Interdomain interactions of radixin in vitro.

We have assayed the domains of the ERM protein radixin for binding activities in vitro. Affinity columns bearing the amino-terminal domain of radixin selectively bound a small subset of the proteins of the chicken erythrocyte cytoskeleton. Two of those proteins were identified as radixin itself and band 4.1. In contrast, the carboxyl-terminal domain of the molecule bound neither protein, and full-length radixin did not bind band 4.1 (binding of full-length radixin to itself was not evaluated). Columns bearing a mixture of the amino- and carboxyl-terminal domains of radixin also failed to bind radixin and band 4.1. These results suggested that the amino- and carboxyl-terminal sequences can interact with one another either in cis or in trans, and so interfere with radixin's interactions with other ligands. Using affinity co-electrophoresis, we confirmed a direct interaction in solution between the two radixin domains; the data are consistent with the formation of a 1:1 complex with a dissociation constant of approximately 5 x 10(-8) M. Competition between intramolecular and intermolecular interactions may help to explain the provocative and dynamic localization of ERM proteins within cells.