INTERDOMAIN INTERACTIONS OF RADIXIN IN-VITRO

We have assayed the domains of the ERM protein radixin for binding act ivities in vitro. Affinity columns bearing the amino-terminal domain o f radixin selectively bound a small subset of the proteins of the chic ken erythrocyte cytoskeleton. Two of those proteins were identified as radixin itself and band 4.1. In contrast, the carboxyl-terminal domai n of the molecule bound neither protein, and full-length radixin did n ot bind band 4.1 (binding of full-length radixin to itself was not eva luated). Columns bearing a mixture of the amino- and carboxyl-terminal domains of radixin also failed to bind radixin and band 4.1. These re sults suggested that the amino- and carboxyl-terminal sequences can in teract with one another either in cis or in trans, and so interfere wi th radixin's interactions with other ligands. Using affinity co-electr ophoresis, we confirmed a direct interaction in solution between the t wo radixin domains; the data are consistent with the formation of a 1: 1 complex with a dissociation constant of similar to 5 x 10(-8) M. Com petition between intramolecular and intermolecular interactions may he lp to explain the provocative and dynamic localization of ERM proteins within cells.