Abstract
Although human immunodeficiency type 1 (HIV-1) infection induces strong antibody responses to the viral envelope glycoprotein (Env) only a few of these antibodies possess the capacity to neutralize a broad range of strains. The induction of such antibodies represents an important goal in the development of a preventive vaccine against the infection. Among the broadly neutralizing monoclonal antibodies discovered so far, three (2F5, Z13 and 4E10) target the short and hidden membrane proximal external region (MPER) of the gp41 transmembrane protein. Antibody responses to MPER are rarely observed in HIV-infected individuals or after immunization with Env immunogens. To initiate antibody responses to MPER in its membrane-embedded native conformation, we generated expression plasmids encoding the membrane-anchored ectodomain of gp41 with N-terminal deletions of various sizes. Following transfection of these plasmids, the MPER domains are displayed on the cell surface and incorporated into HIV virus like particles (VLP). Transfected cells displaying MPER mutants bound as efficiently to both 2F5 and 4E10 as cells transfected with a plasmid encoding full-length Env. Mice immunized with VLPs containing the MPER mutants produced MPER-specific antibodies, the levels of which could be increased by the trimerization of the displayed proteins as well as by a DNA prime-VLP boost immunization strategy. Although 2F5 competed for binding to MPER with antibodies in sera of some of the immunized mice, neutralizing activity could not be detected. Whether this is due to inefficient binding of the induced antibodies to MPER in the context of wild type Env or whether the overall MPER-specific antibody response induced by the MPER display mutants is too low to reveal neutralizing activity, remains to be determined.
Highlights
The HIV envelope glycoprotein complex mediates virus entry into the cell [1] and represents the only exposed viral protein on the surface of the virion
Display of N-terminal deletion mutants of gp41 on the surface of cells increases 2F5 and 4E10 binding compared to gp41 (Fig. 2). This does not seem to be due to higher expression levels of membrane proximal external region (MPER) display mutants, since the virus like particles (VLP) contain at least as much of gp41 as of the MPER display mutants (Fig. 3)
To allow direct comparison of the properties of gp41 with those of the MPER display mutants, the gp41 was expressed with an N-terminal leader sequence, the trimerization domain, the ollas-tag, and the flexible linker region
Summary
The HIV envelope glycoprotein complex mediates virus entry into the cell [1] and represents the only exposed viral protein on the surface of the virion. In the six helix bundle situation and in the presence of the fusion-peptide proximal region, Buzon and coworkers described a helical rod-like structure on the N-terminal side of MPER with a ,90u turn of the MPER chain at Asn 687 (numbering according to Gen bank entry AF128126.3). Liu and co-workers described a labile ahelical trimeric structure of the MPER spanning residues 672–693 [6]. These observations suggest that the MPER within the functional envelope spike adopts a complex structure including trimerized and non-trimerized parts as suggested by Zhu and coworkers [7,8], that may influence the epitope conformation recognized by MPER-targeted neutralizing antibodies
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