Strong evolutionary convergence of receptor-binding protein spike between COVID-19 and SARS-related coronaviruses

Yonghua Wu

Given the selection intensification of S gene in 2019-nCoV since its evolutionary divergence from RaTG13, we conducted comparative sequence analyses between the two. We found that there were more than 20 amino acid differences between them, and most of them were located within RBD, especially RBM (Fig. S2) , suggesting a high variability of RBM. Given the importance of RBM for receptor binding, we further conducted a comparative sequence analysis among all the coronavirus strains studied to examine its variability. The results showed high sequence variability, with insertion and/or deletion and amino acid substitutions among the coronavirus strains studied (Fig. 2) . Despite the high variability of RBM, strikingly, we found that SARS-CoV and its phylogenetic relatives-including WIV16 (KT444582), Rs4231 (KY417146), Rs7327 (KY417151), Rs9401 (KY417152), and BtRs-BetaCoV/YN2018B (MK211376), called SARS-related CoV here, shared many identical or nearly identical amino acids with their phylogenetically distant coronavirus strains, including 2019-nCoV and RaTG13, which we called COVID-19-related CoV (Fig. 2) . These shared amino acids were clearly distinct from bat SARS-like CoV that were phylogenetic intermediates between them (Fig. 2) . Further analyses showed that such identical amino acids shared between SARS-related CoV and COVID-19-related CoV were not restricted to RBM, but rather, they were scattered throughout the spike protein, with a total of 32 such sites, which were centered on RBD (28 sites in total, Fig. S3 ). To further examine whether such similarity occurred in other proteins, we analyzed all 11 genes studied among these coronaviruses, and we found that one additional gene, ORF3a, contained eight such sites (Fig. 1) . The existence of these shared amino acids between SARS-related CoV and COVID-19-related CoV may suggest their high sequence similarity. In support of this, we reconstructed maximum likelihood and neighbor-joining phylogenies using full-length RBD protein sequences, and both showed that SARS-related CoV and COVID-19-related CoV were grouped in the same clade, with relatively high support, which is consistent with two previous studies 6,7 , at the same time, their phylogenetic intermediates were clustered in distinct clades (Fig. 3, Fig. S4 ). The phylogenetic uniting of SARS-related CoV and COVID-19-related CoV provide evidence of their high similarity of RBD protein sequences.