Tropomyosin rotational dynamics in thin filaments

摘要

We have used phosphorescence anisotropy from erythrosin-5-iodoacetamide covalently attached to rabbit skeletal muscle tropomyosin (Tm) to characterize the molecular dynamics of Tm on the surface of F-actin. Although the probe was loosely bound to Tm, it was able to monitor the molecular dynamics of the F-actin/Tm complex on the microsecond time scale. The steady-state phosphorescence anisotropy of Tm in the complex was 0.025±0.005 at 20°C. Since the anisotropy of actin in a similar complex was 0.11±0.01, Tm may be moving independently on the filament surface. Studies of the effect of temperature and solution viscosity provided additional evidence for independent tropomyosin motions. A modified Perrin plot of 1/r versus rT/η (where r is the steady-state phosphorescence anisotropy, τ is the phosphorescence lifetime, T is the absolute temperature, and η is solution viscosity) provided evidence for two distinct modes of motion for Tm on F-actin; these modes corresponded to the rotation of segments with significantly different volumes. While the volume of the fast component was comparable to that of a single tropomyosin molecule, providing evidence that Tm moves on the surface of F-actin, the volume of the slow component corresponded to only a small portion of the entire complex.