A protein activator of Ca(2+)-calmodulin-dependent protein kinase Ia.
A protein activator of Ca(2+)-calmodulin-dependent protein kinase Ia (CaM kinase Ia) was purified to near homogeneity from pig brain. In the final step of purification, sucrose density gradient centrifugation, CaM kinase Ia activating activity correlated with the presence of a approximately 52-kDa protein band detected by SDS-polyacrylamide gel electrophoresis. Comparison of this value with estimations of its molecular mass under nondenaturing conditions indicated that CaM kinase Ia activator is a slightly asymmetric monomer. After removal of endogenous CaM kinase Ia activator, the activity of CaM kinase Ia was 2% of its activity in the presence of a maximally stimulating concentration (15 nM) of the purified activator. In its activated state, CaM kinase Ia retained complete dependence of its activity upon Ca(2+)-CaM. The activation of CaM kinase Ia was rapid (t1/2 < 1 min) and required the combined presence of CaM kinase Ia activator, Ca(2+)-CaM, and MgATP. Similarly, in addition to MgATP, the phosphorylation of CaM kinase Ia required CaM kinase Ia activator and Ca(2+)-CaM. CaM kinase Ia activator was capable of Ca(2+)-dependent binding to CaM-Sepharose. The requirement of the combined presence of CaM kinase Ia activator, Ca(2+)-CaM, and MgATP for both the activation and phosphorylation of CaM kinase Ia is discussed in terms of potential mechanisms for CaM kinase Ia activation.
Lee
J C
JC
Department of Pharmacology and Therapeutics, State University of New York at Buffalo, New York 14214.
Edelman
A M
AM
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002499:Centrifugation, Density Gradient; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000138:chemical synthesis; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013552:Swine
D007527:Isoenzymes; D010455:Peptides; D011506:Proteins; D011494:Protein Kinases; C106686:(CaM)-dependent protein kinase Ia kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases