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Abstract

The binding between F-actin and myosin has been studied by analyzing the amount of radio-actively labelled (by means of 14C-NEM) F-actin and myosin in the formed unsoluble actomyosin pellet after centrifugation of the reaction mixture. In the absence of ATP, the amount of actin bound to myosin varies, depending on the amounts of actin and myosin present, between 0.18 mg actin/mg myosin (2 actin units per 1 myosin molecule) and about 2 mg actin/mg myosin (each actin fibril only uncompletely saturated with myosin). In 0.03 ᴍ KCl ATP decreases, if at all, the affinity of actin towards myosin only slightly; but less actin is bound to myosin in the presence of MgATP and in low ionic strength, indicating that myosin is now more densily distributed over fewer actin fibrils leaving the rest of fibrils free. For precipitation of the total amount of myosin (myosin alone is soluble in MgATP) incomplete saturation of myosin with actin suffices; obviously actin promotes filament formation of myosin. The activation of myosin ATPase by actin depends in a similar manner as actin binding does on actin concentration, hence the enzymatic interaction between actin and myosin is accompanied by true actin-myosin binding. An actin-tropomyosin-troponin preparation, whose reduced viscosity is lower than that of F-actin and which consists of about 30% actin, activates myosin ATPase to the same extent as F-actin does. It competes with F-actin for the same binding sites on myosin and can in the presence of MgATP be displaced from myosin by those preparations of F-actin which have a strong tendency to become fully saturated with myosin. The activation of myosin ATPase by actin-tropomyosin-troponin is reduced after tryptic digestion of actin-tropomyosin-troponin, which affects, according to SDS gel electro­ phoresis, mainly two components of troponin with molecular weights of about 32 000 and 25 000, respectively.