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. 2014 Jan;88(2):1249-58.
doi: 10.1128/JVI.02664-13. Epub 2013 Nov 13.

Mechanism of HIV-1 neutralization by antibodies targeting a membrane-proximal region of gp41

Jia Chen  1 , Gary Frey, Hanqin Peng, Sophia Rits-Volloch, Jetta Garrity, Michael S Seaman, Bing Chen
Affiliations

Mechanism of HIV-1 neutralization by antibodies targeting a membrane-proximal region of gp41

Jia Chen et al. J Virol. 2014 Jan.

Abstract

Induction of broadly neutralizing antibodies (bNAbs) is an important goal for HIV-1 vaccine development. Two autoreactive bNAbs, 2F5 and 4E10, recognize a conserved region on the HIV-1 envelope glycoprotein gp41 adjacent to the viral membrane known as the membrane-proximal external region (MPER). They block viral infection by targeting a fusion-intermediate conformation of gp41, assisted by an additional interaction with the viral membrane. Another MPER-specific antibody, 10E8, has recently been reported to neutralize HIV-1 with potency and breadth much greater than those of 2F5 or 4E10, but it appeared not to bind phospholipids and might target the untriggered envelope spikes, raising the hope that the MPER could be harnessed for vaccine design without major immunological concerns. Here, we show by three independent approaches that 10E8 indeed binds lipid bilayers through two hydrophobic residues in its CDR H3 (third heavy-chain complementarity-determining region). Its weak affinity for membranes in general and preference for cholesterol-rich membranes may account for its great neutralization potency, as it is less likely than other MPER-specific antibodies to bind cellular membranes nonspecifically. 10E8 binds with high affinity to a construct mimicking the fusion intermediate of gp41 but fails to recognize the envelope trimers representing the untriggered conformation. Moreover, we can improve the potency of 4E10 without affecting its binding to gp41 by a modification of its lipid-interacting CDR H3. These results reveal a general mechanism of HIV-1 neutralization by MPER-specific antibodies that involves interactions with viral lipids.

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Figures

FIG 1
FIG 1
Interactions of 10E8 with lipid bilayers. A liposome flotation assay was used to detect binding to HIV-1-like membranes at 37°C by various antibodies, i.e., 17b, 4E10, 10E8, 10E8-F100aA, and 10E8-W100bA. Seven fractions throughout the sucrose gradient (from top [t] to bottom) after centrifugation were collected and analyzed by Western blotting. The heavy and light chains of antibodies are indicated by H and L, respectively.
FIG 2
FIG 2
Interactions of 10E8 with lipid bilayers. Dynamic light scattering analysis was used to monitor cross-linking of liposomes with an HIV-1-like lipid composition at 37°C by various antibodies, i.e., 17b, 4E10, 10E8, 10E8-F100aA, and 10E8-W100bA. PBS was used as a negative control. Regularization histograms plotting percentages of scattering intensity (%intensity) against the hydrodynamic radii of particles (Rh) in nanometers are shown.
FIG 3
FIG 3
Interactions of 10E8 with lipid bilayers. Interactions of lipid bilayers with an HIV-1-like lipid composition (A) and with a PC/CL ratio of 3:1 (B) with various antibodies were measured by SPR at room temperature. Liposomes incorporated with 0.5% biotin-labeled PE were immobilized on the surface of a sensor chip coated with streptavidin, and various antibodies, i.e., 17b, VRC01, 4E10, 10E8, 10E8-F100aA, 10E8-W100bA, and 10E8-FW-AA (10E8-F100aA-W100bA), at 1 μM were passed over the lipid bilayer surfaces. The recorded sensograms are shown.
FIG 4
FIG 4
10E8 targets the pre-hairpin intermediate conformation of HIV-1 gp41. Antibody binding to the HIV-1 92UG037.8 envelope trimer was expressed on 293T cell surfaces by FACS analysis. 293T cells were transfected with either 92UG037.8 gp160 or no DNA as a negative control. Binding of antibodies VRC01, 4E10, 10E8, 4E10 plus 2D CD4, 10E8 plus 2D CD4, 10E8-F100aA, 10E8-W100bA, and 10E8-F100aA-W100bA to the cell surfaces was detected with a phycoerythrin-conjugated goat anti-human secondary antibody. Histograms plotting cell counts against fluorescence intensity are shown. The vertical dashed red lines indicate the peak centers of the untreated 293T cells as a negative control. The experiments were repeated three times with similar results.
FIG 5
FIG 5
10E8 targets the pre-hairpin intermediate conformation of HIV-1 gp41. (A) 10E8 was analyzed by SPR assay for binding to HIV-1 gp140, gp41-inter, and gp41-post in complex with an antibody, 1281, in the presence of 0.1% dodecyl maltoside. 10E8 IgG was captured on the surface of a sensor chip coated with protein A to avoid potential artifacts introduced by protein immobilization. gp140, gp41-inter, and gp41-post–1281 at various concentrations were passed over the antibody surface individually without regeneration for single-cycle kinetic analysis. The recorded sensograms are shown. The molecular masses of the three envelope constructs are also indicated. Since the SPR response is proportional to the molecular mass of the binding analyte, the differences between gp41-inter and gp140 are much greater than those shown. (B) Similar to panel A, 4E10 was analyzed for binding to HIV-1 gp140, gp41-inter, and gp41-post in complex with anti-gp41 antibody 1281. (C) Binding of 10E8 and its variants 10E8-F100aA, 10E8-W100bA, and 10E8-F100aA-W100bA to gp41-inter was compared by single-cycle kinetic analysis in the standard HBS buffer. Binding of 10E8 to gp140 was also performed to evaluate the interaction without dodecyl maltoside. (D) Control experiments by single-cycle kinetic analysis to demonstrate binding of gp140 used in panels A, B, and C to the gp120-specific antibodies 2G12 and VRC01. All of the binding constructs are summarized in Table S2 in the supplemental material.
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References

    1. Harrison SC. 2008. Viral membrane fusion. Nat. Struct. Mol. Biol. 15:690–698. 10.1038/nsmb.1456 - DOI - PMC - PubMed
    1. Chan DC, Kim PS. 1998. HIV entry and its inhibition. Cell 93:681–684. 10.1016/S0092-8674(00)81430-0 - DOI - PubMed
    1. Kilby JM, Eron JJ. 2003. Novel therapies based on mechanisms of HIV-1 cell entry. N. Engl. J. Med. 348:2228–2238. 10.1056/NEJMra022812 - DOI - PubMed
    1. Cohen MS, Shaw GM, McMichael AJ, Haynes BF. 2011. Acute HIV-1 infection. N. Engl. J. Med. 364:1943–1954. 10.1056/NEJMra1011874 - DOI - PMC - PubMed
    1. Richman DD, Wrin T, Little SJ, Petropoulos CJ. 2003. Rapid evolution of the neutralizing antibody response to HIV type 1 infection. Proc. Natl. Acad. Sci. U. S. A. 100:4144–4149. 10.1073/pnas.0630530100 - DOI - PMC - PubMed

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