Rotational and translational motion of troponin C.
Time resolved fluorescence anisotropy and sedimentation velocity has been used 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 peptide (residues 96-116 of troponin I)-ligated states. The fluorescence anisotropy data of both mutants were adequately described by two rotational correlation times, and these are compared with the theoretically expected values based on the rotational diffusion of an idealized dumbbell. These data imply that the motion of the N- and C-terminal domains of troponin C are independent. They also suggest that in the metal-free, calcium-saturated and calcium-saturated troponin I peptide-bound states, troponin C is elongated, having an axial ratio of 4-5. Calcium or magnesium binding to the high affinity sites alone reduces the axial ratio to approximately 3. However, with calcium 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 ion and troponin I peptide-induced length changes in troponin C may play a role in the mechanism by which the regulatory function of troponin C is effected.
Moncrieffe
M C
MC
Department of Biochemistry and Molecular Biology, Mayo Foundation, Rochester, Minnesota 55905, USA.
Eaton
S
S
Bajzer
Z
Z
Haydock
C
C
Potter
J D
JD
Laue
T M
TM
Prendergast
F G
FG
D000818:Animals; D002118:Calcium / Q000737:chemistry; D002645:Chickens; D005454:Fluorescence Polarization; D008274:Magnesium / Q000737:chemistry; D018482:Muscle, Skeletal / Q000737:chemistry; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D019209:Troponin C / Q000737:chemistry*; D019210:Troponin I / Q000737:chemistry; D014364:Tryptophan / Q000737:chemistry; D014461:Ultracentrifugation
D010446:Peptide Fragments; D019209:Troponin C; D019210:Troponin I; D014364:Tryptophan; D008274:Magnesium; D002118:Calcium