Structure and interactions of the carboxyl terminus of striated muscle α-tropomyosin:: It is important to be flexible

Tropomyosin (TM) binds to and regulates the actin filament. We used circular dichroism and heteronuclear NMR to investigate the secondary structure and interactions of the C terminus of striated muscle alpha-TM, a major functional determinant, using a model peptide, TM9a(251-284). The H-1(alpha) and C-13(alpha) chemical shift displacements show that residues 252 to 277 are alpha-helical but residues 278 to 284 are nonhelical and mobile. The H-1(N) and C-13' displacements suggest that residues 257 to 269 form a coiled coil. Formation of an "overlap" binary complex with a 33-residue N-terminal chimeric peptide containing residues 1 to 14 of alpha-TM perturbs the H-1(N) and N-15 resonances of residues 274 to 284. Addition of a fragment of troponin T, TnT(70-170), to the binary complex perturbs most of the H-1(N)-N-15 cross-peaks. In addition, there are many new cross-peaks, showing that the binding is asymmetric. Q263, in a proposed troponin T binding site, shows two sets of side-chain N-15-H-1 cross-peaks, indicating conformational flexibility. The conformational equilibrium of the side chain changes upon formation of the binary and ternary complexes. Replacing Q263 with leucine greatly increases the stability of TlM9a(251-284) and reduces its ability to form the binary and ternary complexes, showing that conformational flexibility is crucial for the binding functions of the C terminus.