Thermal transitions of myosin and its helical fragments. I. Shifts in proton equilibriums accompanying unfolding

Abstract

The thermal transitions of myosin and its helical fragments have been studied with pH as the observable. Heating unbuffered solutions of these proteins near their pI values causes an abrupt rise in pH at a characteristic temperature (the "melting temperature," Tm) which is due to structural changes within the protein. Since the pH shift turns out to be insensitive to the degree of protein aggregation, we have obtained acceptable melting curves even under conditions where the protein coagulates during melting. The melting profiles and Tm vlaues of myosin, myosin rod, and light meromyosin have been found to be remarkably similar (Tm equal to 40 plus or minus 1 degree, 0.5 M KCl, pH 5.9). Proton binding which occurs during melting coincides with the unfolding of a section of myosin rod. Taken in the context of other studies, the proton binding is thought to occur near the "hinge region."