Fig. 3

Single amino acid substitution mutagenesis of the SARS-CoV-2-RBD and SARS-CoV-RBD. a Sequence differences in the SARS-CoV and SARS-CoV-2 RBDs. RBM is in red. Previously, identified critical ACE2-binding residues are shaded in green. The conserved residues are marked with asterisks (*), the residues with similar properties between groups are marked with the colon symbol (:) and the residues with marginally similar properties are marked with the period symbol (.). b ACE2 binding with reciprocal amino acid substitutions in the SARS-CoV-2 RBD. Each value is calculated as the binding relative to that of the WT (%). The mean±S.E.M. of duplicate wells is shown for two independent experiments. The two red dotted lines represent 75% and 125% relative to the WT data, respectively. c, d Structural alignment of SARS-CoV-2-RBD and SARS-CoV-RBD binding with ACE2. The SARS-CoV-RBD complex (PDB ID: 2AJF) is superimposed on the SARS-CoV-2 RBD (PDB ID: 6lzj. grey: ACE2, wheat: SARS-CoV-2. Mutants that weaken the SARS-CoV-2 RBD binding with ACE2 are highlighted in cyan (c). The corresponding residues from SARS-CoV are indicated in green and are illustrated in detail (c left). Mutants that enhance ACE2 binding are highlighted in magenta (d). e ACE2 binding with reciprocal amino acid substitutions in the SARS-CoV RBD. Each value is calculated as the binding relative to that of the WT (%). The mean ± S.E.M. of duplicate wells is shown in two independent experiments. The two red dotted lines represent 75 and 125% relative to the WT data, respectively. f Molecular docking of the SARS-CoV 2 RBD carrying the Q498Y mutant in complex with hACE2. Q498Y formed π-π stacking with Y41 in hACE2: left, Y498; right, Q498