Figures and data
Structure of the glycosylated head region of the ancestral SARS-CoV-2 spike with linoleate bound to the free fatty acid (FA) binding site.
(A) Model of the ectodomain of the glycosylated SARS-CoV-2 spike with linoleate (LA) bound. The spike-LA complex model was built using the cryo-EM structure 7JJI as a reference (23). Each monomer in the spike homotrimer is shown in a different colour: dark blue, light blue and green. The glycans are indicated with grey sticks, and LA molecules are highlighted with magenta spheres. Three FA binding sites exist in the trimer, each located at the interface between two neighbouring monomers. Note that in this model, all three receptor-binding motifs (RBMs) are in the ‘down’ conformation, and the protein is cleaved at the furin recognition site at the S1/S2 interface. (B) Detailed view of the FA binding site. This hydrophobic site is formed by two RBDs, with one providing the hydrophobic pocket for the FA hydrocarbon tail and the other providing polar (Q409) and positively charged (R408 and K417) residues to bind the negatively charged FA headgroup.
Structural response of the glycosylated spike to LA removal.
The average Cα displacements 0.1, 1 and 10 ns after LA removal from the FA binding sites are shown, mapped onto the starting structure for the equilibrium simulations. The norm of the average Cα displacement vector between the D-NEMD apo and equilibrium LA-bound simulations was calculated for each residue using the Kubo-Onsager relation (38, 39, 48, 49). The final displacement values are the averages obtained over the 210 pairs of simulations (Figures S6-S8). The cartoon thickness and structure colours (scale on the right) indicate the average Cα-positional displacement. Each RBD, NTD, furin site and FP are subscripted with their chain ID (A, B or C). Glycans are shown as light grey sticks, whereas the dark grey spheres highlight the position of the LA molecule. The FA site shown in this figure corresponds to FA site 1, which is located at the interface between chains C and A (see Figures S10-S11 for the responses of the other two FA sites, which are generally similar).
Structural responses of functional regions of the spike.
Close-up view of the structural response of the RBD (A), NTD (B) and FP surrounding regions (C) to LA removal. The FA site shown here is FA site 1, located at the interface between chains C and A (see Supplementary Figures S13-S14 for the responses of the other two FA sites, which are similar). Structure colours and cartoon thickness indicate the average Cα displacement values. Each region is subscripted with its chain ID (A, B or C). The dark grey spheres highlight the FA binding site. In the images representing the spike at t=0 ns (left side images A, B and C), the glycans are shown as light grey sticks, whereas the dark grey spheres highlight the FA binding site. Glycans are omitted from the panels showing the responses at t=0.1, 1 and 10 ns, but are present in the simulations. For more detail, see the legend of Figure 2.
Direction of the structural responses of the RBD and FP-surrounding regions to LA removal.
The average Cα displacement vectors at t=10 ns are shown. These vectors were determined by averaging Cα displacement vectors between the equilibrium and nonequilibrium trajectories over the 210 replicas. Vectors with a length ≥0.1 nm are displayed as blue arrows with a scale-up factor of 10. The average displacement magnitudes are represented on a white-yellow-orange-red scale. The dark grey spheres represent the FA site. This figure shows the direction of the responses around FA site 1, which is located at the interface between chains C and A (see Supplementary Figures S18-S19 for the direction of the motions in the other two FA sites).
D-NEMD analysis of displacement vectors shows connection between the FA site and the heme/biliverdin binding site in the NTD.
(A) View of the NTD 10 ns after LA removal, focusing on the heme/biliverdin binding site (56, 57) (which is not occupied in the simulations here). Note that the Q173-Q183 segment, which contains residues forming the heme/biliverdin binding site, shows an outward motion upon LA removal. The magnitudes of the displacements are represented on a white-yellow-orange-red colour scale. Vectors with a length ≥0.1 nm are displayed as blue arrows with a scale-up factor of 10. The dark grey spheres represent the FA site. This figure shows the direction of the structural responses around FA site 1 (see Figure S20 for the direction of the motions in the other two FA sites). (B) Cryo-EM structure showing the biliverdin binding site in the NTD (PDB code: 7NT9)(57). The protein is coloured in grey. The biliverdin molecules are shown with spheres.
