Purification and characterization of Ca2+/calmodulin-dependent protein kinase kinase beta from rat cerebellum.
The existence of isoforms of calmodulin-dependent protein kinase kinase (CaM-kinase kinase) in the rat brain was recently suggested by Northern and Western blot analyses and immunotitration [Okuno, S., Kitani, T., and Fujisawa, H. (1996) J. Biochem. 119, 1176-1181]. In the present study, CaM-kinase kinase beta, distinct from Cam-kinase kinase alpha which had been purified and cloned from rat cerebral cortex, was purified approximately 5,000-fold from rat cerebellum and its properties were examined. The purified CaM-kinase kinase beta gave a doublet at positions corresponding to molecular weights of 66,000 to 67,000 on SDS-PAGE, and neither protein band reacted with antibody against CaM-kinase kinase alpha. Both CaM-kinase kinase alpha and beta markedly activated both CaM-kinase I and IV, but CaM-kinase kinase beta activated CaM-kinase IV more strongly than did CaM-kinase kinase alpha. The maximal extents of the activation of CaM-kinase I and IV by CaM-kinase kinase beta were almost the same as those by CaM-kinase kinase alpha, suggesting that the two CaM-kinase kinases activated CaM-kinase I and IV by the same mechanisms.
Okuno
S
S
Department of Biochemistry, Asahikawa Medical College, Hokkaido.
Kitani
T
T
Fujisawa
H
H
D000595:Amino Acid Sequence; D000818:Animals; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002531:Cerebellum / Q000737:chemistry / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000302:isolation & purification* / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity
D017346:Protein Serine-Threonine Kinases; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517701:Camkk1 protein, rat; C517702:Camkk2 protein, rat