HIV-1 gp41 Envelope Residues 650–685 Exposed on Native Virus Act as a Lectin to Bind Epithelial Cell Galactosyl Ceramide

Abstract

The initial step in the interaction between human immunodeficiency virus (HIV-1) and epithelial cells is the binding of HIV-1 envelope glycoproteins to the epithelial cell galactosyl ceramide (GalCer). Here we show that HIV-1 envelope gp41 residues 650 -685 bind GalCer in a galactose-specific manner. The gp41 residues that display this lectin activity are highly conserved among HIV-1 isolates and constitute three regions: residues 650 -661, which encompass a charged helix; residues 662-667, referred to as the conserved epitope ELDKWA, the epitope recognized by antibodies that neutralize HIV-1 entry in epithelial and CD4؉ -mononucleated cells; and residues 668 -685, a hydrophobic Trp-rich sequence that stabilizes the structure of the galactose binding site. Similar to other galactose-specific lectins, the gp41 lectin site is active only as an oligomer. Finally the orientation of the galactose toward the gp41 lectin site appears to be controlled by the lipid microenvironment of the epithelial membrane. From the experimental data we construct a theoretical model of the interaction between gp41 and GalCer based on thermodynamic considerations. This model integrates the dynamics and the spatial organization of the viral envelope glycoproteins, GalCer organized in raft microdomains in the apical region of the epithelial cell membrane and the interfacial water. Characterization of the minimal sequence and structure of gp41 in direct interaction with GalCer may help unravel the still unknown immunogenic determinant able to elicit antibodies against ELDKWA and target of one of the rare neutralizing antibodies against gp41.

A mucosally targeted subunit vaccine candidate eliciting HIV-1 transcytosis-blocking Abs

“…More recently, it was shown that the region surrounding this epitope (corresponding to the P1 peptide, Fig. 1) mediates viral interaction with the epithelial cell receptor of HIV-1, GalCer (7,10,11). This interaction may be important for the nonfusion processes that enable HIV-1 to cross simple epithelia and can be blocked by the 2F5 IgG, ELDKWA-specific IgA from infected or highly exposed persistently seronegative individuals (20,21).…”

“…Because, the context in which this linear epitope is presented affects its processing by antigen-presenting cells (2), we focused on the minimal region of gp41 that allows GalCer binding (i.e., P1) and is assumed to preserve important features of the native structure (10). Preventing this specific and crucial lectin-like interaction is necessary, and seemingly sufficient, to block HIV-1 transcytosis across the epithelial barrier and infection of CD4 ϩ cells.…”

“…Transcytosis was expressed as percentage of control, i.e., transcytosis observed in absence of Ig fraction. For analysis of the P1 reversion of the neutralizing activity, samples were incubated overnight at 4°C with 125 M P1 [critical concentration for dimerization (10)], extensively dialyzed to remove unbound P1 and assayed as described above. IgA and IgG were depleted from fecal samples by using biotinylated-goat anti-mouse IgA and IgG, respectively, followed by binding to 25 l of streptavidin-agarose beads (Neutravidin, Pierce).…”

“…[12][13][14]. In addition to direct targeting of HIV-1 by neutralizing Abs (15), HIV-1 transcytosis can be blocked by disruption of rafts and by Abs against the ELDKWA epitope (7,8,10,11).…”

“…Recently, the GalCer-binding domain of gp41 was mapped to a highly conserved, partially ␣-helical region of gp41 (''P1'' peptide, residues 649-684, Fig. 1), extending between the gp120 cap and the transmembrane domain (9)(10)(11). This domain spans conserved epitopes including the ELDKWA (''Katinger'') sequence, the target of the broadly neutralizing human monoclonal IgG, 2F5 ( Fig.…”