Intermediate conformations of CD4-bound HIV-1 Env heterotrimers

Abstract

HIV-1 envelope (Env) exhibits distinct conformational changes in response to host receptor (CD4) engagement. Env, a trimer of gp120 and gp41 heterodimers, has been structurally characterized in a closed, prefusion conformation with closely associated gp120s and coreceptor binding sites on gp120 V3 hidden by V1V2 loops1–4 and in fully saturated CD4-bound open Env conformations with changes including outwardly rotated gp120s and displaced V1V2 loops3–9. To investigate changes resulting from substoichiometric CD4 binding, we solved single-particle cryo-electron microscopy (cryo-EM) structures of soluble, native-like heterotrimeric Envs bound to one or two CD4 molecules. Most of the Env trimers bound to one CD4 adopted the closed, prefusion Env state, with a minority exhibiting a heterogeneous partially open Env conformation. When bound to two CD4s, the CD4-bound gp120s exhibited an open Env conformation including a four-stranded gp120 bridging sheet and displaced gp120 V1V2 loops that expose the coreceptor sites on V3. The third gp120 adopted an intermediate, occluded-open state10 that showed gp120 outward rotation but maintained the prefusion three-stranded gp120 bridging sheet with only partial V1V2 displacement and V3 exposure. We conclude that most of the engagements with one CD4 molecule were insufficient to stimulate CD4-induced conformational changes, whereas binding two CD4 molecules led to Env opening in CD4-bound protomers only. The substoichiometric CD4-bound soluble Env heterotrimer structures resembled counterparts derived from a cryo-electron tomography study of complexes between virion-bound Envs and membrane-anchored CD4 (ref. 11), validating their physiological relevance. Together, these results illuminate intermediate conformations of HIV-1 Env and illustrate its structural plasticity.

HIV-1-envelope trimer transitions from prefusion-closed to CD4-bound-open conformations through an occluded-intermediate state

“…1-5). Recently, cryoET of Env in membranes by Li et al 15 unraveled the initial contact of Env with CD4, and cryoEM single particle analysis of engineered Env by Dam et al 14 Overall, our trajectory approach integrates Env dynamics and static structures, expanding our understanding of the structural features of Env trimers in the context of its transition process upon engagement with CD4 and the host (Supplementary Fig. 6).…”

“…Initially 178 trimers with complete gp120 and gp41 were collected and one was excluded due to computational modeling 35 (PDB: 3J70). In addition, ligand free HIV-1 Env trimer 19 (PDB: 4ZMJ), along with one and two CD4 bound HIV-1 Env trimers 14 (PDB: 8FYI, 8FYJ) were included, resulting in a total 180 PDBs participating in the analysis. All 180 PDBs were aligned with each time frame of transition pathway and RMSD analysis was performed using PyMOL (www.pymol.org).…”

“…Recent results from EM-tomograms of viral Env with CD4-VLP provide a context for these structures revealing initial contact to occur between a single-extended CD4 and Env trimer in its prefusion-closed conformation 13,14 when viral membrane and target membranes are ~150 Å apart; more extensive contacts and rearrangement of two or three CD4-binding protomers bound per Env trimer are observed when membranes are ~120 Å apart 15 .…”

Conformational flexibility of HIV-1 envelope glycoproteins modulates transmitted / founder sensitivity to broadly neutralizing antibodies

“…Our mechanistic insights are partially based on structure of SOSIP Env. Although recent studies of SOSIP conformational changes were consistent with data driven form Envs on virions 50,57 , the use of SOSIP may have underestimated Env opening as this soluble format was originally engineered to capture closed, prefusion Env conformation. We analyzed symmetry of only two SOSIPs purified via Galanthus nivalis lectin under identical conditions; we expect that analysis of SOSIP Envs of additional HIV-1 strains, which are planned for future studies, will reveal a range of asymmetries that may relate to the ability of different Envs to “breathe”.…”

Stoichiometry for entry and binding properties of the Env protein of R5 T cell-tropic HIV-1 and its evolutionary variant of macrophage-tropic HIV-1

“…Several structural studies have shown CD4 molecules binding to an Env trimer can give different conformations of the trimer depending on whether one, two, or three CD4 molecules are bound, with the implication being that a trimer must be fully occupied with CD4 to allow the subsequent conformational changes to occur, leading to viral membrane fusion ( 8 , 40 – 42 ). Our modeling of infectivity as a function of surface CD4 density for R5 T-tropic viruses is consistent with this observation in that there is a strong dependence on surface CD4 density for infectivity implying the need to interact with multiple CD4 molecules.…”