Broadening of Neutralization Activity to Directly Block a Dominant Antibody-Driven SARS-Coronavirus Evolution Pathway (original) (raw)
Figure 2
Neutralization of pseudotyped viral infection by anti-GD03 Abs and epitope mapping of GD03 nAb 11A.
A, Five anti-GD03 Abs isolated by phage Ab library screening against purified GD03-RBD were analyzed for neutralizing activity using GD03 (Left panel) or Tor2 (Right panel) spike pseudoyped viruses. B, Kinetic characterization of GD03-RBD binding to 11A-IgG1. Abs were captured on a CM4 chip via immobilized anti-human IgG1 on the chip. GD03-RBD at various concentrations (2 folds serial dilutions, highest concentration was indicated) was injected over the chip surface. Binding kinetics was evaluated using a 1∶1 interaction model. In each panel, the binding response curves (red lines) are overlaid with the fit of the interaction model (black lines). All ka, kd, KD value showed in the table represent the means and standard errors of three experiments. C, Competition of 11A for the binding of GD03-RBD-C9 to 293T-ACE2 cells. GD03-RBD-C9 or control-RBD-C9 (Filled purple) used for staining is at 20 ug/mL and the Abs (256 or 80R negative control) were used at 50 ug/mL to compete for the binding of GD03-RBD to 293T-ACE2 cells. D, Epitope mapping of 11A. Purified proteins of a set of GD03-RBD or Tor2-RBD mutants were coated to ELISA plates at indicated concentrations. 2 µg/ml of 11A-IgG1 followed by HRP-anti-human IgG1 were used to detect the binding of 11A with different mutants. E, Interface of structure of the Tor2- RBD/80R complex 21. S1-RBD is in yellow. CDR loops (H1-H3 and L1-L3) of 80R, amino acids of 80R-CDRL1 and L2 (162–164 and 182) as well as 472L and 480D of S1-RBD are colored as shown.