Structure and stability of wildtype and F29W mutant forms of the N-domain of avian troponin C subjected to high pressures

摘要

The N-domain of troponin C (resdues 1-90) regulates muscle contraction through conformational changes induced by Ca~(2+) binding. A mutant form of this domain of avian troponin C (F29W) has been used in previous studies to observe avian troponin C (F29W) has ben used in previous studies to observe conformational changes that occur upon Ca~(2+) 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 Ca~(2+). We found that the mutant protein at 5 kbar pressures had a destabilized #beta# -sheet between the Ca~(2+) -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 experinced little effect. These results suggest that the surface mutation (F29W) significantly destabilizes the N-dimain of troponin C by altering the packing and dynamics of the hydrophobic core.