Movement of scallop myosin on Nitella actin filaments: regulation by calcium.
In order to determine if Ca2+ regulates scallop myosin movement on actin, we have measured motility of scallop myosin along actin filaments using a direct visual assay. This procedure consists of covalently linking myosin to 1-micron beads and pipetting them onto a parallel array of actin filaments located on the cytoplasmic face of a Nitella internodal cell. In the absence of Ca2+, scallop myosin-coated beads exhibit no directed motion; however, in the presence of pCa2+ of greater than 5.84, these beads undergo linear translocations with average velocities of 2.0 micron/s. This Ca2+ -sensitive motility requires the presence of regulatory light chains on the scallop myosin. Removal of regulatory light chains with 10 mM EDTA produces a "desensitized" myosin, no longer sensitive to Ca2+, which moves at rates of 0.09-0.3 micron in the presence or absence of Ca2+. Readdition of regulatory light chains to preparations of desensitized myosin once again confers Ca2+-sensitive motility. The Ca2+ dependence of scallop-myosin motility shows a sharp transition, consistent with the Ca2+ activation sensitivity of the actin-activated ATPase. Furthermore, relative rates of movement of calcium-regulated myosins from various molluscan species are consistent with their respective rates of ATP hydrolysis. Thus, myosin motility along actin filaments provides a sensitive and direct assay of myosin activity and is suitable for studying myosin regulation.
D000199:Actins / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D000460:Chlorophyta / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D004492:Edetic Acid / Q000494:pharmacology; D008974:Mollusca / Q000032:analysis*; D009218:Myosins / Q000378:metabolism*
D000199:Actins; D004492:Edetic Acid; D000251:Adenosine Triphosphatases; D009218:Myosins; D002118:Calcium