Binding of dystrophin's tandem calponin homology domain to F-actin is modulated by actin's structure

Dystrophin has been shown to be associated in cells with actin bundles, Dys -246, an N-terminal recombinant protein encoding the first 246 residues of dystrophin, includes two calponin-homology (CH) domains, and is similar to a large class of F-actin cross-linking proteins including cr-actinin, fimbr in, and spectrin, It has been shown that expression or microinjection of am ino-terminal fragments of dystrophin or the closely related utrophin result ed in the localization of these protein domains to actin bundles. However, in vitro studies have failed to detect any bundling of actin by either inta ct dystrophin or Dys-246. We show here that the structure of F-actin can be modulated so that there are two modes of Dys-246 binding, from bundling ac tin filaments to only binding to single filaments, The changes in F-actin s tructure that allow Dys-246 to bundle filaments are induced by covalent mod ification of Cys-374, proteolytic cleavage of F-actin's C-terminus, mutatio n of yeast actin's N-terminus, and different buffers. The present results s uggest that F-actin's structural state can have a large influence on the na ture of actin's interaction with other proteins, and these different states need to be considered when conducting in vitro assays.