title: Functional diversity of alternatively spliced isoforms of Drosophila Ca2+/calmodulin-dependent protein kinase II. A role for the variable domain in activation.
abstract: Isoforms of calcium/calmodulin-dependent protein kinase II from Drosophila (R1-R6 and R3A) showed differential activation by two series of mutant calmodulins, B1K-B4K and B1Q-B4Q. These mutant calmodulins were generated by changing a glutamic acid in each of the four calcium binding sites to either glutamine or lysine, altering their calcium binding properties. All mutations produced activation defects, with the binding site 4 and B1Q mutants the most severe. Activation differed substantially between isoforms. R4, R5, and R6 were the least sensitive to mutations in calmodulin, while R1, R3, and R3A were the most sensitive. Activation of R1 and R2 by B4K and activation of R3 and R3A by B2K and B2Q produced significant (6-fold and almost 3-fold, respectively) differences in Kact between isoforms that differ structurally by a single amino acid. These differences could not be accounted for by differential binding, as all isoforms showed almost identical binding patterns with the mutants. High binding affinity did not always correlate with ability to increase enzyme activity, implying that activation occurs in at least two steps. The isoform-specific differences seen in this study reflect a role for the COOH-terminal variable region in activation of CaM kinase II.
authors.affiliation: Department of Biology and Center for Complex Systems, Brandeis University, Waltham, Massachusetts 02254, USA.
authors.forename: B
authors.initials: B
authors.lastname: GuptaRoy
authors.forename: K
authors.initials: K
authors.lastname: Beckingham
authors.forename: L C
authors.initials: LC
authors.lastname: Griffith
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D002118:Calcium / Q000378:metabolism; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism; D004331:Drosophila melanogaster / Q000201:enzymology*; D004789:Enzyme Activation; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D013329:Structure-Activity Relationship
chemical_list: D002147:Calmodulin; D007527:Isoenzymes; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium