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URL: https://pubmed.ncbi.nlm.nih.gov/19515770/

⇱ A conformational switch in human immunodeficiency virus gp41 revealed by the structures of overlapping epitopes recognized by neutralizing antibodies - PubMed

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Abstract

The membrane-proximal external region (MPER) of the human immunodeficiency virus (HIV) envelope glycoprotein (gp41) is critical for viral fusion and infectivity and is the target of three of the five known broadly neutralizing HIV type 1 (HIV-1) antibodies, 2F5, Z13, and 4E10. Here, we report the crystal structure of the Fab fragment of Z13e1, an affinity-enhanced variant of monoclonal antibody Z13, in complex with a 12-residue peptide corresponding to the core epitope (W(670)NWFDITN(677)) at 1.8-A resolution. The bound peptide adopts an S-shaped conformation composed of two tandem, perpendicular helical turns. This conformation differs strikingly from the alpha-helical structure adopted by an overlapping MPER peptide bound to 4E10. Z13e1 binds to an elbow in the MPER at the membrane interface, making relatively few interactions with conserved aromatics (Trp672 and Phe673) that are critical for 4E10 recognition. The comparison of the Z13e1 and 4E10 epitope structures reveals a conformational switch such that neutralization can occur by the recognition of the different conformations and faces of the largely amphipathic MPER. The Z13e1 structure provides significant new insights into the dynamic nature of the MPER, which likely is critical for membrane fusion, and it has significant implications for mechanisms of HIV-1 neutralization by MPER antibodies and for the design of HIV-1 immunogens.

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Figures

👁 FIG. 1.
FIG. 1.
Major features of gp41 include the fusion peptide (FP), NHR, CHR, TM, and cytoplasmic domain (CD). The MPER is located between the CHR and TM regions of gp41. The core epitopes of 2F5 (green), Z13e1 (yellow), and 4E10 (orange) are indicated. The epitope of Z13e1 is located between those of 2F5 and 4E10, but it overlaps more closely with 4E10.
👁 FIG. 2.
FIG. 2.
Crystal structure of Z13e1 with bound peptide. (A) Overview of peptide-Fab Z13e1 interactions showing the location of light and heavy chain CDR loops. Heavy chains and light chains are shown as tube representations. The locations of CDR loops L1 (brown), L2 (purple), L3 (green), H1 (marine), H2 (cyan), and H3 (blue) are shown with respect to bound peptide 178-1 (yellow). (B) Z13e1 mutations (from wild-type Z13) in L3 result in a novel L3 conformation and optimize the packing of the light chain H3 and TrpH47 against heavy chain H3 and TrpH47. Residues 95 to 97 loop out to accommodate a lowering of residues 91 to 94. LeuL94 packs against TrpH47, and a hydrogen bond from TyrH100E is made to be the backbone amide of LeuL92, further stabilizing the interface between VL and VH. LeuL93 is oriented toward L2, but the resulting hydrophobic pocket is not well packed. The light chain is shown in green, the heavy chain in whitish gray, and gp41 peptide in yellow.
👁 FIG. 3.
FIG. 3.
Combining site of Fab Z13e1. (A) Final 2Fo-Fc electron density for bound peptide 178-1 contoured at 1σ. Clear density is seen for all of the peptide epitope residues 670 to 677, whereas the C-terminal polylysine solubility tag is not ordered. (B) Interactions between the conserved aromatic residues TrpP670, TrpP672, and PheP673 and the Z13e1 heavy chain. The bound gp41 peptide 178-1 (yellow) is shown as a tube representation, with select side chain and backbone atoms as stick representations. (C) Interactions between MPER polar residues AspP674, ThrP676, and AsnP677 (hydrophilic face) and the Z13e1 heavy chain.
👁 FIG. 4.
FIG. 4.
gp41 MPER and recognition by antibodies Z13e1 and 4E10. (A) A composite model of the MPER was constructed by the fusion of the 2F5 (green), Z13e1 (yellow), and 4E10 (orange) epitope structures. The fusion of the 2F5 and Z13e1 epitopes results in an N-terminal helix in the MPER consisting of residues 666 to 672. The C terminus of the MPER is modeled using residues 678 to 683 from the 4E10 crystal structure, resulting in a C-terminal helix. The composite model for the MPER consists of two consecutive helices from residues 666 to 683 that are kinked by approximately 90° around Phe673. The apex of H3 (GlyH100A, PheH100B, LeuH100C, and AsnH100D) may adopt a conformation in the presence of the complete epitope different from that observed in complex with peptide 178-1. (B) Proposed recognition of the MPER by Z13e1 with respect to the viral membrane. The composite MPER structure is largely amphipathic; the proposed membrane-facing side of the MPER contains only hydrophobic and mainly aromatic residues, which are expected to insert into the membrane. The MPER surface facing the solvent is largely, though not completely, hydrophilic and includes residues that become deeply inserted into the Z13e1 combining site. No specific immersion depth is implied. (C) On the left is a composite model of the MPER in tube representation with the Z13e1 epitope structure shown in yellow sticks. The view is from the perspective of the antibody looking down toward the membrane surface (perpendicular to the view seen in panel B). Z13e1 binds a Trp672/Phe673-occluded conformation and contacts the hydrophilic face of the MPER and hydrophobic residues in the interior of the elbow region. The Z13e1 conformation is characterized by the capping of the C-terminal helix by Asp674. The structure in the middle shows that 4E10 binds a Trp672/Phe673-accessible conformation that is characterized by the capping of the extended C-terminal helix by Asn671. The extension of the helix then would result in the extraction of Trp672 and Phe673 out of the membrane toward the accessible face of the MPER. On the right is the superposition of the Z13e1 composite MPER model with 4E10 MPER conformation.
👁 FIG. 5.
FIG. 5.
Angle of approach of 2F5, Z13e1, and 4E10 to the MPER. (A) Orientation of the MPER bound by Z13e1 and 4E10. (Left) Z13e1 heavy chain (dark gray) and light chain (whitish gray) are shown in molecular surface representation. Bound peptide (yellow) is shown in stick representation and extends along the heavy chain from the N terminus, located between H1 and H3, toward the C terminus, which terminates at L3. (Right) Depiction of 4E10 variable domains, as described for Z13e1. In 4E10, the bound peptide extends from L3 along a groove in VH that terminates between H3 and H1. Z13e1 and 4E10 bind their peptide epitopes in reverse, resulting in the splaying of their respective light chains to opposite sides of the MPER (see panel B). (B) Superposition of Fab 2F5 and Fab 4E10 onto the Z13e1 MPER composite model. The three Fabs and the composite MPER model are shown as molecular surfaces and a cartoon representation, respectively. 2F5 and 4E10 are colored whitish gray, Z13e1 is colored blue, and the MPER is colored yellow. The superposition indicates that the predicted angles of the approach of 2F5 and 4E10 are too high and would lead to severe clashes with the membrane. The epitopes of 2F5 and 4E10 are, therefore, likely to be presented differently on the trimer than that of Z13e1, mediated by flexibility of the elbow region of the MPER. Alternatively, 2F5 and 4E10 may have binding modes, perhaps extracting their complete epitopes from the membrane (69), that are different from that of Z13e1, for which the postbinding manipulation of the MPER is less likely.

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