Comparison of effects of smooth and skeletal muscle tropomysins on interactions of actin and myosin subfragment 1

Abstract

The ATPase activity of [rabbit] acto.cntdot.myosin subfragment 1 (S-1) was measured in the presence of smooth and skeletal muscle tropomyosins over a wide range of ionic strengths (20-120 mM). In contrast to the 60% inhibitory effect caused by skeletal muscle tropomyosin at all ionic strengths, the effect of smooth muscle tropomyosin was dependent on ionic strength. At low ionic strength (20 mM), smooth muscle tropomyosin inhibits the ATPase activity by 60%, while at high ionic strength (120 mM), it potentiates the ATPase activity 3-fold. All of these ATPase activities were measured at very low ratios of S-1 to actin, under conditions at which a 4-fold increase in S-1 concentration did not change the specific activity of the tropomyosin-acto.cntdot.S-1 ATPase. Therefore, the potentiation of the ATPase activity by smooth muscle tropomyosin at high ionic strength cannot be explained by bound S-1 heads cooperatively turning on the tropomyosin-actin complex. To determine whether the fully potentiated rates are different in the presence of smooth muscle and skeletal muscle tropomyosins, S-1 which was extensively modified by N-ethylmaleimide was added to the ATPase assay to attain high ratios of S-1 to actin. Under all conditions, the fully potentiated rates are the same for both tropomyosins. Therefore, the difference in the effect of smooth muscle and skeletal muscle tropomyosin on the acto.cntdot.S-1 ATPase activity at low ratios of S-1 to actin appears to be due to a greater fraction of the tropomyosin-actin complex being turned on in the absence of S-1 with smooth muscle tropomyosin than with skeletal muscle tropomyosin. This interpretation was supported by the equilibrium binding studies of S-1 to actin at 120 mM ionic strength in the presence of the ATP analog adenyl-5''-yl imidodiphosphate. In the absence of S-1, a greater fraction of the tropomyosin-actin complex is probably in the strong S-1-binding form with smooth muscle tropomyosin than with skeletal muscle tropomyosin. The fraction of tropomyosin-actin complex in the strong S-1-binding form may correlate with the effect of tropomyosin on the actin-activated ATPase rate.