Structure and function of the HIV envelope glycoprotein as entry mediator, vaccine immunogen, and target for inhibitors

Abstract

This chapter discusses the advances of the envelope glycoprotein (Env) structure as related to the interactions of conserved Env structures with receptor molecules and antibodies with implications for the design of vaccine immunogens and inhibitors. The human immunodeficiency virus (HIV) Env binds to cell surface–associated receptor (CD4) and coreceptor (CCR5 or CXCR4) by one of its two non-covalently associated subunits, gp120. The induced conformational changes activate the other subunit (gp41), which causes the fusion of the viral with the plasma cell membranes resulting in the delivery of the viral genome into the cell and the initiation of the infection cycle. As the only HIV protein exposed to the environment, the Env is also a major immunogen to which neutralizing antibodies are directed and a target that is relatively easy to access by inhibitors. A fundamental problem in the development of effective vaccines and inhibitors against HIV is the rapid generation of alterations at high levels of expression during long chronic infection and the resulting significant heterogeneity of the Env. The preservation of the Env function as an entry mediator and limitations on size and expression impose restrictions on its variability and lead to the existence of conserved structures.

Figure 1

Primary structure of HIV‐1 Env glycoprotein and sequence variations in different regions of the Env lead to several HIV‐1 subtypes. (A) A schematic diagram representing different regions of HIV‐1 Env glycoprotein. Approximate locations of the cleavage sites (arrowheads), glycosylation sites (branched symbols), constant (C1–C5) and variable (V1–V5) regions, fusion domain (FD), heptad repeats (HR1 and HR2), and transmembrane domain (TM) are shown along with the numbering scheme of amino acids. The cross‐linking disulfide bonds connecting various segments are indicated as brackets. (B) The phylogenetic tree constructed by using consensus sequences of HIV‐1 M group subtypes A1, A2, B, C, D, F1, F2, G, and H is shown along with evolutionary distances with the maximum value of 0.1.

Figure 2

Sequence variability at each amino acid position of the Env of prominent HIV‐1 subtypes B and C. The x‐axes indicate the positions of amino acids as well as allowed gaps from multiple sequence alignments while the y‐axes denote the value of sequence variation at each position. The variable loops apparently have larger sequence variations comparing to other portions of the Env (see the text).

Figure 3

Crystal structures of gp120 core in the unliganded and liganded states. (A) Ribbon diagram of the unliganded SIV gp120 core is shown as in the same orientation of the liganded HIV gp120 structure. The color codes are in rainbow representation from colors blue to red for the N‐ to C‐terminus. The positions of variable loops and bridging sheets are labeled. (B) Ribbon diagram depicting the 3D‐structure of HIV gp120 core complexed with the first two domains (D1, D2) of CD4 receptor and the Fab fragment of human monoclonal neutralizing antibody 17b (CD4 and 17b are not shown here). The outer domains (in green and yellow) of liganded and unliganded gp120 are relatively conserved while a dramatic change in the inner domain (blue and cyan) occurs. The bridging sheet that connects inner and outer domains is not formed in the unliganded gp120.

Figure 4

Molecular surface diagrams of unliganded (A) and liganded (B) gp120 cores are rendered as viewed from the perspective of CD4 receptor binding. The residues in direct contact with CD4 are in blue; residues contacting the CD4i antibodies, namely, 17b and X5 are in red. The contact residues were selected by limiting interatomic distance of 3.8 Å between gp120 core to the CD4 and CD4i antibodies.

Figure 5

Structures of HIV‐1 gp120 complexes with CD4 receptor and CD4i antibodies, 17b and X5. (A) HIV‐1 gp120 core (green) is bound to the CD4 (orange) and Fab 17b antibody (magenta for heavy and pink for light chains). (B) CDR H3 conformations of antibodies in the free and bound forms are given in stereoviews as crystal structures of 17b and X5 antibodies were available in isolation (PDB codes: 1RZ8 and 1RHH, respectively). (C) HIV‐1 gp120 core with an intact V3 (green) is bound to the CD4 (orange) and Fab X5 antibody (blue for heavy and cyan for light chains). CDR H3 loops are labeled and indicated by arrows. The CDR H3 conformations of 17b antibody (C) are similar in free and bound forms. Notably, the H3 of X5 (D) undergoes a large conformational change with the maximum displacement up to 17 Å (blue in bound form and light blue in free form).

