title: The interaction of F-actin with phosphorylated and unphosphorylated myosins IA and IB from Acanthamoeba castellanii.
abstract: Myosins IA and IB from Acanthamoeba castellanii are single-headed molecules which, upon phosphorylation of their heavy chains by a specific kinase, express actin-activated Mg2+-ATPase activity. These myosins show no tendency to self-associate under assay conditions, a property which allows unambiguous kinetic and actin-binding data to be obtained. Both myosin isoenzymes exhibit a complex dependence of actomyosin ATPase activity on F-actin concentration. A conventional hyperbolic dependence is observed at low concentrations of F-actin but at higher F-actin concentrations, inhibition and then apparent reactivation are seen to occur. From those early portions of the velocity profiles which do not deviate from simple Michaelis-Menten type kinetics, values for the Vmax (10 s-1 for myosin IA, 18 s-1 for myosin IB) and KATPase (0.25 microM for myosin IA, 0.30 microM for myosin IB) were calculated. Similar Vmax values were obtained from the reactivation segment of the kinetic data. The KATPase values are very similar to the directly measured dissociation constants (KD) of 0.10 microM for myosin IA and 0.25 microM for myosin IB. Phosphorylation of the myosin heavy chain, which elicits a greater than 20-fold activation of the actomyosin ATPase, has no effect on the binding of myosin to F-actin. This finding supports the conclusion that phosphorylation of myosins IA and IB accelerates one or more catalytic steps of the actomyosin I ATPase reaction at both low and high concentrations of F-actin.
mesh_terms: D000199:Actins / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000656:Amoeba; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase; D004591:Electrophoresis, Polyacrylamide Gel; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation; D011817:Rabbits; D012995:Solubility
chemical_list: D000199:Actins; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins