FIG. 3.

Mapping of the VP22 phosphorylation sites. Bovine fibroblast cells were transfected with VP22 or its tyrosine-to-phenylalanine mutant constructs for 2 days. The His-tagged VP22 proteins were purified using Ni²⁺ columns. The partially purified proteins were analyzed by immunoblotting and probed with polyclonal antiphosphotyrosine (A) or anti-VP22 (B) antibody. Note that the tyrosine 38-to-phenylalanine mutation abolished the phosphorylation of VP22. (C) The blotted signals were quantified using NIH Image software, and the ratios of phosphorylation were normalized to the amount of VP22. Molecular masses are noted at the left in kilodaltons.