Submillisecond rotational dynamics of spin-labeled myosin heads in myofibrils.
The rotational motion of crossbridges, formed when myosin heads bind to actin, is an essential element of most molecular models of muscle contraction. To obtain direct information about this molecular motion, we have performed saturation transfer EPR experiments in which spin labels were selectively and rigidly attached to myosin heads in purified myosin and in glycerinated myofibrils. In synthetic myosin filaments, in the absence of actin, the spectra indicated rapid rotational motion of heads characterized by an effective correlation time of 10 microseconds. By contrast, little or no submillisecond rotational motion was observed when isolated myosin heads (subfragment-1) were attached to glass beads or to F-actin, indicating that the bond between the myosin head and actin is quite rigid on this time scale. A similar immobilization of heads was observed in spin-labeled myofibrils in rigor. Therefore, we conclude that virtually all of the myosin heads in a rigor myofibril are immobilized, apparently owing to attachment of heads to actin. Addition of ATP to myofibrils, either in the presence or absence of 0.1 mM Ca2+, produced spectra similar to those observed for myosin filaments in the absence of actin, indicating rapid submillisecond rotational motion. These results indicate that either (a) most of the myosin heads are detached at any instant in relaxed or activated myofibrils or (b) attached heads bearing the products of ATP hydrolysis rotate as rapidly as detached heads.
D000199:Actins / Q000032:analysis; D000251:Adenosine Triphosphatases / Q000032:analysis; D000818:Animals; D004578:Electron Spin Resonance Spectroscopy; D046911:Macromolecular Substances; D009132:Muscles / Q000201:enzymology / Q000648:ultrastructure; D009210:Myofibrils / Q000201:enzymology / Q000648:ultrastructure*; D009218:Myosins / Q000032:analysis* / Q000378:metabolism; D011817:Rabbits; D013113:Spin Labels
D000199:Actins; D046911:Macromolecular Substances; D013113:Spin Labels; D000251:Adenosine Triphosphatases; D009218:Myosins