Structure / function analysis of the interaction of PIP2 with actin capping protein: Implications for how capping protein binds the actin filament.
Kim et al. 2007. J. Biol. Chem. 282, 5871-5879. Online Journal
Abstract The heterodimeric actin-capping protein (CP) can be inhibited by polyphosphoinositides, which may be important for actin polymerization at membranes in cells. Here, we have identified a conserved set of basic residues on the surface of CP that are important for the interaction with phosphatidylinositol 4,5-bisphosphate (PIP2). Computational docking studies predicted the identity of residues involved in this interaction, and functional and physical assays with site-directed mutants of CP confirmed the prediction. The PIP2 binding site overlaps with the more important of the two known actin-binding sites of CP. Correspondingly, we observed that loss of PIP2 binding correlated with loss of actin binding among the mutants. Using TIRF (total internal reflection fluorescence) microscopy, we observed that PIP2 rapidly converted capped actin filaments to a growing state, consistent with uncapping. Together, these results extend our understanding of how CP binds to the barbed end of the actin filament, and they support the idea that CP can "wobble" when bound to the barbed end solely by the C-terminal "tentacle" of its beta-subunit.
Supplement Movie 1. Uncapping of capped actin filaments by PIP2. In this typical experiment, 30 images were collected over 60 min. PIP2 is added at the indicated time, and 5 images of text were added to mark that transition. In this movie, these 35 images are played at a rate of one per sec. Click to See Movie
