FIGURE 2.

SpyA preferentially modifies vimentin. Equimolar amounts of recombinant hamster vimentin and bovine actin (877 nm) were incubated with SpyA (0.5 μm) and biotinylated NAD⁺ (A) or [³²P]NAD⁺ (B) for the indicated time. Reactions were subjected to 8–16% SDS-PAGE. A, the gel was transferred to a nitrocellulose membrane and probed for biotin with streptavidin-HRP. B, the gel was subjected to phosphorimaging, and the resulting bands were quantitatively analyzed with ImageJ software. The band intensities in arbitrary units (AU) were plotted against time. Data presented are for a representative experiment. Error bars, S.D. Three independent experiments were conducted.