Analysis of the human immunodeficiency virus type 1 gp41 membrane proximal external region arrayed on hepatitis B surface antigen particles

Structural Virology Section, Vaccine Research Center, NIAID, NIH, Bethesda, MD 20892-3005, USA.
Virology (Impact Factor: 3.32). 04/2008; 373(1):72-84. DOI: 10.1016/j.virol.2007.11.005
Source: PubMed


Vaccine immunogens derived from the envelope glycoproteins of the human immunodeficiency virus type 1 (HIV-1) that elicit broad neutralizing antibodies remain an elusive goal. The highly conserved 30 amino-acid membrane proximal external region (MPER) of HIV gp41 contains the hydrophobic epitopes for two rare HIV-1 broad cross-reactive neutralizing antibodies, 2F5 and 4E10. Both these antibodies possess relatively hydrophobic HCDR3 loops and demonstrate enhanced binding to their epitopes in the context of the native gp160 precursor envelope glycoprotein by the intimate juxtaposition of a lipid membrane. The hepatitis B surface antigen (HBsAg) S1 protein forms nanoparticles that can be utilized both as an immunogenic array of the MPER and to provide the lipid environment needed for enhanced 2F5 and 4E10 binding. We show that recombinant HBsAg particles with MPER (HBsAg-MPER) appended at the C-terminus of the S1 protein are recognized by 2F5 and 4E10 with high affinity compared to positioning the MPER at the N-terminus or the extracellular loop (ECL) of S1. Addition of C-terminal hydrophobic residues derived from the HIV-1 Env transmembrane region further enhances recognition of the MPER by both 2F5 and 4E10. Delipidation of the HBsAg-MPER particles decreases 2F5 and 4E10 binding and subsequent reconstitution with synthetic lipids restores optimal binding. Inoculation of the particles into small animals raised cross-reactive antibodies that recognize both the MPER and HIV-1 gp160 envelope glycoproteins expressed on the cell surface; however, no neutralizing activity could be detected. Prime:Boost immunization of the HBsAg-MPER particles in sequence with HIV envelope glycoprotein proteoliposomes (Env-PLs) did not raise neutralizing antibodies that could be mapped to the MPER region. However, the Env-PLs did raise anti-Env antibodies that had the ability to neutralize selected HIV-1 isolates. The first generation HBsAg-MPER particles represent a unique means to present HIV-1 envelope glycoprotein neutralizing determinants to the immune system.

