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  • Title: Purification and characterization of the CK2alpha'-based holoenzyme, an isozyme of CK2alpha: a comparative analysis.
    Author: Olsen BB, Boldyreff B, Niefind K, Issinger OG.
    Journal: Protein Expr Purif; 2006 Jun; 47(2):651-61. PubMed ID: 16442308.
    Abstract:
    Protein kinase CK2 (former name: "casein kinase 2") is a pivotal and ubiquitously expressed member of the eukaryotic protein kinase superfamily. It predominantly exists as a heterotetrameric holoenzyme composed of two catalytic subunits (CK2alpha) and two regulatory subunits (CK2beta). In higher animals two paralog catalytic chains-abbreviated CK2alpha and CK2alpha'--exist which can combine with CK2beta to three isoforms of the holoenzyme: CK2alpha(2)beta(2), CK2alpha(2)(')beta(2), and CK2alphaalpha(')beta(2). While CK2alpha and the "normal" holoenzyme CK2alpha(2)beta(2) have been extensively characterized in vitro and in vivo, little is known about the enzymological properties of CK2alpha' and the "alternative" holoenzyme CK2alpha(2)(')beta(2) and about their specific physiological roles. A major reason for this lack of knowledge is the fact that so far CK2alpha' rather than CK2alpha has caused serious stability and solubility problems during standard heterologous expression procedures. To overcome them, we developed a preparation scheme for CK2alpha(2)(')beta(2) from Homo sapiens in catalytically active form based on two critical steps: first expression of human CK2alpha' as a well soluble fusion protein with the maltose binding protein (MBP) and second proteolytic cleavage of CK2alpha'-MBP in the presence of human CK2beta so that CK2alpha' subunits are incorporated into holoenzyme complexes directly after their release from MBP. This successful strategy which may be adopted in comparably difficult cases of protein/protein complex preparation is presented here together with evidence that the CK2alpha'-based and the CK2alpha-based holoenzymes are similar concerning their catalytic activities but are significantly different with respect to some well-known CK2 properties like autophosphorylation and supra-molecular aggregation.
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