Ca(2+) causes release of Myosin heads from the thick filament surface on the milliseconds time scale.

摘要

We have used electron microscopy to study the structural changes induced when myosin filaments are activated by Ca(2+). Negative staining reveals that when Ca(2+) binds to the heads of relaxed Ca(2+)-regulated myosin filaments, the helically ordered myosin heads become disordered and project further from the filament surface. Cryo-electron microscopy of unstained, frozen-hydrated specimens supports this finding, and shows that disordering is reversible on removal of Ca(2+). The structural change is thus a result of Ca(2+) binding alone and not an artifact of staining. Comparison of the two techniques suggests that negative staining preserves the structure induced by Ca(2+)-binding. We therefore used a time-resolved negative staining technique to determine the time scale of the structural change. Full disordering was observed within 30ms of Ca(2+) addition, and had started to occur within 10ms, showing that the change occurs on the physiological time scale. Comparison with studies of single heavy meromyosin molecules suggests that an increased mobility of myosin heads induced by Ca(2+) binding underlies the changes in filament structure that we observe. We conclude that the loosening of the array of myosin heads that occurs on activation is real and physiological; it may function to make activated myosin heads freer to contact actin filaments during muscle contraction.