title: Inclining the purine base binding plane in protein kinase CK2 by exchanging the flanking side-chains generates a preference for ATP as a cosubstrate.
abstract: Protein kinase CK2 (casein kinase 2) is a highly conserved and ubiquitously found eukaryotic serine/threonine kinase that plays a role in various cellular key processes like proliferation, apoptosis and circadian rhythm. One of its prominent biochemical properties is its ability to use GTP as well as ATP as a cosubstrate (dual-cosubstrate specificity). This feature is exceptional among eukaryotic protein kinases, and its biological significance is unknown. We describe here a mutant of the catalytic subunit of protein kinase CK2 (CK2alpha) from Homo sapiens (hsCK2alpha) with a clear and CK2-atypical preference for ATP compared to GTP. This mutant was designed on the basis of several structures of CK2alpha from Zea mays (zmCK2alpha) in complex with various ATP-competitive ligands. A structural overlay revealed the existence of a "purine base binding plane" harbouring the planar moiety of the respective ligand like the purine base of ATP and GTP. This purine base binding plane is sandwiched between the side-chains of Ile66 (Val66 in hsCK2alpha) and Met163, and it adopts a significantly different orientation than in prominent homologues like cAMP-dependent protein kinase (CAPK). By exchanging these two flanking amino acids (Val66Ala, Met163Leu) in hsCK2alpha(1-335), a C-terminally truncated variant of hsCK2alpha, the cosubstrate specificity shifted in the expected direction so that the mutant strongly favours ATP. A structure determination of the mutant in complex with an ATP-analogue confirmed the predicted change of the purine base binding plane orientation. An unexpected but in retrospect plausible consequence of the mutagenesis was, that the helix alpha D region, which is in the direct neighbourhood of the ATP-binding site, has adopted a conformation that is more similar to CAPK and less favourable for binding of GTP. These findings demonstrate that CK2alpha possesses sophisticated structural adaptations in favour of dual-cosubstrate specificity, suggesting that this property could be of biological significance.
authors.affiliation: Syddansk Universitet, Institut for Biokemi og Molekylaerbiologi, Campusvej 55, DK-5230 Odense, Denmark.
authors.forename: Christina W
authors.initials: CW
authors.lastname: Yde
authors.forename: Inessa
authors.initials: I
authors.lastname: Ermakova
authors.forename: Olaf-Georg
authors.initials: OG
authors.lastname: Issinger
authors.forename: Karsten
authors.initials: K
authors.lastname: Niefind
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D047390:Casein Kinase II / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011687:Purines / Q000378:metabolism*; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D021122:Protein Subunits; D011687:Purines; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D047390:Casein Kinase II