Identification of Mutations in Protein Kinase CKIIβ Subunit That Affect Its Binding to Ribosomal Protein L41 and Homodimerization

  • Ahn, Bong-Hyun (Institutions Department of Biochemistry, College of Natural Sciences, Kyungpook National University) ;
  • Lee, Ji-Hoon (Institutions Department of Biochemistry, College of Natural Sciences, Kyungpook National University) ;
  • Bae, Young-Seuk (Institutions Department of Biochemistry, College of Natural Sciences, Kyungpook National University)
  • Received : 2003.01.30
  • Accepted : 2003.02.14
  • Published : 2003.07.31


Protein kinase CKII is composed of two catalytic ($\alpha$ or $\alpha$') subunits and two regulatory ($\beta$) subunits. The $CKII{\beta}$ subunit is thought to mediate the tetramer formation and interact with other target proteins. However, its physiological function remains obscure. In this study, point mutants of $CKII{\beta}$ that are defective for the L41 binding were isolated by using the reverse two-hybrid system. A sequence analysis of the point mutants revealed that Asp-26, Met-52, and Met-78 of $CKII{\beta}$ are critical for L41 binding; Asn-67 (and/or Lys-139) and Met-52 are important for $CKII{\beta}$ homodimerization. Two point mutants, R75 and R83, of $CKII{\beta}$ interacted with L5, topoisomerase $II{\beta}$, and CKBBP1/SAG, but not with the wild-type $CKII{\beta}$. This indicates that $CKII{\beta}$ homodimerization is not a prerequisite for its binding to target proteins. These $CKII{\beta}$ point mutants may be useful in exploring the biochemical physiological functions of $CKII{\beta}$.


Protein kinase CKII;Protein-protein interaction;Random PCR mutagenesis;Reverse two-hybrid system


