology; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D009052:Leukemia Virus, Murine / Q000276:immunology; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D002679:Pan troglodytes; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000276:immunology
chemical_list: D000939:Epitopes; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins