Several approaches have been used to study the interactions of the subunits of protein kinase CK2. The inactive mutant of CK2alpha that has Asp 156 mutated to Ala (CK2alphaA156) is able to bind the CK2beta subunit and to compete effectively in this binding with wild-type subunits alpha and alpha'. The interaction between CK2alphaA156 and CK2beta was also demonstrated by transfection of epitope-tagged cDNA constructs into COS-7 cells. Immunoprecipitation of epitope-tagged CK2alphaA156 coprecipitated the beta subunit and vice-versa. The assay of the CK2 activity of the extracts obtained from cells transiently transfected with these different subunits yielded some surprising results: The CK2 specific phosphorylating activity of these cells transfected with the inactive CK2alphaA156 was considerably higher than the control cells transfected with vectors alone. Assays of the immunoprecipitated CK2alphaA156 expressed in these cells, however, demonstrated that the mutant was indeed inactive. It can be concluded that transfection of the inactive CK2alphaA156 affects the endogenous activity of CK2. Transfection experiments with CK2alpha and beta subunits and CK2alphaA156 were also used to confirm the interaction of CK2 with the general CDK inhibitor p21WAF1/CIP1 co-transfected into these cells. Finally a search in the SwissProt databank for proteins with properties similar to those derived from the amino acid composition of CK2beta indicated that CK2beta is related to protein phosphatase 2A and to other phosphatases as well as to a subunit of some ion-transport ATPases.