Fig. 5: Partially open early intermediates and a stepwise mechanism for CD4-induced Env opening.

A CD4,VRC34.01-bound BG505 SOSIP Env with rotation, bridging sheet (red) and α0 helix (green) formation observed in a single gp120. B CD4,VRC34.01-bound BG505 SOSIP Env with rotation, bridging sheet (red) and α0 helix (green) formation observed in two gp120 subunits. C A structure-guided mechanism for stepwise Env opening along the HIV-1 entry pathway. Top panel structures were determined previously, bottom panel structures were determined in this study. Stepwise transitions are marked with numbers within a circle on top of each structure starting from (1) the binding of a single CD4 to a closed Env trimer (PDB: 5U1F, 8FYI). This is followed by (2) opening of the Env trimer (EMD-29292) that allows additional CD4 molecules to bind. (5) A partially open Env conformation was described bound to CD4, a coreceptor (Co-R) mimicking antibody, and the gp120/gp41 interface targeting antibody 8ANC195 (PDB: 6CM3, 6EDU) where the FP was buried within a gp41 cavity. The CD4-induced opening of the HIV-1 Env culminates in the complete rotation of all the gp120 subunits that are accompanied by gp41 conformational changes and resulting in the burial of FP. This state is numbered (8) in this schematic. This study showed that the geometry of the functional entry intermediate (5) that was also visualized on membrane-associated Env (EMD-29294), was compatible with the FP being either buried or exposed, and thus, in this conformation the FP was accessible to antibodies. Further, this study filled in mechanistic gaps between (2) and (5) by showing stepwise gp120 rotations to reach this functional entry intermediate. Finally, this study visualized a stepwise mechanism for how the functional entry intermediate (5) may transition to the fully open Env (8), yet again by stepwise opening of the each gp120 subunit from its partially rotated to the fully rotated conformation, which was accompanied by burial of the FP in the corresponding protomer. Schematics of membranes and the host receptors were created in BioRender. Acharya, P. (2025) https://BioRender.com/7tpbllk.