Characterization of adenosinetriphosphatase of squid actomyosin.

Abstract

Adenosinetriphosphatase (ATPase) activity of actomyosin of the obliquely striated mantle muscle of squid was studied to confirm and supplement the previous work. ATPase activity was assayed at ionic strength of 0.15 or 0.6, Tris-maleate buffer 30 mM (pH 7.0-7.5), protein 0.1mg/ml, ATP 3.2 mM, CaCl2, MgCl2 or EDTA 3.2 mM, and 25°C; for 10 min. Effects of Ca2+ and Mg2+ were tested in the function of temperature. Both Ca2+ and Mg2+ revealed marked activation at eithe ionic strength of 0.15 or 0.6, but EDTA none. At ionic strength 0.15, Ca2+-ATPase showed a steep peak at 30°C, while Mg2+-ATPase demonstrated a gradual peak with the top at 35°C. At ionic strenght of 0.6, the macimum of the former appeared at 25°C and that of the latter at 40°C. Mg2+ ion seemed to stabilize the ATPase. When the activety was assayed at various ratios of actomyosin and ATP, the full activity levels were attained below the ratio of 0.1-0.2 mg/ml protein per 3 mM ATP. Addition of Mg2+ to Ca2+-activated ATPase, reduced the activity to the level activated with Mg2+ alone, while addition of Ca2+ to Mg2+-activated ATPase, did not alter the activity. Effects of Mg2+ appeared to overcome that of Ca2+. During 60 min incubation of enzymic reaction mixture at 25°C prior to adding ATP, the activity was reduced appreciably. This seemed to satand with the so far reported lability of the squid actomyosin which was illustrated in changes of several physiconchemical properties.