Fig 4.

Binding of CHIKV and SFV DIII proteins to membrane-inserted CHIKV DI/II core trimers. CHIKV DI/II protein (50 μg/ml) was mixed with liposomes and a 1:1 molar ratio of the indicated SFV (A) or CHIKV (B) DIII proteins, with truncations and nomenclature as shown in Fig. 5A. The samples were treated at pH 5.75 at room temperature for 30 min, adjusted to pH 8.0, and separated on sucrose flotation gradients. Gradients were fractionated and pooled into the top (T), middle (M), and bottom (B) fractions in panel A or into the top (T) and bottom (B) fractions in panel B. Pooled fractions were acid precipitated and analyzed by SDS-PAGE and Western blotting to detect E1 DI/II and DIII as indicated. Both panels are representative examples of 3 independent experiments, with the percentage of DIII protein in the top fraction (average ± standard deviation) listed below each panel.