Fig. 2.

The amino acid position 43 is critical for spike binding onto tracheal tissue. Binding of recombinant S1 spike proteins on tissue sections of mature tracheal tissue was examined using protein histochemistry (A). Four S1 spike proteins were used for the binding experiments namely, the primary ArkDPI S1 spike (DPI), the ArkDPI spike with the Y43H change (Y43H), the ArkDPI spike with a deletion at position 344 (∆344), and finally, the ArkDPI spike with both Y43H and ∆344 changes. StrepTactin-tagged GFP protein (GFP-Strep) was used as a negative control. To detect binding, we pre-complexed the S1 spike proteins with horseradish peroxidase-conjugated StrepTactin and developed a brown signal using DAB as a substrate. Quantification of the signals was performed by densitometry using the ImageJ software (B). Highest binding to ciliated epithelium of the trachea was observed for Y43H spike. Statistical analysis was done in GraphPad Prism software.*Indicates p <0.05 when compared to other groups.