Structure and function of palladin's actin binding domain

Abstract

Here, we report the NMR structure of the actin-binding domain contained in the cell adhesion protein palladin. Previously, we demonstrated that one of the immunoglobulin domains of palladin (Ig3) is both necessary and sufficient for direct filamentous actin binding in vitro. In this study, we identify two basic patches on opposite faces of Ig3 that are critical for actin binding and cross-linking. Sedimentation equilibrium assays indicate that the Ig3 domain of palladin does not self-associate. These combined data are consistent with an actin cross-linking mechanism that involves concurrent attachment of two actin filaments by a single palladin molecule by an electrostatic mechanism. Palladin mutations that disrupt actin binding show altered cellular distributions and morphology of actin in cells, revealing a functional requirement for the interaction between palladin and actin in vivo.

Keywords: 3D; 4′,6-diamidino-2-phenylindole; ABP; DAPI; F-actin; FLNa; G-actin; GFP; HSQC; PDB; Protein Data Bank; RASREC; VASP; actin binding protein; actin-binding protein; crosslinking; electrostatics; filamentous actin; filamin A; globular or monomeric actin; green fluorescent protein; heteronuclear single quantum correlation; immunoglubulin-like domain; palladin; resolution-adapted structural recombination; three-dimensional; vasodilator-stimulated phosphoprotein.