Effects of nucleotide on skeletal muscle myosin unfolding in myofibrils by DSC.

The thermal unfolding of myosin in skeletal muscle myofibrils was studied by differential scanning calorimetry (DSC). In the absence of nucleotide two major transitions with Tm of 52 degrees C and 58 degrees C, and a minor transition with Tm of 19 degrees C were detected. The unfolding can be characterized with a total enthalpy of -90 +/- 6.1 mJ/g protein. The major transition with Tm of 58 degrees C was independent of the presence of nucleotide and orthovanadate (Vi), and it can be assigned to the unfolding of the alpha-helical rod part of myosin and partly to actin. In the presence of MgADP, the minor transition shifted to higher temperature, indicating changes between the heavy chain of subfragment-1 and the LC-2 light chain. The transition with Tm of 52 degrees C exhibited a significant broadening and a small shift to lower temperature. It indicates an internal domain or segmental rearrangement of the myosin motor. Upon addition of MgADP and Vi, a shift to higher temperature was observed for the lower major transition, evidencing that with trapped ADP and Vi the intermolecular interactions stabilized the myosin head region.