Induced Fit in HIV-Neutralizing Antibody Complexes: Evidence for Alternative Conformations of the gp120 V3 Loop and the Molecular Basis for Broad Neutralization † , ‡

Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100, Israel.
Biochemistry (Impact Factor: 3.02). 06/2005; 44(19):7250-8. DOI: 10.1021/bi047387t
Source: PubMed


Human monoclonal antibody (mAb) 447-52D neutralizes a broad spectrum of HIV-1 isolates, whereas murine mAb 0.5beta, raised against gp120 of the X4 isolate HIV-1(IIIB), neutralizes this strain specifically. Two distinct gp120 V3 peptides, V3(MN) and V3(IIIB), adopt alternative beta-hairpin conformations when bound to 447-52D and 0.5beta, respectively, suggesting that the alternative conformations of this loop play a key role in determining the coreceptor specificity of HIV-1. To test this hypothesis and to better understand the molecular basis underlying an antibody's breadth of neutralization, the solution structure of the V3(IIIB) peptide bound to 447-52D was determined by NMR. V3(IIIB) and V3(MN) peptides bound to 447-52D exhibited the same N-terminal strand conformation, while the V3(IIIB) peptide revealed alternative N-terminal conformations when bound to 447-52D and 0.5beta. Comparison of the three known V3 structures leads to a model in which a 180 degrees change in the orientation of the side chains and the resulting one-residue shift in hydrogen bonding patterns in the N-terminal strand of the beta-hairpins markedly alter the topology of the surface that interacts with antibodies and that can potentially interact with the HIV-1 coreceptors. Predominant interactions of 447-52D with three conserved residues of the N-terminal side of the V3 loop, K312, I314, and I316, can account for its broad cross reactivity, whereas the predominant interactions of 0.5beta with variable residues underlie its strain specificity.

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    • "n insight into the conformation of the V3 region presented to the immune system by HIV - 1 , the structures of V3 peptides bound to HIV - 1 neutralizing antibodies were determined by X - ray crystallography ( Bell et al . , 2008 ; Dhillon et al . , 2008 ; Stanfield et al . , 2003 ; Stanfield et al . , 2004 ; Stanfield et al . , 2006 ) and by NMR ( Rosen et al . , 2005 ; Sharon et al . , 2003 ; Tugarinov et al . , 1999 ; Tugarinov et al . , 2000 ) . All bound V3 structures revealed a β - hairpin conformation with an RMSD between the hairpin regions of any two V3 structures in the different complexes ranging between 1 . 2 and 2 . 5 Å ."
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    ABSTRACT: Synthetic peptides offer an attractive option for development of a V3-directed vaccine. However, immunization with flexible linear peptides may result in an immune response to multiple conformations, many of which differ from the native conformation of the corresponding region in the protein. Here we show that optimization of the location of a disulfide bond in peptides constrained to mimic the beta-hairpin conformation of the V3, yields an immunogen that elicits a 30-fold stronger HIV-1 neutralizing response in rabbits compared with the homologous linear V3 peptide. The HIV-1 neutralizing response elicited by the optimally constrained peptide is also significantly stronger than that elicited by a gp120 construct in which the V3 is exposed. Neutralization of an HIV-1 strain that shares only 72% identity with the immunizing peptide was demonstrated. The most effective immunogen was also able to neutralize primary isolates that are more resistant to neutralization such as SS1196 and 6535.
    Full-text · Article · Mar 2010 · Virology
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    • "effects of the amino acid substitutions , replacing Arg 315 with either Ala or Gln reduced antibody binding affinity substantially ( Table 4 ) . This observation is in agreement with previous mapping studies ( Gorny et al . , 1992 ; Keller et al . , 1993 ) as well as NMR and crystallographic analyses of this antibody in complex with a V3 peptide ( Rosen et al . , 2005 ; Stanfield et al . , 2004 ) . Alanine substitution of residues at positions 304 - 309 , 313 , and 319 - 320 moderately diminished the binding affinity of 447 - 52D . Pro 313 in the V3 sequence is important for proper 447 - 52D binding interactions ( Stanfield et al . , 2004 ) and , thus , a reduction in binding affinity upon mutation to"
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    ABSTRACT: The identification of cross-neutralizing antibodies to HIV-1 is important for designing antigens aimed at eliciting similar antibodies upon immunization. The monoclonal antibody (mAb) F425-B4e8 had been suggested previously to bind an epitope at the base of V3 and shown to neutralize two primary HIV isolates. Here, we have assessed the neutralization breadth of mAb F425-B4e8 using a 40-member panel of primary HIV-1 and determined the epitope specificity of the mAb. The antibody was able to neutralize 8 clade B viruses (n=16), 1 clade C virus (n=11), and 2 clade D viruses (n=6), thus placing it among the more broadly neutralizing anti-V3 antibodies described so far. Contrary to an initial report, results from our scanning mutagenesis of the V3 region suggest that mAb F425-B4e8 interacts primarily with the crown/tip of V3, notably Ile(309), Arg(315), and Phe(317). Despite the somewhat limited neutralization breadth of mAb F425-B4e8, the results presented here, along with analyses from other cross-neutralizing anti-V3 mAbs, may facilitate the template-based design of antigens that target V3 and permit neutralization of HIV-1 strains in which the V3 region is accessible to antibodies.
    Full-text · Article · Sep 2007 · Virology
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    • "How then, might V3 influence which coreceptor is used? While our findings do not exclude a role for conformational changes, available data concerning (1) overall similarity of structures adopted by V3 peptides in complex with neutralizing antibodies (Rosen et al., 2005; Stanfield et al., 2003 "
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    ABSTRACT: The V3 region of the HIV-1 envelope complex is known to play a pivotal role in directing the choice between the chemokine receptors CCR5 and CXCR4 for use as coreceptors, but the underlying structural mechanism is as yet unknown (reviewed in Hartley et al. [2005]). Sharon et al. have proposed a model in which V3 drives coreceptor choice by adopting structures that mimic β-hairpin loops (40's loops) from chemokines selective for either CCR5 or CXCR4 (Sharon et al., 2003). The model is centered on evidence that structures adopted by V3-derived peptides in complex with V3-directed neutralizing antibodies show significant structural homology with 40's loops from (1) CCR5 binding chemokines in the case of the proposed "CCR5-tropic conformation" (1NIZ [Sharon et al., 2003]), and (2) the CXCR4 binding chemokine SDF-1 (CXCL12) in the case of the proposed "CXCR4-tropic conformation" (1QNZ [Tugarinov et al., 2000]). While the structural analogy model has attracted interest because of its relevance to the development of vaccine strategies based on HIV-1 V3 (Rosen et al., 2005; Zolla-Pazner, 2004), a number of theoretical objections have been raised (Hartley et al., 2005; Lusso, 2003). Here we present new data concerning the extent of structural similarity between antibody-complexed V3 structures and 40's loop regions from all of the chemokines for which structural data is available. Our results challenge the Sharon et al. (2003) model by showing that the degree of similarity between the structures of V3 and those of β-hairpins from HIV-1 coreceptor-selective chemokines is too low to be of biological significance.
    Full-text · Article · May 2006 · Structure
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