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    • "It is also possible that the presence of complete C-terminal MPER region in gp140L exerted immune suppression due to mimicry of this region to the self-protein cardiolipin [47]. It was shown earlier that the MPER region is weakly immunogenic when it was presented to the immune system on particulate Hepatititis B surface antigen particles [48]. However, recently analysis of human monoclonal antibody 10E8, isolated from an HIV-1 infected individual with high neutralization titers, demonstrated that the breadth of neutralizing antibody response of this antibody is mediated by recognition of MPER [49]. "
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    ABSTRACT: The development of a vaccine against human immunodeficiency virus-1 (HIV-1) capable of inducing broad humoral and cellular responses at both the systemic and mucosal levels will be critical for combating the global AIDS epidemic. We previously demonstrated the ability of Newcastle disease virus (NDV) as a vaccine vector to express oligomeric Env protein gp160 and induce potent humoral and mucosal immune responses. In the present study, we used NDV vaccine strain LaSota as a vector to compare the biochemical and immunogenic properties of vector-expressed gp160, gp120, and two versions of gp140 (a derivative of gp160 made by deleting the transmembrane and cytoplasmic domains), namely: gp140L, which contained the complete membrane-proximal external region (MPER), and gp140S, which lacks the distal half of MPER. We show that, similar to gp160, NDV-expressed gp140S and gp120, but not gp140L, formed higher-order oligomers that retained recognition by conformationally sensitive monoclonal antibodies. Immunization of guinea pigs by the intranasal route with rLaSota/gp140S resulted in significantly greater systemic and mucosal antibody responses compared to the other recombinants. Immunization with rLaSota/140S, rLaSota/140L rLaSota/120 resulted in mixed Th1/Th2 immune responses as compared to Th1-biased immune responses induced by rLaSota/160. Importantly, rLaSota/gp140S induced neutralizing antibody responses to homologous HIV-1 strain BaL.26 and laboratory adapted HIV-1 strain MN.3 that were stronger than those elicited by the other NDV recombinants. Additionally, rLaSota/gp140S induced greater CD4+ and CD8+ T-cell responses in mice. These studies illustrate that rLaSota/gp140S is a promising vaccine candidate to elicit potent mucosal, humoral and cellular immune responses to the HIV-1 Env protein.
    PLoS ONE 10/2013; 8(10):e78521. DOI:10.1371/journal.pone.0078521 · 3.23 Impact Factor
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    • "Unfortunately, many attempts at eliciting broadly neutralizing antibodies by targeting neutralizing epitopes of MPER failed [22] [23] [24] [25] [26] [27]. Two recent studies reported that presenting the ELDKWA epitope by human rhinovirus [28] or a whole MPER domain with mutations (T569A and I675V) by NCM antigen [29] successfully induced anti-HIV-1 cross-clade neutralizing antibodies. "
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    ABSTRACT: Two conserved epitopes, located in the membrane-proximal external region (MPER) of the human immunodeficiency virus type 1 (HIV-1) gp41, are recognized by two HIV-1 broadly neutralizing antibodies 2F5 and 4E10, and are promising targets for vaccine design in efforts to elicit anti-HIV-1 broadly neutralizing antibodies. Since most HIV-1 infections initiate at mucosal surfaces, induction of mucosal neutralizing antibodies is necessary and of utmost importance to counteract HIV-1 infection. Here, we utilized a mucosal vaccine vector, bovine papillomavirus (BPV) virus-like particles (VLPs), as a platform to present HIV-1 neutralizing epitopes by inserting the extended 2F5 or 4E10 epitope or the MPER domain into D-E loop of BPV L1 respectively. The chimeric VLPs presenting MPER domain resembled the HIV-1 natural epitopes better than the chimeric VLPs presenting single epitopes. Oral immunization of mice with the chimeric VLPs displaying the 2F5 epitope or MPER domain elicited epitope-specific serum IgGs and mucosal secretory IgAs. The induced antibodies specifically recognized the native conformation of MPER in the context of HIV-1 envelope protein. The antibodies induced by chimeric VLPs presenting MPER domain are able to partially neutralize HIV-1 viruses from clade B and clade C.
    Vaccine 09/2013; 31(46). DOI:10.1016/j.vaccine.2013.09.003 · 3.62 Impact Factor
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    • "Thus the structure, function and immunogenicity of the MPER have been a focus of intense study for many years, however the design of vaccines that elicit 2F5- and 4E10-like antibodies has proven difficult (e.g. [33,34]). "
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    ABSTRACT: Background The disulfide-bonded region (DSR) of HIV-1 gp41 mediates association with gp120 and plays a role in transmission of receptor-induced conformational changes in gp120 to gp41 that activate membrane fusion function. In this study, forced viral evolution of a DSR mutant that sheds gp120 was employed to identify domains within gp120-gp41 that are functionally linked to the glycoprotein association site. Results The HIV-1AD8 mutant, W596L/K601D, was serially passaged in U87.CD4.CCR5 cells until replication was restored. Whereas the W596L mutation persisted throughout the cultures, a D601H pseudoreversion in the DSR partially restored cell-free virus infectivity and virion gp120-gp41 association, with further improvements to cell-free virus infectivity following a 2nd-site D674E mutation in the membrane-proximal external region (MPER) of gp41. In an independent culture, D601H appeared with a deletion in V4 (Thr-394-Trp-395) and a D674N substitution in the MPER, however this MPER mutation was inhibitory to W596L/K601H cell-free virus infectivity. While cell-free virus infectivity was not fully restored for the revertant genotypes, their cell-to-cell transmission approached the levels observed for WT. Interestingly, the functional boost associated with the addition of D674E to W596L/K601H was not observed for cell-cell fusion where the cell-surface expressed glycoproteins function independently of virion assembly. The W596L/K601H and W596L/K601H/D674E viruses exhibited greater sensitivity to neutralization by the broadly reactive MPER directed monoclonal antibodies, 2F5 and 4E10, indicating that the reverting mutations increase the availability of conserved neutralization epitopes in the MPER. Conclusions The data indicate for the first time that functional crosstalk between the DSR and MPER operates in the context of assembled virions, with the Leu-596-His-601-Glu-674 combination optimizing viral spread via the cell-to-cell route. Our data also indicate that changes in the gp120-gp41 association site may increase the exposure of conserved MPER neutralization epitopes in virus.
    Retrovirology 04/2013; 10(1):44. DOI:10.1186/1742-4690-10-44 · 4.19 Impact Factor
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