Kinase experiment:

Ca2+/CaM-dependent phosphodiesterase is assayed at 30℃. The reaction mixture contains 40 mM Tris-HCl, pH 8.0, 5 mM magnesium acetate, 1 mM calcium chloride, 30 μM cGMP, 0.15 pCi of [3H]cGMP, 1 mM dithiothreitol, 20% glycerol, 0.64 mg/mL bovine serum albumin, 2.38 nM CaM, 50 pM CaM-deficient phosphodiesterase, and various concentrations of Calmodulin-Dependent Protein Kinase II (290-309). Following preincubation of the reaction mixture at 30℃, the reaction is initiated by the addition of substrate. After 50 min the reaction is terminated by boiling. Conditions are selected in which the reactions are linear with respect to time[1].

Calmodulin-Dependent Protein Kinase II (290-309) is a potent CaMK antagonist with an IC50 of 52 nM for inhibition of Ca2+/calmodulin-dependent protein kinase II.

Peptide 290 to 309 is found to be a potent calmodulin antagonist with an IC50 of 52 nM for inhibition of Ca2+/calmodulin-dependent protein kinase. Neither truncation from the amino terminus (peptide 296-309) nor extension in the carboxyl-terminal direction (peptide 294-319) markedly affects calmodulin binding, whereas shortening the peptide from the carboxyl terminus (peptide 290-302) or from both ends (peptide 295-304) results in the elimination of this activity[1].

[1]. Payne ME, et al. Calcium/calmodulin-dependent protein kinase II. Characterization of distinct calmodulin binding and inhibitory domains. J Biol Chem. 1988 May 25;263(15):7190-5.