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. 2013 Jul 23;110(30):12438-43.
doi: 10.1073/pnas.1307382110. Epub 2013 Jun 11.

Molecular architecture of the uncleaved HIV-1 envelope glycoprotein trimer

Youdong Mao  1 , Liping Wang, Christopher Gu, Alon Herschhorn, Anik Désormeaux, Andrés Finzi, Shi-Hua Xiang, Joseph G Sodroski
Affiliations

Molecular architecture of the uncleaved HIV-1 envelope glycoprotein trimer

Youdong Mao et al. Proc Natl Acad Sci U S A. .

Abstract

The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer, a membrane-fusing machine, mediates virus entry into host cells and is the sole virus-specific target for neutralizing antibodies. Binding the receptors, CD4 and CCR5/CXCR4, triggers Env conformational changes from the metastable unliganded state to the fusion-active state. We used cryo-electron microscopy to obtain a 6-Å structure of the membrane-bound, heavily glycosylated HIV-1 Env trimer in its uncleaved and unliganded state. The spatial organization of secondary structure elements reveals that the unliganded conformations of both glycoprotein (gp)120 and gp41 subunits differ from those induced by receptor binding. The gp120 trimer association domains, which contribute to interprotomer contacts in the unliganded Env trimer, undergo rearrangement upon CD4 binding. In the unliganded Env, intersubunit interactions maintain the gp41 ectodomain helical bundles in a "spring-loaded" conformation distinct from the extended helical coils of the fusion-active state. Quaternary structure regulates the virus-neutralizing potency of antibodies targeting the conserved CD4-binding site on gp120. The Env trimer architecture provides mechanistic insights into the metastability of the unliganded state, receptor-induced conformational changes, and quaternary structure-based strategies for immune evasion.

Keywords: cryo-EM; membrane protein; retrovirus; spike; vaccine immunogen.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Architecture of the HIV-1 Env trimer. (A) Cryo-EM map of the HIV-1JR-FL Env trimer in a surface representation, viewed from a perspective parallel to the viral membrane. (B) Cryo-EM map of the HIV-1JR-FL Env trimer, viewed from the perspective of the target cell. (C and D) Domain organization of the Env protomer, revealed by segmentation of the density map. The gp120 domains are colored as follows: outer domain, blue; inner domain, orange; and TAD, red. The gp41 domains are colored as follows: ectodomain, green; and transmembrane region, cyan.
Fig. 2.
Fig. 2.
The unliganded gp120 subunit and CD4-induced changes. (A and B) Crystal structure of the outer domain of the gp120 core (17) was fitted into the cryo-EM map of the unliganded Env trimer and is shown from two perspectives parallel to the viral membrane. The approximate location of the V4 variable region, which was not resolved in the crystal structure, is indicated by a broken line. (C and D) Secondary structure organization of the inner domain of gp120 was approximated in the cryo-EM map of the unliganded Env trimer, which is viewed from two perspectives parallel to the viral membrane. (E and F) For comparison of the unliganded precursor state (E) and the CD4-bound state (F) of the gp120 core, the gp120 outer domains (blue) are aligned in the same orientation. The gp120 core in the unliganded precursor state is derived as described in A–D above, and the CD4-bound gp120 core structure is from an X-ray crystal structure (Protein Data Bank ID: 3JWD).
Fig. 3.
Fig. 3.
Architecture of the gp120 TAD. (A) Secondary structure elements in the gp120 TAD, viewed from a perspective perpendicular to the Env trimer axis. The TAD comprises an α-helix (red), a minibarrel structure (green), and a β-sheet-like element (yellow). (B) Three helical elements (red) pointing toward the central minibarrel structure (green), viewed from the center of mass of the trimer. (C) The TAD, viewed from the perspective of the target cell. The points at which the V1/V2 stem and V3 loop enter the TAD from the gp120 inner domain and outer domain, respectively, are indicated. The map segmentations of the gp120 TAD are shown as blue meshwork for a lower level of contour and as solid surfaces for slightly higher levels of contour. The potential α-helical elements shown as worm tubes are schematically illustrated and are not intended to represent definitive backbone traces.
Fig. 4.
Fig. 4.
gp41 subunit in the Env trimer. (A) Cryo-EM map of the three gp41 subunits in the Env trimer, viewed from an angle of ∼30° with respect to the viral membrane. (B) gp41 membrane-interactive region viewed from a perspective parallel to the viral membrane. (C) gp41 ectodomain, viewed from the perspective of the virus. The torus-like topology of the three gp41 subunits in the Env trimer is evident from this perspective. (D) gp41 ectodomain, viewed from a perspective parallel to the viral membrane. (E) Comparison of the gp41 ectodomain in the unliganded precursor state and the postfusion state with respect to the tertiary organization of helices. The perspective is identical to that shown in D, with a reduction in scale. For simplicity, only one of the three gp41 glycoprotein subunits in the Env trimer is depicted.
Fig. 5.
Fig. 5.
Env glycosylation. (A and B) Glycan-associated densities on the gp120 surface, viewed from two perspectives parallel to the viral membrane. The gp120 outer domain ribbon is colored blue and the asparagine residues associated with potential N-linked glycosylation sites are shown in Corey-Pauling-Koltun (CPK) representation and labeled. The blue meshwork shows the overall map segment associated with the gp120 outer domain. The yellow densities highlight the potential glycan-associated map segments in a surface representation. (C) Schematic showing the five glycan core residues associated with N-linked glycosylation. The trimannosyl core, which is common to high-mannose, complex, and hybrid oligosaccharides, is colored.
Fig. 6.
Fig. 6.
Effect of quaternary structure on virus neutralization by CD4BS antibodies. (A–E) Crystal structures of the gp120 core in complex with the CD4BS antibodies, VRC01 (A), VRC03 (B), b12 (C), b13 (D), and F105 (E) were superposed on the unliganded Env precursor map, with the Env trimer axis shown in A. The conformationally rigid gp120 outer domain was fitted to the Env trimer density map, and the outer domains in the crystal structures were aligned with the fitted outer domain (represented as a blue ribbon). The heavy and light chains of the CD4BS antibodies are colored red and yellow, respectively. The dashed red circle in C–E marks the approximate diameter of the space in the neighboring gp120 subunit that is overlapped by the antibodies, highlighting the quaternary steric hindrance that the antibodies experience when approaching their binding site on the gp120 outer domain. (F) Using the volume of the quaternary clash in the case of b12 as a normalization metric, the degree of quaternary steric hindrance encountered by each antibody as it accesses the Env trimer was quantified and was plotted against the geometric mean IC50 of the neutralization of multiple HIV-1 strains by the antibody (–, –45). The Protein Data Bank IDs of the antibodies in complex with gp120 core structures are 3NGB (A), 3SE8 (B), 2NY7 (C), 3IDX (D), and 3HI1 (E). The red line shows a linear regression of the relationship (rS: 0.9985, slope: 5.5, SE: 0.082).
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