Orientation of paramagnetic probes attached to gizzard regulatory light chain bound to myosin heads in rabbit skeletal muscle.
The orientation of the myosin neck was monitored using electron paramagnetic resonance (EPR) spectroscopy. Gizzard regulatory light chain was labeled with a nitroxide spin probe and exchanged for the native subunit, located in the myosin neck, in rabbit psoas muscle fibers. The EPR spectra of rigor fibers indicated a substantial degree of probe immobilization and showed a strong dependence on the orientation of the fiber axis relative to the magnetic field, indicating that the neck was ordered in this state. Spectra of relaxed fibers at 24 degrees C showed that the neck was disordered, but the spectra of relaxed fibers at 4 degrees C indicated that the neck was partially ordered. Active fibers at the two temperatures produced spectra identical to relaxed fibers, indicating that no novel angles could be seen in the neck during the powerstroke. Proteolytic fragments of myosin, S1 and HMM, were exchanged with labeled light chains and bound to thin filaments in unlabeled fibers. The distribution of probe orientations for HMM was identical to that of labeled rigor fibers, while S1 showed a slightly different distribution, suggesting that the neck is distorted (by a few degrees) by the interactions of the two heads of myosin when bound to actin.
Zhao
L
L
Department of Biochemistry & Biophysics, University of California, San Francisco 94143-0448, USA.
Gollub
J
J
Cooke
R
R
D000818:Animals; D002645:Chickens; D004578:Electron Spin Resonance Spectroscopy; D005895:Gizzard, Avian; D066298:In Vitro Techniques; D009119:Muscle Contraction; D018485:Muscle Fibers, Skeletal / Q000502:physiology; D018482:Muscle, Skeletal / Q000502:physiology*; D009130:Muscle, Smooth / Q000502:physiology*; D018994:Myosin Light Chains / Q000737:chemistry / Q000502:physiology*; D015879:Myosin Subfragments / Q000737:chemistry / Q000502:physiology*; D011817:Rabbits; D013113:Spin Labels; D014336:Troponin; D019209:Troponin C
D018994:Myosin Light Chains; D015879:Myosin Subfragments; D013113:Spin Labels; D014336:Troponin; D019209:Troponin C