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 polarization signals at high time resolution from rhodamine probes bound to myosin regulatory light chains in skeletal muscle fibers. Motions of the cross-bridges were partially synchronized either by applying quick length changes to the fibers during active contractions or by activating the 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 from the muscle fiber axis. After photorelease of ATP from caged ATP, changes in polarization signals are consistent with broadening of the distribution of probe orientations. The signal deflections occur when ATP binds to actomyosin or when the cross-bridges detach, but the orientational distribution changes surprisingly little during active force development. In contrast, when staircases of quick releases are applied to labeled fibers during active contractions, the fluorescence polarization signals suggest a concerted rotation of the probes. The results indicate that the light chain region of myosin tilts during the quick release and/or during the tension recovery phase within the next few ms.
Allen
T S
TS
Pennsylvania Muscle Institute, University of Pennsylvania, Philadelphia 19104, USA.
Sabido-David
C
C
Ling
N
N
Irving
M
M
Goldman
Y E
YE
D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000737:chemistry; D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D005454:Fluorescence Polarization; D066298:In Vitro Techniques; D008954:Models, Biological; D009119:Muscle Contraction / Q000502:physiology; D009218:Myosins / Q000737:chemistry* / Q000502:physiology; D010782:Photolysis; D016658:Psoas Muscles / Q000737:chemistry* / Q000502:physiology; D011817:Rabbits; D012235:Rhodamines
D012235:Rhodamines; C029273:P(3)-1-(2-nitro)phenylethyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D009218:Myosins