Recent evidence indicates that a group of proteins can function as anchors for a variety of protein kinases. These anchoring proteins are proposed to be targeting subunits for the kinases which bring the enzymes within proximity of their physiological substrates. Targeting of protein kinases to discrete subcellular compartments would restrict access to potential substrates and render a degree of specificity of cellular responses to hormonal stimulation. The anchoring proteins may also function as regulatory subunits which could modify the catalytic properties of the kinases bound to them or they may function to anchor additional regulatory proteins and enzymes, other than kinases and phosphates to the cytoskeleton. Cyclic AMP-dependent protein kinase (PKA) is a multifunctional enzyme with a broad substrate specificity. PKA is ubiquitously expressed in all cells and it is responsive to a large variety of hormones and neurotransmitters. Type II PKA is associated with specific subcellular structures through the interaction of its regulatory (RII) subunit with A-kinase anchor proteins (AKAPs). During the past few years several AKAPs have been identified and cloned. All of these proteins appear to have the potential to form an amphipathic helix as predicted from computer assisted analysis of 14-18 amino acids which form part of the RII binding site. There is some reason to believe that the binding of RII to AKAPs is more complicated than just recognition of an amphipathic helix. We were never able to generate a peptide smaller than 29 amino acids of the MAP2 sequence which bound RII suggesting that more than the 14 amino acids proposed by Scott and his associates were involved in the binding reaction. Scanning mutagenesis studies on AKAP75 performed by Rubin and colleagues showed that residues containing long aliphatic side chains are essential for the high affinity binding of RII to AKAP75. Substitiution of Ala for Leu and Ile residues greatly diminished the RII-binding activity of AKAKP75. These substitutions were not predicted to significantly alter the hydrophobicity or helicity of the protein. Recently, we cloned testis-specific RII-binding protein which has no sequences that have a propensity to form an amphipathic helix as predicted by helical wheel analysis. These observations strongly suggest that helical structure, although potentially important, is not by itself the sole determining factor for high affinity interaction between RII, AKAP peptides and the truncated RII bound to AKAP peptides using NMR analyses can be a potentially strong method to solving the 3-D structure of the RII-AKPA interaction.