title: Peptide mimotopes selected with HIV-1-blocking monoclonal antibodies against CCR5 represent motifs specific for HIV-1 entry.
abstract: CCR5 is a chemokine receptor that mediates entry of human immunodeficiency virus-1 (HIV-1). Two monoclonal antibodies (mAbs) that block HIV-1 entry, 3A9 and 5C7, were used to select peptide mimotopes of sequences on CCR5 from phage displayed peptide libraries. The selected mimotofpes comprised motifs at the N-terminus and on the first and third extracellular loops (ECL1 and ECL3) of CCR5. Amino acids in these motifs were exchanged for alanines by site-directed mutagenesis (sdm) in the cDNA for human CCR5. Ensuing effects on antibody binding to CCR5, cellular entry of HIV-1 and chemokine-induced signalling were analysed by transfection of mutant cDNAs into HEK293.CD4 cells. For both mAbs, fluorescence-activated cell sorting analysis was used to define overlapping conformational epitopes on CCR5 at the N-terminus, on ECL1 and ECL3. Mutation of the N-terminal motif 10YD11 prevented HIV-1 entry into transfected cells as judged by single round infection assays with R5 and R5X4 HIV-1 isolates, as did mutation of the motif 96FG97 in ECL1, whereas mutation of the motif 274RLD276 in ECL3 had only a minor effect. None of the motifs in CCR5 relevant to HIV-1 entry disrupted chemokine-induced signalling. Thus, peptide mimotopes of conformational contact sites of CCR5 with the paratope of mAbs 3A9 and 5C7 represent sites on CCR5 that are essential for HIV-1 entry. Structural knowledge of these mimotopes could help elucidate the nature of the interaction between CCR5 and HIV-1, and thus the derivation of specific inhibitors of entry of HIV-1 into susceptible cells without interference with chemokine signalling.
authors.affiliation: Georg-Speyer-Haus, Institute for Biomedical Research, Frankfurt am Main, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Königs
authors.forename: Anette
authors.initials: A
authors.lastname: Pustowka
authors.forename: James
authors.initials: J
authors.lastname: Irving
authors.forename: Christoph
authors.initials: C
authors.lastname: Kessel
authors.forename: Katharina
authors.initials: K
authors.lastname: Klich
authors.forename: Valérie
authors.initials: V
authors.lastname: Wegner
authors.forename: Merrill J
authors.initials: MJ
authors.lastname: Rowley
authors.forename: Ian R
authors.initials: IR
authors.lastname: Mackay
authors.forename: Wolfhart
authors.initials: W
authors.lastname: Kreuz
authors.forename: Christian
authors.initials: C
authors.lastname: Griesinger
authors.forename: Ursula
authors.initials: U
authors.lastname: Dietrich
mesh_terms: D020816:Amino Acid Motifs / Q000235:genetics / Q000276:immunology; D019138:Antibodies, Blocking / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001666:Binding Sites, Antibody / Q000276:immunology; D002478:Cells, Cultured; D018604:Epitope Mapping; D005434:Flow Cytometry; D015497:HIV-1 / Q000276:immunology* / Q000378:metabolism / Q000502:physiology; D006801:Humans; D008954:Models, Biological; D018716:Molecular Mimicry / Q000502:physiology*; D009154:Mutation; D010446:Peptide Fragments / Q000276:immunology / Q000302:isolation & purification*; D019713:Receptors, CCR5 / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D053586:Virus Internalization
chemical_list: D019138:Antibodies, Blocking; D000911:Antibodies, Monoclonal; D010446:Peptide Fragments; D019713:Receptors, CCR5