Characterization of the catalytic subunit of casein kinase II expressed in escherichia coli and regulation of activity

The catalytic (α) subunit of casein kinase II from Drosophila, cloned and expressed in Escherichia coli (Saxena, A., Padmanabha, R., and Glover, C. V. C., (1987) Mol. Cell. Biol. 7, 3409-3417), has been purified and characterized, and the properties have been compared to those of the holoenzyme. The catalytic subunit exhibits protein kinase activity with casein as substrate and is autophosphorylated. The specific activity of the purified subunit is 6% of the activity of the holoenzyme from reticulocytes or from Drosophila. The α subunit is a monomer, eluting at M_r = 40,000 upon gel filtration in high salt, but as part of an aggregate in low salt. The α subunit has been purified to apparent homogeneity by sequential chromatography on DEAE-cellulose, Mono S, and Mono Q. A single band, M_r = 37,000, is detected by silver staining following polyacrylamide gel electrophoresis. The isolated α subunit displays apparent K_m values for β casein, ATP, and GTP similar to those of the holoenzyme. The activity of the α subunit is inhibited by heparin with an I₅₀ of 0.1-0.3 μg/ml, a value similar to that observed for the holoenzyme; autophosphorylation is also inhibited by heparin. Polylysine has no stimulatory effect on the activity of the catalytic subunit, as measured with casein and by autophosphorylation, but stimulates both activities with the holoenzyme. When physiological substrates for casein kinase II are examined, glycogen synthase and eukaryotic initiation factor 3 (elF-3) (p120) are phosphorylated by the α subunit at a rate equivalent to that of the holoenzyme, while phosphorylation of elF-3 (p67) is reduced 9-fold and elF-2β is not modified. From these data, it can be concluded that the α subunit of casein kinase II is sufficient for catalysis, is autophosphorylated, and can be directly inhibited by heparin, whereas the β subunit mediates the effects of basic stimulatory compounds and is involved in recognition and/or binding to specific physiological substrates.