Structure and Stability of Wildtype and F29W Mutant Forms of the N-Domain of Avian Troponin C Subjected to High Pressures

The N-domain of troponin C (residues 1–90) regulates muscle contraction through conformational changes induced by Ca2+ binding. A mutant form of this domain of avian troponin C (F29W) has been used in previous studies to observe conformational changes that occur upon Ca2+ binding, and pressure and temperature changes. In this study we examined the effect of the point mutation on the protein structure and its stability to pressure. We performed 1-D and 2-D 1H-NMR experiments at 300, 400, and 500 MHz on the wildtype and F29W mutant forms of the N-domain of chicken troponin C in the absence of Ca2+. We found that the mutant protein at 5 kbar pressures had a destabilized βl-sheet between the Ca2+-binding loops, an altered environment near Phe 26, and reduced local motions of Phe 26 and Phe 75 in the core of the protein, probably due to a higher compressibility of the mutant. Under the same pressure conditions, the wildtype protein experienced little effect. These results suggest that the surface mutation (F29W) significantly destabilizes the N-domain of troponin C by altering the packing and dynamics of the hydrophobic core.