SOLUTION SECONDARY STRUCTURE OF CALCIUM-SATURATED TROPONIN-C MONOMER DETERMINED BY MULTIDIMENSIONAL HETERONUCLEAR NMR-SPECTROSCOPY

The solution secondary structure of calcium-saturated skeletal troponi n C (TnC) in the presence of 15% (v/v) trifluoroethanol (TFE), which h as been shown to exist predominantly as a monomer (Slupsky CM, Kay CM, Reinach FC, Smillie LB, Sykes ED, 1995, Biochemistry 34, forthcoming) , has been investigated using multidimensional heteronuclear nuclear m agnetic resonance spectroscopy. The H-1, N-15, and C-13 NMR chemical s hift values for TnC in the presence of TFE are very similar to values obtained for calcium-saturated NTnC (residues 1-90 of skeletal TnC), c almodulin, and synthetic peptide homodimers. Moreover, the secondary s tructure elements of TnC are virtually identical to those obtained for calcium-saturated NTnC, calmodulin, and the synthetic peptide homodim ers, suggesting that 15% (v/v) TFE minimally perturbs the secondary an d tertiary structure of this stably folded protein. Comparison of the solution structure of calcium-saturated TnC with the X-ray crystal Str ucture of half-saturated TnC reveals differences in the phi/psi angles of residue Glu 41 and in the linker between the two domains. Glu 41 h as irregular phi/psi angles in the crystal structure, producing a kink in the B helix, whereas in calcium-saturated TnC, Glu 41 has helical phi/psi angles, resulting in a straight B helix. The linker between th e N and C domains of calcium-saturated TnC is flexible in the solution structure.