Supported by : Korea Science & Engineering Foundation


  1. Aim, B. -H., Kim, T. -H. and Bae, Y. -S. (2001) Mapping of the interaction domain of the protein kinase CKII $\beta$ subunit with target proteins. Mol. Cells 12, 158-163.
  2. Appel, K, Wagner, P., Boldyreff, B., Issinger, O. G. and Montenarh, M. (1995) Mapping of the interaction sites of the growth suppressor protein p53 with the regulatory $\beta$ subunit of protein kinase CK2. Oncogene 11, 1971-1978.
  3. Boldyreff, B., Meggio, F, Pinna, L. A. and Issinger, O. G. (1994) Efficient autophosphorylation and phosphorylation of the $\beta$ subunit by casein kinase-2 require the integrity of an acidic cluster 50 residues downstream from the phosphoacceptor site. J. Biol. Chem. 269, 4827-4832.
  4. Boldyreff, B. and Issinger, O. G. (1997) A-Raf kinase is a new interacting partner of protein kinase CK2 $\beta$ subunit. FEBS Lett. 403, 197-199.
  5. Chen, M., Li, D., Krebs, E. G. and Cooper, J. A. (1997) The casein kinase II $\beta$ subunit binds to Mos and inhibits Mos activity. Mol. Cell. Biol. 17, 1904-1912.
  6. Feilotter, H. E., Hannon, G. J., Ruddell, C. J. and Beach, D. (1994) Construction of an improved host strain for two hybrid screening. Nucleic Acids Res. 22, 1502-1503.
  7. Fields, S. and Song, O. (1989) A novel genetic system to detect protein-protein interactions. Nature 340, 245-246.
  8. Gietz, R. D., Graham, K. C. and Litchfield, D. W. (1995) Interactions between the subunits of casein kinase II. J. Biol. Chem. 270, 13017-13021.
  9. Hagemann, C, Kalmes, A., Wixler, V., Wixler L., Schuster, T. and Rapp, U. R. (1997) The regulatory subunit of protein kinase CK2 is a specific A-Raf activator. FEBS Lett. 403, 200-202.
  10. Hanna, D. E., Rethinaswamy, A. and Glover, C. V. C. (1995) Casein kinase II is required for cell cycle progression during G1 and G2/M in Saccharomyces cerevisiae. J. Biol. Chem. 270, 25905-25914.
  11. Issinger, O. G. (1993) Casein kinases: pleiotropic mediators of cellular regulation. Pharmacol. & Ther. 59, 1-30.
  12. Jakobi, R. and Traugh, J. A. (1992) Characterization of the phosphotransferase domain of casein kinase II by site-directed mutagenesis and expression in Escherichia coli. J. Biol. Chem. 267, 23894-23902.
  13. Kim, I. -M., Cha, I. -Y, Marshak, D. R. and Bae, Y -S. (1996) Interaction of the $\beta$ subunit of casein kinase II with the ribosomal protein L5. Biochem. Biophys. Res. Commun. 226, 180-186.
  14. Kim, M. -S., Lee, Y. -T., Kim, J. -M., Cha, J. -Y. and Bae, Y. -S. (1998) Characterization of protein interaction among subunits of protein kinase CKII in vivo and in vitro. Mol. Cells 8, 43-48.
  15. Kim, Y. -S., Ha, K. -S., Kim, Y. -H. and Bae, Y. -S. (2002) The Ring-H2 finger motif of CKBBP1/SAG is necessary for interaction with protein kinase CKII and optimal cell proliferation. J. Biochem. Mol. Biol. 35, 629-636.
  16. Lee, I. -H., Kim, I. -M., Kim, M. -S., Lee, Y -T., Marshak, D. R. and Bae, Y -S. (1997) The highly basic ribosomal protein L41 interacts with the $\beta$ subunit of protein kinase CKII and stimulates phosphorylation of DNA topoisomerase Ifx by CKII. Biochem. Biophys. Res. Commun. 238, 462-467.
  17. Leung, D. W., Chen, E. and Goeddel, D. V. (1989) A method for random mutagenesis of a defined DNA segment using a modified polymerase chain reaction. Technique 1, 11-15.
  18. Lin, W. -J., Tuazon, P. T. and Traugh, J. A. (1991) Characterization of the catalytic subunit of casein kinase II expressed in Escherichia coli and regulation of activity J. Biol. Chem. 266, 5664-5669.
  19. Litchfield, D. W. (2003) Protein kinase CK2: structure, regulation and role in cellular decisions of life and death. Biochem. J. 369, 1-15.
  20. Meggio, F., Boldyreff, B., Marin, O., Pinna, L. A. and Issinger, O. -G. (1992) Role of the $\beta$ subunit of casein kinase-2 on the stability and specificity of the recombinant reconstituted holoenzyme. Eur. J. Biochem. 204, 293-297.
  21. Meggio, R, Boldyreff, B., Issinger, O. G. and Pinna, L. A. (1994) Casein kinase 2 down-regulation and activation by polybasic peptides are mediated by acidic residues in the 55-64 region of the $\beta$ subunit. A study with calmodulin as phosphorylatable substrate. Biochemistry 33, 4336-4342.
  22. Padmanabha, R., Chen-Wu, J. L., Hanna, D. E. and Glover, C. V.(1990) Isolation, sequencing, and disruption of the yeast CKA2 gene: casein kinase II is essential for viability in Saccharomyces cerevisiae. Mol. Cell. Biol. 10, 4089-4099.
  23. Park, K. -H., Lee, Y. -T. and Bae, Y. -S. (2001) Stimulation of human DNA topoisomerase II activity by its direct association with the $\beta$ subunit of protein kinase CKII. Mol. Cells 11, 82-88.
  24. Park, G. -H. and Bae, Y -S. (2001) Mapping of the interaction domain of DNA topoisomerase $Il{\alpha}$ and $III{\beta}$ with extracellular signal-regulated kinase 2. J. Biochem. Mol. Biol. 34, 85-89.
  25. Pinna, L. A. (1990) Casein kinase 2: an ‘eminence grise’ in cellular regulation? Biochem. Biophys. Acta 1054, 267-284.
  26. Pinna, L. A. (1997) Protein kinase CK2. Int. J. Biochem. Cell Biol. 29, 551-554.
  27. Raman, C., Kuo, A., Deshane, J., Litchfield, D. W. and Kimberly,R. P. (1998) Regulation of casein kinase 2 by direct interaction with cell surface receptor CD5. J. Biol. Chem. 273, 19183-19189.
  28. Rose, M. D., Winston, F. and Hieter, P. (1990) Laboratory Courses Manual for Methods in Yeast Genetics, Cold Spring Harbor Laboratory Press, New York, USA.
  29. Seldin, D. C. and Leder, P. (1995) Casein kinase Ila transgene-induced murine lymphoma: relation to theileriosis in cattle. Science 267, 894-897.
  30. Son, M. -Y., Park, J. -W., Kim, Y. -S., Kang, S. -W., Marshak, D.R., Park, W. and Bae, Y. -S. (1999) Protein kinase CKII interacts with and phosphorylates the SAG protein containing ring-H2 finger motif. Biochem. Biophys. Res. Commun. 263, 743-748.
  31. Vidal, M., Brachmann, R. K., Fattaey, A., Harlow, E. and Boeke,J. D. (1996) Reverse two-hybrid and one-hybrid systems to detect dissociation of protein-protein and DNA-protein interactions. Proc. Natl. Acad. Sci. USA 93, 13896-13901.

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