Modulation of actomyosin ATPase by thin filament-associated proteins

Abstract

Phosphorylation of the myosin light chain is a prerequisite for actin-activation of the Mg-ATPase of smooth muscle myosin. However, maximal activation of the Mg-ATPase by actin requires stoichiometric binding of tropomyosin to actin filaments and Ca2+ at free Mg2+ below 3 mM. The requirement for Ca2+ for actin-activation is not due to a calcium-mediated binding of tropomyosin to actin since the binding of tropomyosin to actin is not dependent on Ca2+. Caldesmon, an actin and calmodulin binding protein, at caldesmon:actin molar ratio of 1:18, binds equally to pure actin and actin containing stoichiometric amounts of bound tropomyosin. The Mg-ATPase of myosin reconstituted with actin is not affected by the caldesmon; on the other hand, the activity of actomyosin containing tropomyosin is inhibited. The inhibition of activity by the caldesmon is reversed by the addition of calmodulin (caldesmon:calmodulin molar ratio, 1:8) in the presence of Ca2+. The amount of caldesmon bound to actin in the presence of calcium-calmodulin is 50% more when actin filaments contain tropomyosin, indicating that the release of inhibition of the activity inhibited by caldesmon does not require complete release of caldesmon from actin.