The isolation of microquantities of myosin from Amoeba proteus and Chaos carolinensis

Myosin has been isolated from the large carnivorous amoebae Chaos carolinensis and Amoeba proteus to 90% purity with the aid of microtechniques. These techniques for electrophoresis, dialysis, colorimetric protein, and ATPase assays and centrifugation permitted the isolation, purification, and characterization of microgram quantities of myosin from less than 1 ml of packed cells. The purification utilized sucrose extraction of the actomyosin, low ionic strength precipitation, and final separation of the actin and myosin by gel filtration in the presence of KI. The properties of carnivorous amoeba myosin are similar to those of several other cytoplasmic myosins. Its heavy chain molecular weight is greater than that of the heavy chain of rabbit skeletal muscle myosin. The ATPase activities of myosin from C. carolinensis and A. proteus were almost identical and demonstrated inhibition by EDTA and Mg⁺⁺ and activation by Ca⁺⁺ in the presence of 0.6 m KCl and the absence of actin. Actin activated the Mg⁺⁺ ATPase activity by an average factor of 6. No cofactor protein was required for this activity. These properties are similar to those of myosin isolated from Starfish eggs, Physarum, and Dictyostelium. Carnivorous amoeba myosin bound rabbit muscle F actin in the absence but not in the presence of Mg⁺⁺ and ATP.