Preparation and characterization of calmodulin-dependent protein kinase IV (CaM-kinase IV) free of CaM-kinase IV kinase from rat cerebral cortex.
Our earlier finding [Kameshita, I. and Fujisawa, H. (1993) J. Biochem. 113, 583-590] that calmodulin-dependent protein kinase IV (CaM-kinase IV) from rat cerebral cortex is markedly activated through autophosphorylation was reexamined in the light of a more recent study [Okuno, S. and Fujisawa, H. (1993) J. Biochem. 114, 167-170] suggesting the involvement of Ca2+/calmodulin-dependent CaM-kinase IV kinase in the activation of CaM-kinase IV in rat brain. Further purification of the previous CaM-kinase IV preparation by HPLC on a DEAE-NPR column abolished its marked activation on incubation under Ca2+/calmodulin-dependent phosphorylation conditions, and the marked activation was restored by the addition of the fraction separated on a DEAE-NPR column, suggesting that CaM-kinase IV was separated from CaM-kinase IV kinase involved in its activation by HPLC on DEAE-NPR. The HPLC-purified CaM-kinase IV still underwent a slow autophosphorylation, which was an intramolecular reaction. Some kinetic properties of CaM-kinase IV before and after autophosphorylation were examined for comparison. In addition, the purification by HPLC on DEAE-NPR revealed that a novel Ca(2+)-dependent calmodulin-binding protein, designated as CaMBP64, is present as a major contaminant in the previous purified CaM-kinase IV preparation.
Kameshita
I
I
Department of Biochemistry, Asahikawa Medical College, Hokkaido.
Fujisawa
H
H
D000595:Amino Acid Sequence; D000818:Animals; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000302:isolation & purification*; D002384:Catalysis; D002540:Cerebral Cortex / Q000201:enzymology*; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004789:Enzyme Activation; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D051381:Rats
D017346:Protein Serine-Threonine Kinases; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054734:Calcium-Calmodulin-Dependent Protein Kinase Type 4; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517686:Camk4 protein, rat