Figure 5
From: West Nile virus methyltransferase domain interacts with protein kinase G
The MTase domain interacts with PKGIβ through the αΕ helix. HEK293T cells were transfected with plasmid encoding wild-type PKGIβ-Flag and (a) WNV MTase or (b) WNV RdRp. Cell lysates were subjected to immunoprecipitation using α-Flag. The cell lysates and immunoprecipitated proteins were subjected to Western blot analysis using α-actin, α-WNV MTase, α-Flag (PKG), or α-HA (RdRp). (c) WNV MTase (wild-type (WT) or chimeric mutant with LGTV sequence at αΕ (Mut)) and LGTV MTase (WT) were purified onto Ni-NTA beads and incubated with 35S-labeled human PKGIβ-Flag in an in vitro binding assay. Beads were washed, and eluted proteins were separated by SDS-PAGE. An autoradiograpH(upper panel) shows PKG that precipitated with MTase. The lower panel is the Coomassie-stained SDS-PAGE gel.