3.4 Structure of SARS-CoV-2 S glycoprotein trimer with upRBD bound to mvACE2 receptor
One of the three mvACE2-bound spike trimers captured had a strong density for mvACE2 bound to two of the RBD at 69.2°. After multiple rounds of additional 2D and 3D classification analysis, we determined the complex structure of mink S glycoprotein bound to two mvACE2 at 3.36 Å (Fig. 5A, S1B, and S1D). The local resolution of the protein interface between the complex was low due to its flexibility in the region. Nonetheless, the overall architecture of the mvACE2-mink S glycoprotein complex was comparable to the previously published structure. To visualize the key interaction at the protein interface between the complex, we performed local refinement with the mask covering the RBD-ACE2 interface, returning the improved resolution of the density map at 3.82Å (Fig. 5B, S1G, and S1I). To analyze the binding interface between the complex, we selected residues located less than 4.0 Å apart40 (Fig 5C).
Overall, our structure largely agreed with the previous Y453F RBD-mink ACE2 in interface residues8,9 (Fig. 5D-F). Notable interactions include a salt bridge between mink S D405 with mvACE2 H354 and S E484 with mvACE2 H79; hydrogen bond between S T500 with mvACE2 Y41, S G502 with mvACE2 K353, and S Y505 with mvACE2 R393; π-cation interaction between S Y489 with mvACE2 K31; and π-π stacking between S F453 with mvACE2 Y34 and S Y505 with mvACE2 H354. F453 in S glycoprotein is unique to the mink variant among other S variants (Fig. 3), and as indicated in our structural and binding affinity studies, as well as noted in previous studies8,9, Y453F mink S glycoprotein enhances the interaction with mvACE2 Y34.