Internal motions in myosin. 2.

The high-resolution 1H NMR detected internal motions in myosin and myosin subfragment 1 (S1) [Highsmith, S., Akasaka, K., Konrad, M., Goody, R., Holmes, K., Wade-Jardetzky, N., & Jardetzky, O. (1979) Biochemistry 18, 4238--4244] were unperturbed by induced changes in the rate of protein tumbling, and the mobile regions proved inaccessible to added surface-directed paramagnetic probes. The rate of tumbling was changed by changing the solvent viscosity for S1 or by aggregation to thick filaments for myosin. Neither manipulation caused a measurable broadening of the narrow lines in the spectrum. Sulfhydryl-directed covalently attached nitroxide spin-labels, soluble nitroxide spin-labels, and MnCl2 were used to probe the surface. Unique labeling at the fastest reacting thiol of S1 had no effect on the NMR spectrum. Multiple labeling of thiols caused a small but detectable broadening of the narrow peaks. Soluble spin-labels and MnCl2 had a very small effect on the narrow bands even in great excess. The results substantiate the notion that myosin has internal motions that are independent of the overall rate of rotation and suggest that the mobile structure is mainly in the interior of the S1 moiety. This supports a model in which actin quenches the internal motions of myosin by changing the structure of myosin upon binding.