EPR Studies of Signal Transduction in Muscle Contraction.

Abstract The signal transduction pathway (Ca2+ activation) was investigated by observing the orientation and dynamics of the labeled regulatory proteins reconstituted into the thin filaments of muscle fibers. Troponin C labeled at Cys-97 was strongly immobilized by interaction with other regulatory proteins, which allowed an accurate determination of the orientation during activation with Ca and/or myosin heads. The addition of Ca to reconstituted fibers resulted in a 20° rotation of the labeled domain accompanied by an increase of orientational disorder, whereas myosin head attachment induced a 30° rotation with similar disordering. The difference in the degree of rotation reflects the different modes of thin filament activation by Ca and by myosin heads. Tropomyosin labeled with MSL at Cys-197 and Cys-36 was reconstituted into ghost fibers. Mobility of the labeled domains was not inhibited by the interaction with the thin filament. The large scale mobility of tropomyosin was modulated to a small extent by interaction with troponin, Ca, caldesmon, or myosin heads. These findings suggest weak, nonstereospecific interactions of tropomyosin and actin, with tropomyosin hovering over the surface of the thin filament.