The interplay between cardiac sarcoplasmic Ca2+ATPase and phospholamban is a key regulating factor of contraction and relaxation in the cardiac muscle. In heart failure, aberrations in the inhibition of sarcoplasmic Ca2+ATPase by phospholamban are associated with anomalies in cardiac functions. In experimental heart failure models, modulation of the interaction between these two proteins has been shown to be a potential therapeutic approach. The aim of our research was to find molecules able to interfere with the inhibitory activity of phospholamban on sarcoplasmic Ca2+ATPase. For this purpose, a portion of phospholamban was synthesized and used as target for a phage-display peptide library screening. The cyclic peptide C-Y-W-E-L-E-W-L-P-C-A was found to bind to phospholamban (1–36) with high specificity. Its functional activity was tested in Ca2+uptake assays utilizing preparations from cardiac sarcoplasmic reticulum. By synthesizing and testing a series of alanine point-mutated cyclic peptides, we identified which amino acid was important for the inhibition of the phospholamban function. The structures of active and inactive alanine-mutated cyclic peptides, and of phospholamban (1–36), were determined by NMR. This structure–activity analysis allowed building a model of phospholamban –cyclic peptide complex. Thereafter, a simple pharmacophore was defined and used for the design of small molecules. Finally, examples of such molecules were synthesized and characterized as phospholamban inhibitors.
Molecules able to interfere with the inhibitory activity of phospholamban (PLB) on SERCA2a were discovered. PLB was used as target for a phage display peptide library screening. The cyclic peptide C-Y-W-E-L-EW-L-P-C-A was found to bind to PLB and interfere with its activity. Its structure was determined and a model of this interaction with PLB was built. A pharmacophore was defined and used for the design of small molecules. Examples of such molecules were synthesized and characterized as phospholamban inhibitors.