Phospholipid-sensitive Ca2+-dependent protein kinase inhibition by R-24571, a calmodulin antagonist.
R-24571 (calmidazolium), a derivative of the antimycotic agent miconazole, inhibited phospholipid-sensitive Ca2+-dependent protein kinase (PL-Ca-PK), with an IC50 (the concentration causing 50% inhibition) of 5.3 microM. It also inhibited the calmodulin/Ca2+-stimulated enzymes, with IC50 values of 1.6 and 0.1 microM for myosin light chain kinase (MLCK) and phosphodiesterase respectively. Analysis of inhibition by R-24571 of PL-Ca-PK and MLCK revealed complex kinetics, suggesting that the agent interacted with the cofactors, the enzyme, and/or the cofactor-enzyme complexes. At saturating concentrations of the cofactors, R-24571 inhibited PL-Ca-PK and MLCK noncompetitively with their respective cofactors. Inhibition of MLCK by R-24571 was completely overcome by phosphatidylserine, indicating a strong hydrophobic interaction between R-24571 and the phospholipid in the presence of calmodulin. R-24571 also inhibited phosphorylation of various endogenous proteins in brain stimulated specifically by phosphatidylserine/Ca2+ or calmodulin/Ca2+. The present findings inducated that R-24571 has little specificity in inhibiting two types of Ca2+-dependent protein kinases sensitive to phospholipid or calmodulin.
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000037:antagonists & inhibitors*; D002417:Cattle; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D007700:Kinetics; D007948:Leukemia, Monocytic, Acute / Q000201:enzymology; D009219:Myosin-Light-Chain Kinase; D010743:Phospholipids / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D013552:Swine
D002147:Calmodulin; D007093:Imidazoles; D010743:Phospholipids; D047428:Protein Kinase Inhibitors; C031938:calmidazolium; D009219:Myosin-Light-Chain Kinase; D002118:Calcium