Exploration and Exploitation of Secondary Sites of Protein Kinase CK2

• Applicants: Professor Dr. Joachim Jose; Professor Dr. Karsten Niefind
• Subject Area: Structural Biology ; Biochemistry
• Term: since 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 505676396

Protein kinase CK2, an acidophilic, pleiotropic and highly conserved member of the eukaryotic protein kinase superfamily, mainly exists as a heterotetrameric holoenzyme consisting of a central dimer of regulatory subunits (CK2β) with two attached catalytic chains. In humans, two paralogous isoforms of the catalytic CK2 subunit – designated as CK2α and CK2α’ – are present. CK2 is upregulated in many types of cancer and thus an attractive pharmaceutical target. The typical target region for inhibition of CK2 and other protein kinases is the ATP cavity, therefore often referred to as “primary” site. Due to the conservation of the ATP site, ATP-competitive inhibitors are limited in their selectivity. A strategy to overcome this issue is the use of a “secondary” binding site that can be addressed either alone or in combination with the ATP site by a so-called bivalent inhibitor. Typical secondary sites are the substrate binding region and allosteric sites located remote of the active centre. Allosteric sites can be preformed or “cryptic”, i.e. not open in the apo-form and becoming accessible only in the presence of a suitable ligand. An attractive secondary site of CK2α and CK2α’ is the αD pocket, a cryptic site accessible only after conformational adaptations of the helix αD region. In the XPLOR_CK2 project, the αD pocket will be exploited to construct highly potent, selective and cell permeable bivalent CK2 inhibitors with indenoindole-based ATP-competitive anchor groups. If possible, the selectivity shall be boosted even to a level that allows the distinction of the two isoforms CK2α and CK2α’. Selected bivalent inhibitors from these activities will be used to determine high-resolution crystal structures of the two CK2 holoenzymes based on either CK2α or CK2α’. Further secondary sites in the focus of the XPLOR_CK2 project are the substrate recognition region, the heparin site and the N-terminal segment site. A recently described substrate peptide will be used to solve the first CK2α/CK2α’ structure with a visible peptide substrate attached to the substrate binding region. The heparin site was discovered in CK2α; now, it shall be detected in CK2α’ as well and exploited to construct indenoindole-based bivalent inhibitors, which address parts of the heparin site. The N-terminal segment site is new and cryptic: it was unexpectedly found in CK2α’ crystals after soaking with a putative bisubstrate inhibitor. In ExExCK2, its existence shall be confirmed with CK2α and in a non-crystalline state; in case of validation, its overlap with the substrate recognition region shall be exploited to generate a selective substrate-competitive CK2 inhibitor.

• DFG Programme: Research Grants
• International Connection: France
• Partner Organisation: Agence Nationale de la Recherche / The French National Research Agency
• Cooperation Partner: Professor Dr. Marc Le Borgne