The Broadly Neutralizing Anti-Human Immunodeficiency Virus Type 1 4E10 Monoclonal Antibody Is Better Adapted to Membrane-Bound Epitope Recognition and Blocking than 2F5

ABSTRACT The broadly neutralizing 2F5 and 4E10 monoclonal antibodies (MAbs) recognize epitopes within the membrane-proximal external region (MPER) that connects the human immunodeficiency virus type 1 (HIV-1) envelope gp41 ectodomain with the transmembrane anchor. By adopting different conformations that stably insert into the virion external membrane interface, such as helical structures, a conserved aromatic-rich sequence within the MPER is thought to participate in HIV-1-cell fusion. Recent experimental evidence suggests that the neutralizing activity of 2F5 and 4E10 might correlate with the MAbs' capacity to recognize epitopes inserted into the viral membrane, thereby impairing MPER fusogenic activity. To gain new insights into the molecular mechanism underlying viral neutralization by these antibodies, we have compared the capacities of 2F5 and 4E10 to block the membrane-disorganizing activity of MPER peptides inserted into the surface bilayer of solution-diffusing unilamellar vesicles. Both MAbs inhibited leakage of vesicular aqueous contents (membrane permeabilization) and intervesicular lipid mixing (membrane fusion) promoted by MPER-derived peptides. Thus, our data support the idea that antibody binding to a membrane-inserted epitope may interfere with the function of the MPER during gp41-induced fusion. Antibody insertion into a cholesterol-containing, uncharged virion-like membrane is mediated by specific epitope recognition, and moreover, partitioning-coupled folding into a helix reduces the efficiency of 2F5 MAb binding to its epitope in the membrane. We conclude that the capacity to interfere with the membrane activity of conserved MPER sequences is best correlated with the broad neutralization of the 4E10 MAb.

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Specific Interactions between the Viral Coreceptor CXCR4 and the Biguanide-Based Compound NB325 Mediate Inhibition of Human Immunodeficiency Virus Type 1 Infection

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00866-08 ◽

2008 ◽

Vol 53 (2) ◽

pp. 631-638 ◽

Cited By ~ 18

Author(s):

Nina Thakkar ◽

Vanessa Pirrone ◽

Shendra Passic ◽

Wei Zhu ◽

Vladyslav Kholodovych ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

T Lymphocytes ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Epitope Recognition ◽

Flow Cytometric ◽

Immunodeficiency Virus ◽

Loop 2 ◽

Hiv 1

ABSTRACT The present studies were conducted to better define the mechanism of action of polyethylene hexamethylene biguanide (PEHMB) (designated herein as NB325), which was shown in previous studies to inhibit infection by the human immunodeficiency virus type 1 (HIV-1). Fluorescence-activated flow cytometric analyses of activated human CD4+ T lymphocytes exposed to NB325 demonstrated concentration-dependent reductions in CXCR4 epitope recognition in the absence of altered recognition of selected CD4 or CD3 epitopes. NB325 also inhibited chemotaxis of CD4+ T lymphocytes induced by the CXCR4 ligand CXCL12. However, NB325 did not cause CXCR4 internalization (unlike CXCL12) and did not interfere with CXCL12 binding. Additional flow cytometric analyses using antibodies with distinct specificities for extracellular domains of CXCR4 demonstrated that NB325 specifically interfered with antibody binding to extracellular loop 2 (ECL2). This interaction was confirmed using competitive binding analyses, in which a peptide derived from CXCR4 ECL2 competitively inhibited NB325-mediated reductions in CXCR4 epitope recognition. Collectively, these results demonstrate that the biguanide-based compound NB325 inhibits HIV-1 infection by specifically interacting with the HIV-1 coreceptor CXCR4.

