Structural and molecular analysis of a cGMP-Kinase/ IRAG1-Macrocomplex

• Applicants: Dr. Leonhard Jakob; Professor Dr. Jens Schlossmann
• Subject Area: Pharmacology
• Term: since 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 547855469

Signaling by NO/cGMP/cGMP-dependent kinase (PKG) regulates various (patho)physiological processes e. g. in the vascular and gastrointestinal system and in the hemostasis. For the PKG 1-isoform PKG 1ß was a ternary macro-complex identified which additionally consists of the inositoltrisphosphate receptor Type I (IP3R1) and the IP3R1-associated cGMP-Kinase-substrate protein (IRAG1). This complex modulates intracellular calcium and thereby regulates e. g. relaxation of smooth muscle and the inhibition of platelet aggregation. However, the structural and functional characteristics of the ternary complex and the dynamics of the macro-complex to inhibit the IP3R1 are unknown so far. The aim of the project is the detailed elucidation of the cGMP-kinase/IRAG1-macrocomplex to comprehend the structure and dynamic of the complex and thereby its regulation of the IP3R1-calcium channel. The structure of the purified complex will be analyzed by cryo-electron microscopy and compared in its basal and active form. Finally, it will be investigated whether an IRAG1-mutation, which is caused by a polymorphism and is associated with a vasculopathy, alters the molecular structure of the complex and thereby leads to a functional defect. The dynamics of PKG 1 and of the complex in the basal and active state will tested by single molecule-FRET-analysis (SLAM-FRET). With the applied project we expect to get a detailed information of the structural foundation and the dynamic effects of the NO/cGMP/PKG 1ß-signaling cascade und hence of its (patho)physiological actions.

• DFG Programme: Research Grants