Myosin catalytic domain flexibility in MgADP.
Conventional EPR studies of muscle fibers labeled with a novel alpha-iodoketo spin label at Cys-707 of the myosin head revealed substantial internal domain reorganization on the addition of ADP to rigor fibers. The spin probes that are well-ordered in the rigor state become disordered and form two distinct populations. These orientational changes do not correspond to rotation of the myosin catalytic domain as a whole because other probes (maleimide and iodoacetamide nitroxides attached to the same Cys-707 of myosin head) report only a small (5-10 degrees) torsional rotation and little or no change in the tilt angle [Ajtai et al. (1992) Biochemistry 31, 207-17; Fajer (1994) Biophys. J. 66, 2039-50]. In the presence of ADP, the labeled domain becomes more flexible and executes large-amplitude microsecond motions, as measured by saturation-transfer EPR with rates (tau r = 150 microseconds) intermediate between the rotations of detached (tau r = 7 microseconds) and rigor heads (tau r = 2500 microseconds). This finding contrasts with an absence of global motion of the myosin head in ADP (tau r = 2200 microseconds) as reported by the maleimide spin label. Our results imply that the myosin head in a single chemical state (AM.ADP) is capable of attaining many internal configurations, some of which are dynamic. The presence of these slow structural fluctuations might be related to the slow release of the hydrolysis products of actomyosin ATPase.
Raucher
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Institute of Molecular Biophysics, Florida State University, Tallahassee 32306-3015.
Sár
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CP
Hideg
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Fajer
P G
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D000244:Adenosine Diphosphate / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy; D009218:Myosins / Q000737:chemistry*; D011487:Protein Conformation; D011817:Rabbits; D013113:Spin Labels
C090627:3-(2-iodopropionyl)-1-oxyl-2,5-dihydro-2,2,5,5-tetramethyl-1H-pyrrole; D003497:Cyclic N-Oxides; D013113:Spin Labels; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D009218:Myosins