Scorpion-Toxin Mimics of CD4 in Complex with Human Immunodeficiency Virus gp120: Crystal Structures, Molecular Mimicry, and Neutralization Breadth

Vaccine Research Center, National Institutes of Health, Bethesda, Maryland 20892, USA.
Structure (Impact Factor: 5.62). 06/2005; 13(5):755-68. DOI: 10.1016/j.str.2005.03.006
Source: PubMed


The binding surface on CD4 for the HIV-1 gp120 envelope glycoprotein has been transplanted previously onto a scorpion-toxin scaffold. Here, we use X-ray crystallography to characterize atomic-level details of gp120 with this transplant, CD4M33. Despite known envelope flexibility, the conformation of gp120 induced by CD4M33 was so similar to that induced by CD4 that localized measures were required to distinguish ligand-induced differences from lattice variation. To investigate relationships between structure, function, and mimicry, an F23 analog of CD4M33 was devised. Structural and thermodynamic analyses showed F23 to be a better molecular mimic of CD4 than CD4M33. F23 also showed increased neutralization breadth, against diverse isolates of HIV-1, HIV-2, and SIVcpz. Our results lend insight into the stability of the CD4 bound conformation of gp120, define measures that quantify molecular mimicry as a function of evolutionary distance, and suggest how such evaluations might be useful in developing mimetic antagonists with increased neutralization breadth.

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Available from: Richard Wyatt, Oct 03, 2015
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    • "It was found that only three regions (31–35; 40–48 and 58–64 residues) are the most important for gp120 binding [11] [12]. The 27-amino acid-length F23 peptide is an efficient mimic of the CD4 domain to study the CD4–gp120 binding interaction [8]. The purpose of our research was application of the F23 peptide for detection of gp120 protein in solution. "
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    ABSTRACT: To explore a low-cost novel probe for HIV detection, we designed and prepared a 50-amino acid-length short fusion peptide (FP-50) via Escherichia coli in vivo expression. It was employed as a novel probe to detect HIV-1 gp120 protein. The detectable level of gp120 protein using the FP-50 peptide was approximately 20-200 times lower than previously published methods that used a pair of monoclonal antibodies. Thus, this short peptide is a very promising component for detection of gp120 protein during early stages of HIV infection.
    Biochemical and Biophysical Research Communications 12/2013; 443(1). DOI:10.1016/j.bbrc.2013.11.095 · 2.30 Impact Factor
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    • "Since diverse relationships exist among epitope specificity, neutralization and effector function [47], generating broader ‘protective’ antibody responses to the CD4BS, V1V2, C1 and C5, and CD4i epitopes will contribute to improving efficacy of vaccine(s) against HIV. In addition, as opposed to the restrictive function of CD4i-directed antibodies suggested earlier [48], several recent studies have emphasized the role of antibodies targeting the CD4i epitope [41], [49] and demonstrated the diversity in specificity and functionality of these antibodies [50]. "
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    ABSTRACT: Entry of HIV-1 into target cells requires binding of the viral envelope glycoprotein (Env) to cellular receptors and subsequent conformational changes that culminates in fusion of viral and target cell membranes. Recent structural information has revealed that these conformational transitions are regulated by three conserved but potentially flexible layers stacked between the receptor-binding domain (gp120) and the fusion arm (gp41) of Env. We hypothesized that artificial insertion of a covalent bond will 'snap' Env into a conformation that is less mobile and stably expose conserved sites. Therefore, we analyzed the interface between these gp120 layers (layers 1, 2 and 3) and identified residues that may form disulfide bonds when substituted with cysteines. We subsequently probed the structures of the resultant mutant gp120 proteins by assaying their binding to a variety of ligands using Surface Plasmon Resonance (SPR) assay. We found that a single disulfide bond strategically inserted between the highly conserved layers 1 and 2 (C65-C115) is able to 'lock' gp120 in a CD4 receptor bound conformation (in the absence of CD4), as indicated by the lower dissociation constant (Kd) for the CD4-induced (CD4i) epitope binding 17b antibody. When disulfide-stabilized monomeric (gp120) and trimeric (gp140) Envs were used to immunize rabbits, they were found to elicit a higher proportion of antibodies directed against both CD4i and CD4 binding site epitopes than the wild-type proteins. These results demonstrate that structure-guided stabilization of inter-layer interactions within HIV-1 Env can be used to expose conserved epitopes and potentially overcome the sequence diversity of these molecules.
    PLoS ONE 10/2013; 8(10):e76139. DOI:10.1371/journal.pone.0076139 · 3.23 Impact Factor
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    • "Cc-CD4M33F23 was generated and compared to Cc-CD4 using clade B and C pseudoviruses. CD4M33F23 is a CD4 mimetic that has been shown to be more effective than CD4 and to interact with gp120 and inhibit infection of a wide range of HIV-1 isolates [31]. Co-incubating Cc-CD4 with HIV-1 pseudovirus representing clade C resulted in a significant inhibition of infection of TZM-bl cells (Fig. 3). "
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    ABSTRACT: The HIV/AIDS pandemic remains an enormous global health concern. Despite effective prevention options, 2.6 million new infections occur annually, with women in developing countries accounting for more than half of these infections. New prevention strategies that can be used by women are urgently needed. Topical microbicides specific for HIV-1 represent a promising prevention strategy. Conceptually, using harmless bacteria to display peptides or proteins capable of blocking entry provides an inexpensive approach to microbicide development. To avoid the potential pitfalls of engineering commensal bacteria, our strategy is to genetically display infection inhibitors on a non-native bacterium and rely on topical application of stabilized bacteria before potential virus exposure. Due to the high density cell-surface display capabilities and the inherent low toxicity of the bacterium, the S-layer mediated protein display capabilities of the non-pathogenic bacterium Caulobacter crescentus has been exploited for this approach. We have demonstrated that C. crescentus displaying MIP1α or CD4 interfered with the virus entry pathway and provided significant protection from HIV-1 pseudovirus representing clade B in a standard single cycle infection assay. Here we have expanded our C. crescentus based microbicide approach with additional and diverse classes of natural and synthetic inhibitors of the HIV-1 entry pathway. All display constructs provided variable but significant protection from HIV-1 infection; some with protection as high as 70%. Further, we describe protection from infection with additional viral clades. These findings indicate the significant potential for engineering C. crescentus to be an effective and readily adaptable HIV-1 microbicide platform.
    PLoS ONE 06/2013; 8(6):e65965. DOI:10.1371/journal.pone.0065965 · 3.23 Impact Factor
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