Orientation of spin-labeled light chain 2 of myosin heads in muscle fibers.
Electron paramagnetic resonance (e.p.r.) spectroscopy has been used to monitor the orientation of spin labels attached rigidly to a reactive SH residue on the light chain 2 (LC2) of myosin heads in muscle fibers. e.p.r. spectra from spin-labeled myosin subfragment-1 (S1), allowed to diffuse into unlabeled rigor (ATP-free) fibers, were roughly approximated by a narrow angular distribution of spin labels centered at 66 degrees relative to the fiber axis, indicating a uniform orientation of S1 bound to actin. On the other hand, spectra from spin-labeled heavy meromyosin (HMM) were roughly approximated by two narrow angular distributions centered at 42 degrees and 66 degrees, suggesting that the LC2 domains of the two HMM heads have different orientations. In contrast to S1 or HMM, the spectra from rigor fibers, in which LC2 of endogenous myosin heads was labeled, showed a random orientation which may be due to distortion imposed by the structure of the filament lattice and the mismatch of the helical periodicities of the thick and thin filaments. However, spectra from the fibers in the presence of ATP analog 5'-adenylyl imidodiphosphate (AMPPNP) were approximated by two narrow angular distributions similar to those obtained with HMM. Thus, AMPPNP may cause the LC2 domain to be less flexible and/or the S2 portion to be more flexible, so as to release the distortion of the LC2 domain and make it return to its natural position. At high ionic strength, AMPPNP disoriented the spin labels as ATP did under relaxing conditions, suggesting that the myosin head is detached from and/or weakly (flexibly) attached to a thin filament.
Arata
T
T
Department of Biology, Faculty of Science, Osaka University, Japan.
D000255:Adenosine Triphosphate; D000266:Adenylyl Imidodiphosphate; D000818:Animals; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D009132:Muscles / Q000032:analysis* / Q000648:ultrastructure; D015879:Myosin Subfragments; D009218:Myosins; D009994:Osmolar Concentration; D011817:Rabbits; D013113:Spin Labels
D015879:Myosin Subfragments; D013113:Spin Labels; D000266:Adenylyl Imidodiphosphate; D000255:Adenosine Triphosphate; D009218:Myosins