The C-terminal tail of the gp41 transmembrane envelope glycoprotein of HIV-1 clades A, B, C, and D may exist in two conformations: an analysis of sequence, structure, and function

Abstract

In addition to the major ectodomain, the gp41 transmembrane glycoprotein of HIV-1 is now known to have a minor ectodomain that is part of the long C-terminal tail. Both ectodomains are highly antigenic, carry neutralizing and non-neutralizing epitopes, and are involved in virus-mediated fusion activity. However, data have so far been biologically based, and derived solely from T cell line-adapted (TCLA), B clade viruses. Here we have carried out sequence and theoretically based structural analyses of 357 gp41 C-terminal sequences of mainly primary isolates of HIV-1 clades A, B, C, and D. Data show that all these viruses have the potential to form a tail loop structure (the minor ectodomain) supported by three, beta-sheet, membrane-spanning domains (MSDs). This means that the first (N-terminal) tyrosine-based sorting signal of the gp41 tail is situated outside the cell membrane and is non-functional, and that gp41 that reaches the cell surface may be recycled back into the cytoplasm through the activity of the second tyrosine-sorting signal. However, we suggest that only a minority of cell-associated gp41 molecules - those destined for incorporation into virions - has 3 MSDs and the minor ectodomain. Most intracellular gp41 has the conventional single MSD, no minor ectodomain, a functional first tyrosine-based sorting signal, and in line with current thinking is degraded intracellularly. The gp41 structural diversity suggested here can be viewed as an evolutionary strategy to minimize HIV-1 envelope glycoprotein expression on the cell surface, and hence possible cytotoxicity and immune attack on the infected cell.

Fig. 1

Summary of the conservation of gp41 C-terminal amino acid residues 690–793 of HIV-1 clades A to D, using a 7-residue moving window.

Fig. 2

Consensus sequences for gp41 C-terminal amino acid residues 690–793 of HIV-1 clades A to D combined (top line), and for clades A to D individually. The potential tyrosine-dependent sorting signals are in red. A potential di-leucine signal is pink. Residues common to the majority of sequences are in black, while residues that vary are in green. Where two residues are equally represented between the consensus sequences of the four clades, the overall consensus sequence is based on the larger number of sequences analyzed. The proposed first, second, and third MSDs are overlined. Underlined is the antigenically active Kennedy sequence.

Fig. 3

Analysis of the structure of the gp41 C-terminal residues 690–793 of HIV-1 clade A using various algorithms: (a) hydropathy prediction, where high values represent the more hydrophobic regions; (b) α-helix prediction, where high values indicate a greater probability of α-helical structure; (c) β-sheet prediction, where high values indicate a greater probability of β-sheet structure; (d) β-turn prediction, where high values indicate a greater probability of a β-turn; (e) polarity prediction, where high values suggest high polarity; and (f) buried residue prediction, where high values suggest buried residues.

Fig. 4

Schematic showing the suggested membrane insertion of a monomer of (a) the 1-MSD and (b) the 3-MSD forms of gp41 of HIV-1 clades A to D. The positions of the two potential tyrosine-sorting signals and the minor ectodomain containing the Kennedy sequence are shown. The major ectodomain at the N-terminus of gp41 and gp120 have been omitted. The distance shown in (b) between MSD 2 and 3 (reading from left to right) is conjectural.

Fig. 5

Representation of some possible interactions of the minor ectodomain (residues 713–753) of the 3-MSD form of gp41 with other elements of the Env trimer. Shown are residues 645–863, comprising part of the main ectodomain, the MSDs, the minor ectodomain, and the entire C-terminal tail of the gp41 of HIV-1 NL-4.3 (clade B). Only a monomer is represented. There may be interactions between MSD 1, 2, and 3, between the minor ectodomain and the gp41 major ectodomain, between the minor ectodomain and elements of gp120, or with the other gp41 monomers that form the trimer (arrows). The nine MSDs of the trimer could also interact with each other. The antigenically active Kennedy sequence (731–752) containing neutralizing and non-neutralizing epitopes is shown as the outer face of the minor ectodomain.