cGMP-dependent protein kinase (PKG, cGPK) is a serine-threonine kinase of the AGC-kinase family. Although PKG type Ia plays a prominent role in the control of blood-flow and blood pressure, no current hypertension therapies target this enzyme.1 A PKG-targeted therapy would establish a first-in-class treatment for patients with uncontrolled blood pressure and may provide a clinically relevant alternative to existing antihypertensive therapies.
Dostmann et al. developed novel peptides derived from the crystal structure of PKG Ia, which are helical in solution and found to activate the kinase independent of cGMP and were demonstrated to lower blood pressure in vivo.2,3 Two consecutive phenylalanine residues were identified as a central element of the putative pharmacophore within the full-length peptide. However, a shortened peptide derivative, called S1.5 (here SP), was found to have increased potency and thus has been suggested as a potential lead compound for further development. The aim of this thesis is to reevaluate the pharmacophore and specifically the role of the phenylalanine residues in context of the SP peptide.
In this study, analogs of SP were designed using an alanine-scanning approach to analyze the role of specific amino acids in the activation mechanism. By employing recombinant PKG Ia, kinetic parameters were determined using a specific radiometric assay.4,5
We found that substituting either or both phenylalanine residues for alanine did not impair the peptide’s potency or efficacy. However, deleting one phenylalanine greatly diminished activity. This phenotype could be rescued by substituting either of two positively charged lysine residues N-terminal to the pharmacophore for alanine. These results indicate that the interactions between the kinase and these novel synthetic peptides are more complex and involve unidentified amino acids. Furthermore, the results presented here will serve to evaluate the role of N-terminal residues in peptide binding and the rescue phenotype discovered in this study
