Cryo-EM structures of MERS-CoV and SARS-CoV spike glycoproteins reveal the dynamic receptor binding domains

Yuan, Yuan; Cao, Duanfang; Zhang, Yanfang; Ma, Jun; Qi, Jianxun; Wang, Qihui; Lu, Guangwen; Wu, Ying; Yan, Jinghua; Shi, Yi; Zhang, Xinzheng; Gao, George F.

NATURE COMMUNICATIONS | DOI: 10.1038/ncomms15092 ARTICLE obtained three distinguished class average images as references. A total number of B530,000 particles were picked in 1,810 micrographs and processed by no reference 2D classification using RELION program. About 260,000 particles in the good classes representing the S1/S2 trimers ( Supplementary Fig. 4a) were kept for further 3D classification. The HKU1 S trimer density map was low-pass filtered to 60 Ã… and rescaled as a reference map for 3D classification without imposing any symmetry. All the particles were classified into six classes and a 3D model within each class was reconstructed. Among the six reconstructions, two of them having the most accurate rotational alignment have reasonable rod-like densities in the middle representing central helices of S2. Class one containing about 55,000 particles has two RBDs in a standing state and one RBD in a lying state. Class two containing about 40,000 particles has one RBD in a standing state and two RBDs in a lying state. Further classification could not identify other conformations such as all three RBDs in a standing state or all three RBDs in a lying state, probably because of the small population of particles with these conformations. The rest part of the S protein monomer kept the same in a trimer. Thus, for better alignment, a 4.1 Ã… reconstruction with three-fold symmetry imposed containing all the particles from these two classes was calculated. However, the density of RBD region became quite low due to the average of RBD density between lying state and standing state. Further particle based motion correction and particle shinning process improve the resolution to 3.7 Ã… of the three-fold symmetry reconstruction by 0.143 criterion in the gold standard Fourier Shell correlated Coefficient (Supplementary Fig. 4d ). In addition, the shiny particles in class one and two were used to calculate a 4.1 and a 4.2 Ã… map without imposing any symmetry, respectively. The orientation distribution ( Supplementary Fig. 4c ) of MERS S protein trimer in the three-fold symmetry reconstruction was similar to that of HKU1 (ref. 27) . The local resolution of the three maps was calculated using program ResMap 47 .