AMINO-TERMINAL SUBSTITUTIONS IN THE CCR5 CORECEPTOR IMPAIR GP120 BINDING AND HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 ENTRY

The CC-chemokine receptor CCR5 is required for the efficient fusion of macrophage (M)-tropic human immunodeficiency virus type 1 (HIV-1) str ains with the plasma membrane of CD4(+) cells and interacts directly w ith the viral surface glycoprotein gp120. Although receptor chimera st udies have provided useful information, the domains of CCR5 that funct ion for HIV-1 entry, including the site of gp120 interaction, have not been unambiguously identified, Here, we use site-directed, alanine-sc anning mutagenesis of CCR5 to show that substitutions of the negativel y 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 a nd JR-FL M-tropic strains and the DH123 dual-tropic strain. These muta tions also impair Env-mediated membrane fusion and the gp120-CCR5 inte raction. Of these three residues, only D11 is necessary for CC-chemoki ne-mediated inhibition of HIV-1 entry, which is, however, also depende nt on other extracellular CCR5 residues. Thus, the gp120 and CC-chemok ine binding sites on CCR5 are only partially overlapping, and the form er site requires negatively charged residues in the amino-terminal CCR 5 domain.