title: Amino-terminal substitutions in the CCR5 coreceptor impair gp120 binding and human immunodeficiency virus type 1 entry.
abstract: The CC-chemokine receptor CCR5 is required for the efficient fusion of macrophage (M)-tropic human immunodeficiency virus type 1 (HIV-1) strains with the plasma membrane of CD4+ cells and interacts directly with the viral surface glycoprotein gp120. Although receptor chimera studies have provided useful information, the domains of CCR5 that function for HIV-1 entry, including the site of gp120 interaction, have not been unambiguously identified. Here, we use site-directed, alanine-scanning mutagenesis of CCR5 to show that substitutions of the negatively charged aspartic acid residues at positions 2 and 11 (D2A and D11A) and a glutamic acid residue at position 18 (E18A), individually or in combination, impair or abolish CCR5-mediated HIV-1 entry for the ADA and JR-FL M-tropic strains and the DH123 dual-tropic strain. These mutations also impair Env-mediated membrane fusion and the gp120-CCR5 interaction. Of these three residues, only D11 is necessary for CC-chemokine-mediated inhibition of HIV-1 entry, which is, however, also dependent on other extracellular CCR5 residues. Thus, the gp120 and CC-chemokine binding sites on CCR5 are only partially overlapping, and the former site requires negatively charged residues in the amino-terminal CCR5 domain.
authors.affiliation: Aaron Diamond AIDS Research Center, The Rockefeller University, New York, New York 10016, USA. tdragic@adarc.org
authors.forename: T
authors.initials: T
authors.lastname: Dragic
authors.forename: A
authors.initials: A
authors.lastname: Trkola
authors.forename: S W
authors.initials: SW
authors.lastname: Lin
authors.forename: K A
authors.initials: KA
authors.lastname: Nagashima
authors.forename: F
authors.initials: F
authors.lastname: Kajumo
authors.forename: L
authors.initials: L
authors.lastname: Zhao
authors.forename: W C
authors.initials: WC
authors.lastname: Olson
authors.forename: L
authors.initials: L
authors.lastname: Wu
authors.forename: C R
authors.initials: CR
authors.lastname: Mackay
authors.forename: G P
authors.initials: GP
authors.lastname: Allaway
authors.forename: T P
authors.initials: TP
authors.lastname: Sakmar
authors.forename: J P
authors.initials: JP
authors.lastname: Moore
authors.forename: P J
authors.initials: PJ
authors.lastname: Maddon
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D001665:Binding Sites / Q000235:genetics; D065100:CCR5 Receptor Antagonists; D015496:CD4-Positive T-Lymphocytes / Q000502:physiology / Q000821:virology; D002459:Cell Fusion; D002460:Cell Line; D018925:Chemokines / Q000494:pharmacology; D004563:Electrochemistry; D015699:HIV Envelope Protein gp120 / Q000378:metabolism*; D015497:HIV-1 / Q000472:pathogenicity* / Q000502:physiology*; D006801:Humans; D008264:Macrophages / Q000502:physiology / Q000821:virology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D019713:Receptors, CCR5 / Q000235:genetics* / Q000502:physiology*
chemical_list: D000596:Amino Acids; D065100:CCR5 Receptor Antagonists; D018925:Chemokines; D015699:HIV Envelope Protein gp120; D019713:Receptors, CCR5