Abstract
In order to elucidate the physiological role of calcium binding protein, calmodulin in platelet aggregation phenomenon, we determined the effect of the calmodulin-interacting agent, which selectively binds to calmodulin and inhibits Ca2+-calmodulin activated enzymes, on the endogenous phosphorylation of myosin light chain from human platelets. An actomyosin enrich fraction contained approximately 50 units of calmodulin per mg protein. N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W-7) as a calmodulin-interacting agent produced dose-dependent inhibition of 32P incorporation into 20, 000 dalton myosin light chain from [γ-32P] ATP in the presence of calcium ions (100μM) with the IC50 value of 75μM. However within the concentration range examined, W-7 was not effective to inhibit the phosphorylation of MLC in the absence of calcium (EGTA 5mM).
Moreover, W-7 was able to inhibit in dose-dependent fashion platelet aggregation induced by various aggregating agents such as adenosine diphosphate (ADP), collagen, epinephrine or restocetin with indistinguishable IC50 values to that observed in phosphorylation inhibition. Our results provide pharmocological evidence that calmodulin-mediated system, especially Ca2+-dependent phosphorylation of platelet myosin light chain play an important role and is common process in platelet aggregation phenomenon induced by any aggregating agents.
