Experimental evidence for the contractile activities of Acanthamoeba myosins IA and IB.
The low-shear viscosity of 5-30 microM F-actin was greatly increased by the addition of 0.1-0.5 microM unphosphorylated Acanthamoeba myosins IA and IB. The increase in viscosity was about the same in 2 mM ADP as in the absence of free nucleotide but was much less in 2 mM ATP. The single-headed monomolecular Acanthamoeba myosins were as effective as an equal molar concentration of two-headed muscle heavy meromyosin and much more effective than single-headed muscle myosin subfragment-1. These results suggest that Acanthamoeba myosins IA and IB can cross-link actin filaments as proposed in the accompanying paper (Albanesi, J. P., Fujisaki, H., and Korn, E. D. (1985) J. Biol. Chem. 260, 11174-11179) to explain the actin-dependent cooperative increase in actin-activated Mg2+-ATPase activity as a function of the concentration of myosin I. Superprecipitation occurred when phosphorylated myosin IA or IB was mixed with F-actin. In addition to myosin I heavy chain phosphorylation, superprecipitation required Mg2+ and ATP. ATP hydrolysis was linear during the time course of the superprecipitation, and inhibitors of ATP hydrolysis inhibited superprecipitation. A small, dense contracted gel was formed when the reaction was carried out in a cuvette, and a birefringent actomyosin thread resulted from superprecipitation in a microcapillary. The rate and extent of superprecipitation depended on the actin and myosin I concentrations with maximum superprecipitation occurring at an actin:myosin ratio of 7:1. These results provide strong evidence for the ability of Acanthamoeba myosins IA and IB to perform contractile and motile functions.
D000199:Actins / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000656:Amoeba / Q000201:enzymology*; D000818:Animals; D007700:Kinetics; D046911:Macromolecular Substances; D008274:Magnesium / Q000378:metabolism; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D014783:Viscosity
D000199:Actins; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D009218:Myosins; D008274:Magnesium