FIG 13.

Identification of the domain of vimentin involved in the interaction with EV71 VP1. (A) Analysis of various truncated vimentin fragments for interaction with VP1. The His-tagged vimentin and its fragments (aa 1 to 466, 1 to 230, 231 to 466, 1 to 115, 116 to 230, 1 to 56, and 57 to 115) were subjected to a pulldown assay using the GST-VP1-coupled glutathione-Sepharose beads or GST beads as described in Materials and Methods. Proteins coprecipitating with the beads were analyzed by immunoblotting using anti-His antibody or anti-GST antibody. (B) Effect of pretreating EV71 inoculum with full-length vimentin and truncated vimentin on the binding of the virus to U251 cells. EV71 was pretreated with vimentin fragments at a concentration of 20 μg ml⁻¹ at 37°C for 1 h prior to infecting U251 cells. The cells were then infected with EV71 at 4°C for 1 h. Quantitative RT-PCR analysis was then performed to determine the EV71 RNA levels as described in Materials and Methods. Control, cells infected with untreated EV71.