Single-headed Scallop Myosin and Regulation

Single-headed scallop myosin (shM) was prepared by papain digestion of filamentous scallop myosin and purified by hydrophobic interaction chromatography. The shM preparation consisted of equimolar amounts of polypeptides corresponding to an intact heavy chain, rod chain, essential light chain, and regulatory light chain. In electron micrographs the shape of shM showed the presence of a single head domain to which a normal looking rod was attached. Myosin and shM bound Ca 2+ with association constants of 5 × 10 6 and 11 × 10 6 M −1 , respectively. The ATPase activity of shM was activated about 3-fold by Ca 2+ . Both heads of myosin and shM had comparable ATPase activities in the presence of Ca 2+ . The activation of the ATPase activity of single-headed scallop myosin by Ca 2+ paralleled closely the Ca 2+ binding, in sharp contrast to the activation of intact myosin by Ca 2+ , which is highly cooperative. Single turnover experiments of myosin with radioactive ATP gave a half-life for the ATPase cycle of ∼3 min in the presence of EGTA, whereas that of single-headed myosin was shorter than ∼30 s, which was the resolution time of these measurements. The results suggest that the presence of two heads, as well as the attachment of the head to the coiled coil rod, contribute to the regulation of scallop myosin by Ca 2+ .