title: A hidden region in the third variable domain of HIV-1 IIIB gp120 identified by a monoclonal antibody.
abstract: The third variable domain (V3 domain) of HIV-1 gp120 is involved in virus neutralization by antibody, in determination of cell tropism, and in syncytium-inducing/non-syncytium-inducing capacity. Antibodies are highly specific tools to delineate the role of different V3 amino acid sequences in these processes, and to dissect events occurring during synthesis of gp120/160, gp120-CD4 interaction, cellular infection, and syncytium formation. We describe here an IgG1 murine monoclonal antibody (MAb), coded IIIB-V3-01, that was raised with a synthetic peptide (FVTIGKIGNMRQAHC) derived from the carboxy-terminal flank of the HIV-1 IIIB V3 domain. The binding site of this antibody was mapped to the sequence IGKIGNMRQ, using Pepscan analysis. In ELISA, this antibody binds to E. coli-derived gp120 from HIV-1 IIIB, which is denatured and not glycosylated. The antibody showed no neutralizing activity against HIV-1 IIIB, MN, SF2, or RF in a virus neutralization assay and in a syncytium formation inhibition assay. In addition, this antibody did not react with gp120 expressed on the surface of IIIB-infected MOLT-3 cells in FACS analysis. To assess whether the epitope defined by MAb IIIB-V3-01 is hidden on native gp120, reactivity of the antibody with SDS-DTT-denatured or DTT-denatured glycosylated gp120 (CHO cell produced) was tested. Both these treatments exposed the epitope for binding. From these data we conclude that the epitope defined by MAB IIIB-V3-01 is hidden on glycosylated recombinant gp120, and is not accessible on gp120 expressed on the membrane of HIV-1, IIIB-infected cells.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Immunology and Medical Microbiology, Medical Biological Laboratory TNO, Rijswijk, The Netherlands.
authors.forename: J D
authors.initials: JD
authors.lastname: Laman
authors.forename: M M
authors.initials: MM
authors.lastname: Schellekens
authors.forename: G K
authors.initials: GK
authors.lastname: Lewis
authors.forename: J P
authors.initials: JP
authors.lastname: Moore
authors.forename: T J
authors.initials: TJ
authors.lastname: Matthews
authors.forename: J P
authors.initials: JP
authors.lastname: Langedijk
authors.forename: R H
authors.initials: RH
authors.lastname: Meloen
authors.forename: W J
authors.initials: WJ
authors.lastname: Boersma
authors.forename: E
authors.initials: E
authors.lastname: Claassen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001666:Binding Sites, Antibody; D005260:Female; D015726:Giant Cells; D015483:HIV Antibodies / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000737:chemistry / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology* / Q000502:physiology; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments