Fig. 5: Confirmation and biochemical characterization of the R3DC23 epitope.

a, b Identification of the R3DC23 binding region on recombinant spike protein by HDX-MS. The panels in A show the HDX-MS uptake plots of the two peptides with high degrees of protection from deuteration in the presence of R3DC23. The asterisk indicates the glycosylated Asn residue. b Difference HDX uptake plot (Woods Plot). Each line is a peptide and the residues spanned by the peptide are indicated by the positioning along the x-axis. The position along the y-axis is the difference in exchange between the apo condition and the R3DC23 bound condition (apo – R3DC23 bound) at an individual exchange time. The color of each line indicates the exchange time. c R3DC23 binding to cells that express the SARS-CoV-2 spike (left panel) or HR2 (right panel) at their surface. d R3DC23 does not recognize cells that express the SARS-CoV-2 6HB or 5HB at their surface. e R3DC23 binds to trimerized HR2 coiled-coil constructs. The graphs show binding of huR3DC23-Fc (left panel) or palivizumab (right panel) to various HR2 constructs, that are fused to His-SUMO (monomeric) or foldon (trimeric). f R3DC23 VHHs bind with a 3:1 stoichiometry to recombinant SARS-CoV-2 spike trimers (S-2P). The BLI sensorgram shows the immobilization of S-2P trimer on a biolayer interferometry biosensor followed by association with 20 nM R3DC23 monomer in solution. Raw data (no reference subtraction) of one of triplicate experiments is shown. The sensongrams of the two other runs are shown in Supplementary Fig. 6D. Source data are provided as a source data file.