FIGURE 5.

Revertant analysis of cavity B mutant virus, I863A. A, plaque morphology and sequencing chromatogram from the I863A P2 and P7 viruses. The engineered mutant nucleotides are underlined. B, validation of the I863V revertant mutation. The I863V genome-length RNA was electroporated into BHK-21 cells. The transfected cells were monitored for E protein expression at 24, 48, and 72 h post-transfection. Plaque morphology of the I863V virus (P0) is shown. C, comparison of de novo RdRp activities among WT, I863V, and I863A NS5 proteins. The relative de novo RdRp activities (WT set as 100%) are indicated below the denaturing gel. D, comparison of elongation RdRp activities among WT, I863V, and I863A NS5 proteins. The relative elongation activities were shown in parentheses (WT set as 100%). Results are the average of two independent experiments performed in duplicate.