Rotational and translational motion of troponin C

Time resolved fluorescence anisotropy and sedimentation velocity has been u sed to study the rotational and translational hydrodynamic behavior of two mutants of chicken skeletal troponin C bearing a single tryptophan residue at position 78 or 154 in the metal-free-, metal-bound-, and troponin I pept ide (residues 96-116 of troponin I)-ligated states. The fluorescence anisot ropy data of both mutants were adequately described by two rotational corre lation times, and these are compared with the theoretically expected values based on the rotational diffusion of an idealized dumbbell. These data imp ly that the motion of the N- and C-terminal domains of troponin C are indep endent. They also suggest that in the metal-free, calcium-saturated and cal cium-saturated troponin I peptide-bound states, troponin C is elongated, ha ving an axial ratio of 4-5, Calcium or magnesium binding to the high affini ty sites alone reduces the axial ratio to approximately 3. However, with ca lcium bound to sites III and IV and in the presence of a 1:1 molar ratio of the troponin I peptide, troponin C is approximately spherical. The metal i on and troponin I peptide-induced length changes in troponin C may play a r ole in the mechanism by which the regulatory function of troponin C is effe cted.