Abstract
Changes in the orientation of tropomyosin on actin are important for the regulation of striated muscle contraction and could also be important for smooth muscle regulation. We showed earlier that acrylodan-labeled skeletal muscle tropomyosin reports the kinetics of the reversible transitions among the active, intermediate, and inactive states when S1 is rapidly detached from actin-tropomyosin. We now show that acrylodan-labeled smooth muscle tropomyosin reports similar transitions among states of actin-tropomyosin. When S1 was rapidly detached from actin-smooth muscle tropomyosin, there was a rapid decrease in acrylodan-tropomyosin fluorescence as the intermediate state became populated. The rate constant for this process was >600 s(-1) at temperatures near 5 °C. In the presence of skeletal troponin and EGTA, the decrease in fluorescence was followed by the redevelopment of fluorescence as the inactive state became populated. The apparent rate constant for the fluorescence increase was 14 s(-1) at 5 °C. Substituting smooth muscle caldesmon for skeletal muscle troponin produced a similar decrease and re-increase in fluorescence, but the apparent rate constant for the increase was >10 times that observed with troponin. Furthermore, the fluorescence increase was correlated with an increase in the extent of caldesmon attachment as S1-ATP dissociated. Although the measured rate constant appeared to reflect the rate-limiting transition for inactivation, it is unclear if the fluorescence change resulted from caldesmon binding, the movement of tropomyosin over actin, or both.