TYPE-II REGULATORY SUBUNIT (RII) OF THE CAMP-DEPENDENT PROTEIN-KINASEINTERACTION WITH A-KINASE ANCHOR PROTEINS REQUIRES ISOLEUCINE-3 AND ISOLEUCINE-5

Compartmentalization of the type II cAMP-dependent protein kinase is m aintained by association of the regulatory subunit (RII) with A-Kinase Anchor Proteins (AKAPs). In previous studies (Scott, J. D., Stofko, R .E., McDonald, J. R., Comer, J. D., Vitalis, E. A., and Mangili J. (19 90) J. Biol. Chem. 265, 21561-21566) we have shown that dimerization o f RII alpha was required for interaction with the cytoskeletal compone nt microtubule-associated protein 2. In this report we show that the l ocalization and dimerization domains of RII alpha are contained within the first thirty residues of each RII protomer, RII des-5 (an amino t erminal deletion mutant lacking residues 1-5) was unable to bind AKAPs but retained the ability to dimerize. RII alpha I3,I5A (a mutant wher e isoleucines 3 and 5 were replaced with alanine) was unable to bind a variety of AKAPs. Mutation of both isoleucines decreased AKAP binding without affecting dimerization, cAMP binding, or the overall secondar y structure of the protein. Measurement of RII alpha I3A,I5A interacti on with the human thyroid AKAP, Ht 31, by two independent methods sugg ests that mutation of isoleucines 3 and 5 decreases affinity by at lea st 6-fold. Therefore, we propose that two isoleucine side chains on ea ch RII protomer are principle sites of contact with the conserved amph ipathic helix binding domain on AKAPs.