Selective modulation of calcium-dependent myosin phosphorylation by novel protein kinase inhibitors, isoquinolinesulfonamide derivatives.
Ca2+-dependent myosin phosphorylation by Ca2+/calmodulin-dependent myosin light chain kinase (MLC-kinase) and protein kinase C were studied using selective inhibitors, isoquinolinesulfonamide derivatives. Both protein kinases were potently inhibited by 1-(8-chloro-5-isoquinolinesulfonyl)piperazine (HA-156) and its derivatives. Kinetic analysis indicated that HA-156 inhibited both enzymes competitively with respect to ATP, and Ki values of HA-156 for MLC-kinase and protein kinase C were 7.3 and 7.2 microM, respectively. To clarify molecular mechanisms of the isoquinolinesulfonamides to inhibit the Ca2+-dependent protein kinases, we examined the structure-activity relationships of HA-156 and its derivatives. The dechlorinated analogues, HA-100 and HA-142, markedly decreased the affinity for MLC-kinase, suggesting that the inhibitory effect of isoquinolinesulfonamide derivatives depends upon hydrophobicity of the compounds. There is a good correlation between MLC-kinase inhibition and hydrophobicity determined by reverse phase chromatography. In contrast, HA-140 and HA-142 showed weak inhibition of protein kinase C, suggesting that the electron density of the nitrogen in the isoquinoline ring of the compounds correlates with the potency to inhibit protein kinase C activity. These pairs of isoquinolinesulfonamides will aid in elucidating the biological roles of Ca2+-dependent myosin phosphorylation in intact cells. HA-156 and HA-140 inhibited myosin light chain phosphorylation in platelets exposed to collagen, whereas HA-142 and HA-100 did not, significantly. These isoquinolinesulfonamide derivatives should prove to be useful tools for distinguishing between the biological functions of Ca2+-activated, phospholipid-dependent, and Ca2+/calmodulin-dependent myosin light chain phosphorylation, in vivo.
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors; D000818:Animals; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology*; D002645:Chickens; D005895:Gizzard, Avian; D007546:Isoquinolines / Q000494:pharmacology*; D007700:Kinetics; D009130:Muscle, Smooth / Q000201:enzymology; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D047428:Protein Kinase Inhibitors; D011817:Rabbits; D013329:Structure-Activity Relationship; D013449:Sulfonamides / Q000494:pharmacology*
D007546:Isoquinolines; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D011493:Protein Kinase C; D009219:Myosin-Light-Chain Kinase; D000251:Adenosine Triphosphatases; D002118:Calcium