FIGURE 1:

Capu-NT inhibits the polymerization activity of Capu-CT. (A) Domain organization of Capu along with diagrams of the constructs used in this study. Previously characterized domains are shown as boxes, including FH1 and FH2 domains (orange and green, respectively) and the Capu-tail (violet). (B) Purified Capu-NT and truncations visualized on a Coomassie-stained SDS–PAGE gel. (C) Pyrene-actin polymerization assays with Capu-CT and Capu-NT. Polymerization of 4 μM actin (5% pyrene labeled) was induced by adding Mg²⁺ and KCl (see Materials and Methods). Addition of 9–300 nM Capu-NT (shown with increasing shades of blue) to 20 nM Capu-CT (alone in red) before mixing with actin inhibited the polymerization activity of Capu-CT in a dose-dependent manner. Addition of 225 nM Capu-NT to actin (green) does not change polymerization kinetics (black). (D) From the pyrene assay in C, the barbed-end concentration at time until half-maximal polymerization (t_1/2) was calculated and plotted for each concentration of Capu-NT. Data were fitted with a quadratic binding equation, yielding an inhibition constant (K_i) of 10 nM.