The targeting of ion channels to particular membrane microdomains and their organization in macromolecular complexes allow excitable cells to respond efficiently to extracellular signals. In this study, we describe the formation of a complex that contains two scaffolding proteins: caveolin-3 (Cav-3) and a membrane-associated guanylate kinase (MAGUK), SAP97. Complex formation involves the association of Cav-3 with a segment of SAP97 localized between its PDZ2 and PDZ3 domains. In heterologous expression systems, this scaffolding complex can recruit Kv1.5 to form a tripartite complex in which each of the three components interacts with the other two. These interactions regulate the expression of currents encoded by a glycosylation-deficient mutant of Kv1.5. We conclude that the association of Cav-3 with SAP97 may constitute the nucleation site for the assembly of macromolecular complexes containing potassium channels.