TRANSIENTS OF FLUORESCENCE POLARIZATION IN SKELETAL-MUSCLE FIBERS LABELED WITH RHODAMINE ON THE REGULATORY LIGHT-CHAIN

Structural changes of the myosin heads were correlated with mechanical events in the cross-bridge cycle by measuring fluorescence polarizati on signals at high time resolution from rhodamine probes bound to myos in regulatory light chains in skeletal muscle fibers. Motions of the c ross-bridges were partially synchronized either by applying quick leng th changes to the fibers during active contractions or by activating t he fibers from rigor by photolysis of caged ATP in the presence of Ca2 +. With fibers in rigor, the fluorescence polarization values indicate that the probe dipoles are quite well ordered and are directed away f rom the muscle fiber axis. After photorelease of ATP from caged ATP, c hanges in polarization signals are consistent with broadening of the d istribution of probe orientations, The signal deflections occur when A TP binds to actomyosin or when the cross-bridges detach, but the orien tational distribution changes surprisingly little during active force development. In contrast, when staircases of quick releases are applie d to labeled fibers during active contractions, the fluorescence polar ization signals suggest a concerted rotation of the probes. The result s indicate that the light chain region of myosin tilts during the quic k release and/or during the tension recovery phase within the next few ms.