Figure 6

Crystal structure trimeric gp41 fragment. (A) A schematic view of gp41 Env showing the locations of functional regions corresponding to the N36 and C34 peptide fragments. (B) The peptides N36–C34 complex forms a stable α‐helical domain of six‐helix bundle structure. The N36 (green) and C34 (red) helices point to each other in the opposite directions; N36 forms the inner core of the trimeric structure while C34 warps the core. (C) The bottom view of the trimer clearly depicts the arrangement of N36–C34 complex.

Figure 7

Diagrams illustrate 3D structures of Env spikes as revealed from cryoelectron microscopy. (A) The model obtained at ∼3.2‐nm resolution by Zhu et al. has a head structure comprising trimeric gp120 in three lobes, which is supported by three separate legs in a tripodlike arrangement. The model fitting based on the available gp120 crystal structures suggests carbohydrates on the top; CD4 on the periphery appears closer to the variable loops which may shield the conserved regions of gp120 and gp41. (B) The Env spike model at 2.8‐nm resolution as presented by Zanetti et al. is similar in having a three‐lobed head supported by stalk as seen by Zhu et al. but with a subtly different compact stalk with no obvious separation as three legs at the gp41 stem. Model fitting using the gp120 core structures indicates the exposed receptor binding sites, which are protected by the sugars and variable loops. The bridging sheet is either hidden at the trimer‐g41 interface or protected by the V3 loop.

Figure 8

Conformations of CDR H3s from b12, m18, m14, and F105 antibodies. Residues Arg94 and Trp103 from the framework regions play critical role in maintaining the H3 conformations by involving specific salt bridges at the bases. The differences in H3s are markedly noticed along the torso and tip regions.

Figure 9

Two different antigen‐binding sites and binding modes CDRs. (A) In gp120–Fab X5 antibody interaction, the long CDR H3 protrudes into the CD4i binding site. (B) Conversely, in the SARS Env–Fab m396 antibody interaction, the antibody CDRs form like a canyon around the protruding binding site.

Figure 10

Steric restriction of access to CD4i epitopes on CD4 binding. (A) The sketch with molecules shown describes the attachment of HIV‐1 from viral membrane to the cell surface CD4 receptor. The binding of CD4 induces conformational changes resulting into the exposure of coreceptor binding site, which is sterically restricted for the CD4i antibodies. Taken into considerations of the dimensions derived from structures of gp120, CD4, and possible flexibility of CD4 molecule, a total distance of about 85 Å between the gp120 and target cell membrane is measured. (B) Dimensions of antibodies in different formats, Fv, Fab, and IgG molecules, are also shown. This clearly shows that CD4i antibodies of scFvs and Fabs have better access to the restricted binding site for competing with the coreceptor than IgGs have.

Figure 11

Antibody interactions at the membrane‐proximal region of gp41. (A) Schematic diagram of gp41 shows the different important regions, FD, fusion domain, HR1, HR2‐heptad repeats, and TM, transmembrane domain. The location of membrane‐proximal region containing the core 2F5 and 4E10 epitopes on the Trp‐rich region of gp41 is indicated along with amino acids sequence. Sequence numbering corresponds to HXB2 scheme. Crystal structures of Fab 2F5 (B) and 4E10 (C) in complex with peptides from the MPER. The H3s of the antibodies are shown in green.

Figure 12

Mimicry of receptor CD4 by miniprotein CD4M33. The binding of gp120 (green) to the CD4 (first domain, D1 is only shown) on left and the miniprotein CD4M33 on right are depicted in ribbon diagrams.

Figure 13

Comparisons of CD4 D1 domain with VH domains of CD4bs antibodies b12 and m18. (A) D1 domain CD4 (green) with Phe43 in sticks. (B) VH domain of b12 antibody (cyan) with Tyr53 at the CDR H2 in sticks. (C) VH domain of m18 antibody (blue) with Phe99 at the CDR H2 in sticks. (D) Backbone skeletal views of the CDR2‐like region of CD4 and the H3 of m18 indicate a common β‐hairpin structure with a phenylalanine residue at the tip.