Rational design of a selective antagonist of ε protein kinase C derived from the selective allosteric agonist, pseudo-RACK peptide

We have previously shown that domains involved in binding of protein kinase C (PKC) isozymes to their respective anchoring proteins (RACKs) and short peptides derived from these domains are PKC isozyme-selective antagonists. We also identified PKC isozyme-selective agonists, named psi RACK peptides, derived from a sequence within each PKC with high homology to its respective RACK. We noted that all the psi RACK sequences within each PKC isozyme have at least one non-homologous amino acid difference from their corresponding RACK that constitutes a charge change. Based on this information, we have devised here a new approach to design an isozyme-selective PKC antagonist, derived from the psi RACK sequence. We focused on epsilon PKC psi RACK peptide, where the pseudo-epsilon RACK sequence (psi epsilon RACK; H (D) under bar APIGYD; corresponding to epsilon PKC85-92) is different in charge from the homologous RACK-derived sequence (N (N) under bar VALGYD; corresponding to epsilon RACK285-292) in the second amino acid. Here we show that changing the charge of the psi epsilon RACK peptide through a substitution of only one amino acid (aspartate to asparagine) resulted in a peptide with an opposite activity on the same cell function and a substitution for aspartate with an alanine resulted in an inactive peptide. These data support our hypothesis regarding the mechanism by which pseudo-RACK peptide activates PKC in heart cells and suggest that this approach is applicable to other signaling proteins with inducible protein-protein interactions. (c) 2007 Elsevier Inc. All rights reserved.