A CK2α' mutant indicating why CK2α and CK2α', the isoforms of the catalytic subunit of human protein kinase CK2, deviate in affinity to CK2β.

Protein kinase CK2 (casein kinase 2) mainly exists as heterotetrameric holoenzyme with two catalytic subunits (CK2α or CK2α') bound to a homodimer of non-catalytic subunits (CK2β). With and , the human genome contains two paralogs encoding catalytic CK2 subunits. Both gene products, called CK2α and CK2α', strongly interact with CK2β. An earlier report that CK2α' has a lower CK2β affinity than CK2α is confirmed via isothermal titration calorimetry in this study. Furthermore, we show with a fluorescence-anisotropy assay that a CK2β-competitive peptide binds less strongly to CK2α' than to CK2α. The reason for the reduced affinity of CK2α' to CK2β and CK2β competitors is puzzling: both isoenzymes have identical amino acid compositions at their CK2β interfaces, but the β4β5 loop, a component of this interface, is conformationally less adaptable in CK2α' than in CK2α due to intramolecular constraints. To release these constraints, we constructed a CK2α' mutant that was equalized to CK2α at the backside of the β4β5 loop. Concerning thermostability, affinity to CK2β or CK2β competitors and 3D-structure next to the β4β5 loop, this CK2α' mutant is more similar to CK2α than to its own wild-type, suggesting a critical role of the β4β5 loop adaptability for CK2β affinity.