Rapid method for evaluation of CK2 enzymatic activity and CK2α/CK2β-interaction in Escherichia coli cell lysates

This study introduces a novel, rapid assay to measure CK2 alpha activity in Escherichia coli cell lysates. By fusing CK2 alpha with the fluorescent protein mScarlet it was possible to quantify CK2 alpha concentration directly in lysates. We used the dose-dependent increase of CK2 alpha activity after addition of CK2 beta 1-193 to determine the dissociation constants (K D ) of the CK2 alpha/CK2 beta-interaction. As a first trial, activity and affinity of the variant CK2 alpha R191Q to CK2 beta 1-193 was investigated using the developed assays. This mutation in the CSNK2A1 gene, encoding CK2 alpha is related to the Okur-Chung Neurodevelopmental Syndrome (OCNDS). Apparent K D values of 13 nM for the CK2 alpha R191Q/CK2 beta interaction and 7.4 nM for the CK2 alpha/CK2 beta interaction were determined using nonlinear regression. Uncertainties with regards to the concentration of both binding partners were propagated through the entire process of nonlinear regression by Monte Carlo simulations. This way, accuracy confidence intervals of the K D -values were derived. This resulted in 96.4 % confidence that the accurate K D -values of the CK2 alpha-CK2 beta and CK2 alpha R191Q-CK2 beta interactions were different. The results suggest potential disruptions in oligomeric assembly caused by the R191Q mutation.