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Antibody Neutralization Escape Mediated by Point Mutations in the Intracytoplasmic Tail of Human Immunodeficiency Virus Type 1 gp41

Journal of Virology ◽

10.1128/jvi.79.4.2097-2107.2005 ◽

2005 ◽

Vol 79 (4) ◽

pp. 2097-2107 ◽

Cited By ~ 46

Author(s):

Vandana Kalia ◽

Surojit Sarkar ◽

Phalguni Gupta ◽

Ronald C. Montelaro

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Neutralizing Antibodies ◽

Point Mutations ◽

Mutant Virus ◽

Gp41 Ectodomain ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT The persistence of human immunodeficiency virus type 1 (HIV-1) infection in the presence of robust host immunity has been associated in part with variation in viral envelope proteins leading to antigenic variation and escape from neutralizing antibodies. Previous studies of natural neutralization escape mutants have predominantly focused on gp120 and gp41 ectodomain sequence variations that alter antibody binding via changes in conformation or glycosylation pattern of the Env, likely due to the immune pressure exerted on the exposed ectodomain component of the glycoprotein. Here, we show for the first time a novel mechanism by which point mutations in the intracytoplasmic tail of the transmembrane component (gp41) of envelope can render the virus resistant to neutralization by monoclonal antibodies and broadly neutralizing polyclonal serum antibodies. Point mutations in a highly conserved structural motif within the intracytoplasmic tail resulted in decreased binding of neutralizing antibodies to the Env ectodomain, evidently due to allosteric changes both in the gp41 ectodomain and in gp120. While receptor binding and infectivity of the mutant virus remained unaltered, the changes in Env antigenicity were associated with an increase in neutralization resistance of the mutant virus. These studies demonstrate the structurally integrated nature of gp120 and gp41 and underscore a previously unrecognized potentially critical role for even minor sequence variation of the intracytoplasmic tail in modulating the antigenicity of the ectodomain of HIV-1 envelope glycoprotein complex.

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Analysis of Neutralization Specificities in Polyclonal Sera Derived from Human Immunodeficiency Virus Type 1-Infected Individuals

Journal of Virology ◽

10.1128/jvi.01992-08 ◽

2008 ◽

Vol 83 (2) ◽

pp. 1045-1059 ◽

Cited By ~ 201

Author(s):

Yuxing Li ◽

Krisha Svehla ◽

Mark K. Louder ◽

Diane Wycuff ◽

Sanjay Phogat ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Protein Characterization ◽

Viral Neutralization ◽

Formidable Challenge ◽

Immunodeficiency Virus ◽

Env Protein ◽

Hiv 1

ABSTRACT During human immunodeficiency virus type 1 (HIV-1) infection, patients develop various levels of neutralizing antibody (NAb) responses. In some cases, patient sera can potently neutralize diverse strains of HIV-1, but the antibody specificities that mediate this broad neutralization are not known, and their elucidation remains a formidable challenge. Due to variable and nonneutralizing determinants on the exterior envelope glycoprotein (Env), nonnative Env protein released from cells, and the glycan shielding that assembles in the context of the quaternary structure of the functional spike, HIV-1 Env elicits a myriad of binding antibodies. However, few of these antibodies can neutralize circulating viruses. We present a systematic analysis of the NAb specificities of a panel of HIV-1-positive sera, using methodologies that identify both conformational and continuous neutralization determinants on the HIV-1 Env protein. Characterization of sera included selective adsorption with native gp120 and specific point mutant variants, chimeric virus analysis, and peptide inhibition of viral neutralization. The gp120 protein was the major neutralizing determinant for most sera, although not all neutralization activity against all viruses could be identified. In some broadly neutralizing sera, the gp120-directed neutralization mapped to the CD4 binding region of gp120. In addition, we found evidence that regions of the gp120 coreceptor binding site may also be a target of neutralizing activity. Sera displaying limited neutralization breadth were mapped to the immunogenic V3 region of gp120. In a subset of sera, we also identified NAbs directed against the conserved, membrane-proximal external region of gp41. These data allow a more detailed understanding of the humoral responses to the HIV-1 Env protein and provide insights regarding the most relevant targets for HIV-1 vaccine design.

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