X-ray diffraction studies on the structural changes of rigor muscles induced by binding of phosphate analogs in the presence of MgADP.
To clarify the structure of the ATP hydrolysis intermediates (ADP.Pi bound state) formed by actomyosin crossbridges, the effects of various phosphate analogs in the presence of MgADP on the structures of the thin and thick filaments in glycerinated rabbit psoas muscle fibers in the rigor state have been investigated by X-ray diffraction with a short exposure time using synchrotron radiation. When MgADP and phosphate analogs such as metallofluorides (BeFx = 3,4 and AlF4) and vanadate (VO4(Vi)) were added to rigor fibers in the presence of the ATP-depletion backup system, the intensities of the actin-based layer lines were markedly weakened. The greatest effect (approximately 50% decrease in intensity) was observed in the presence of BeFx among the analogs examined. The intensity distribution of the 5.9 nm actin-based layer line shifted towards that observed in the Ca(2+)-activated fibers, while the first actin layer line at approximately 1/36.7 nm-1 retained a rigor-like profile with an intensity weakened by approximately 50%. The intensity of the equatorial 10 reflection increased while that of the 11 reflection changed little, resulting in only a small increase (approximately 1.7 fold) in the intensity ratio of the 10 to the 11 reflection. No resting-like pattern appeared upon the addition of MgADP and BeFx. These results indicate that a substantial fraction (approximately 40%) of the myosin heads dissociate from actin but the detached heads remain in the vicinity of the actin filaments when MgADP and BeFx bind. The states produced by binding phosphate analogs to a rigor muscle differ from the resting-like state produced by adding them to a contracting muscle (Takemori et al., J. Biochem. (Tokyo) 117 (1995) 603-608). Our conclusion put forward to explain the data is that one of the two heads of a crossbridge is detached and the other retains a rigor-like attachment.
Kim
D S
DS
Division of Biophysical Engineering, Graduate School of Engineering Science, Osaka University, Japan.
Takezawa
Y
Y
Ogino
M
M
Kobayashi
T
T
Arata
T
T
Wakabayashi
K
K
D000244:Adenosine Diphosphate / Q000737:chemistry*; D000818:Animals; D006868:Hydrolysis; D018482:Muscle, Skeletal / Q000737:chemistry*; D010710:Phosphates / Q000737:chemistry*; D011817:Rabbits; D012298:Rigor Mortis / Q000378:metabolism*; D014961:X-Ray Diffraction
D010710:Phosphates; D000244:Adenosine Diphosphate