Chymotryptic Subfragments of Troponin T from Rabbit Skeletal Muscle. Interaction with Tropomyosin, Troponin I and Troponin C

Abstract

The binding of the chymotryptic troponin T subfragments to tropomyosin, troponin I, and troponin C was semiquantitatively examined by using affinity chromatography, and also by co-sedimentation with F-actin and polyacrylamide gel electrophoresis in 14mM Tris/90mM glycine. Circular dichroism spectra of the subfragments were measured to confirm that the subfragments retained their conformational structures. Based on these results, the binding sites of tropomyosin, troponin I, and troponin C on the troponin T sequence were elucidated. Tropomyosin bound mainly to the region of troponin T₁ (residues 1-158) with the same binding strength as to the original troponin T. The C-terminal region of troponin T (residues 243-259) was the second binding site to tropomyosin under physiological conditions. The binding site of troponin I was concluded to be the region including residues 223-227. The binding of troponin C was dependent on Ca²⁺ ion concentration. The C-terminal region of troponin T₂ (residues 159-259) was indicated to be the Ca²⁺-independent troponin C-binding site and the N-terminal side of troponin T₂ to be the Ca²⁺-dependent site.