The human immunodeficiency virus gp120 binding site on CD4: delineation by quantitative equilibrium and kinetic binding studies of mutants in conjunction with a high-resolution CD4 atomic structure.

Abstract

SummaryThe first immunoglobulin V-like domain of CD4 contains the binding site for human immunodeficiency virus gp120. Guided by the atomic structure of a two-domain CD4 fragment, we have examined gp120 interaction with informative CD4 mutants, both by equilibrium and kinetic analysis. The binding site on CD4 appears to be a surface region of about 900 A 2 on the C" edge of the domain. It contains an exposed hydrophobic residue, Phe43, on the C" strand and four positively charged residues, Lys29, Lys35, LYS46, and Arg59, on the C, C', C", and D strands, respectivdy. Replacement of Phe43 with Ala or Ile reduces affinity for gp120 by more than 500-fold; Tyr, Trp, and Leu substitutions have smaller effects. The four positively charged side chains each make significant contributions (7-50-fold). This CD4 site may dock into a conserved hydrophobic pocket bordered by several negatively charged residues in gpi20. Class II major histocompatibility complex binding includes the same region on CD4; this overlap needs to be considered in the design of inhibitors of the CD4-gp120 interaction.T he CD4 transmembrane glycoprotein participates in key steps of thymocyte differentiation (1-3) and T lymphocyte activation (4-9) by interacting with nonpolymorphic regions of class II MHC molecules (10--12). It is expressed on most thymocytes (1) and a subset of T lymphocytes, including helper T cells and class II MHC-specific cytotoxic T cells (5-7, D, 14). CD4 in humans is also the receptor for HIV (15-18). It is required for viral attachment and subsequent entry into ceils (19), and these functions account for HIV T cell tropism. The virus binds through a contact made by the gp120 component of its envelope glycoprotein (20). The binding of gp120 to CD4 has been implicated in the cellular depletion of CD4 + lymphocytes in AIDS (21-24), and circulating free gp120 is thought to interfere with normal immune function (12, 25). The central role of the CD4-gp120 interaction in the pathogenesis of AIDS justifies a detailed analysis in molecular terms, and inhibition of that interaction is also an attractive target for antiviral intervention.The extracellular part of CD4 contains four Ig-like do= mains (26). The gp120 binding site lies in the most NH2 terminal of these domains (27,28). A number of studies eraploying escape mutants (29), insertion mutagenesis (30), homology scanning mutagenesis (12,28,31), and alanine scanning mutagenesis (32) have helped to restrict the binding site to a segment surrounding the region homologous to the CDR2 of an Ig V domain.The atomic structure of a fragment containing the first two domains of CD4 has been determined by x-ray crystallography (33, 34). The first domain contains two antiparallel sheets, as predicted by its Ig V homology. The mutations that affect gp120 binding lie mostly in the antiparalld strands, denoted C' and C", that form a ridge at one edge of the domain as well as in the adjacent D strand. The C'C" loop is longer than the CDR2 loop of a typical Ig V domain, and it projects prominently ...

Structure of an HIV gp120 envelope glycoprotein in complex with the CD4 receptor and a neutralizing human antibody

“…Residues in contact are concentrated in the span from 25 to 64 of CD4, but they are distributed over six segments of gp120 (Figs 2d and 3i): one residue from the V1/V2 stem, loop ℒD, the β15-α3 excursion, the β20-β21 hairpin, strand β23 and the β24-α5 connection. These interactions are compatible with previous analyses of mutational data on both CD4 (refs 11,12,29 ) and gp120 (refs 3,13,14 ). Other groups are also involved, including some at gp120 sites that have not been tested, but residues identified as critical for binding do indeed interact with one another (Fig.…”

“…Structures of both the N-terminal two domains 8,9 and the entire extracellular portion of CD4 (ref. 10 ) have been determined, and mutagenesis indicates that the CD4 structure analogous to the second complementarity-determining region (CDR2) of immunoglobulins is critical for gp120 binding 11,12 . Conserved gp120 residues important for CD4 binding have likewise been identified by mutagenesis 3,13,14 .…”

HIV-1 Resistance to Maraviroc Conferred by a CD4 Binding Site Mutation in the Envelope Glycoprotein gp120

“…However, when K425 is modeled into the structure using the COOT modeling program, the side chain of K425 is predicted to form a hydrogen bond with the oxygen of S42 CD4 as well as generate a new cation-interaction between the aromatic ring of F43 CD4 and the Nε side chain of K425. Since F43 of CD4 is known to be a critical residue for gp120 binding (42)(43)(44), our model would suggest that enhanced interactions with CD4 may play a role in the resistance mechanism of A74.MVC.21 virus.…”

The Ixodes scapularis salivary protein, salp15, prevents the association of HIV-1 gp120 and CD4

“…The replacement of these residues with Ala decreased the affinity to gp120 by ∼500 and ∼10 fold, respectively [28]. Scyllatoxin is a scorpion toxin with structural similarities in the hairpin region with the CDR2-like loop of CD4.…”