FIGURE 3.

EphrinB2 binding to NiV-G induces secondary structure changes in NiV-G that are correlated with fusogenicity. A, changes in the secondary structure of soluble G (blue) at 1.2 μm were monitored using CD spectroscopy upon the addition of B2 (red) at 2.5 μm. A theoretical spectrum is shown using a black dashed line, and the experimental G:B2 scan is shown in green. The difference between the latter two lines reflects differences in secondary structure. B and C, same experiment as in A, except replacing WT G for soluble 4-5 and GΔstalk proteins, respectively. The GΔStalk protein lacked amino acids 71–154 of the stalk domain. D, the difference between experimental and theoretical data (Δ [θ]) were generated by averaging the CD signals between 223 and 208 nm and subtracting the experimental signal (green line) coming from the indicated G and B2 mixture from the theoretical signal (black dashed line). Data are the average ± S.E., n = 3. E, soluble WT G, 4-5, or GΔStalk mutant proteins coating a maxisorb plate bound soluble B2 similarly in an ELISA enzyme-linked immunosorbent assay format assay. A sigmoidal dose response curve is shown (data are the average ± S.D., performed in triplicate).