Nucleotides increase the internal flexibility of filaments of dephosphorylated Acanthamoeba myosin II.
The actin-activated Mg(2+)-ATPase activity of Acanthamoeba myosin II minifilaments is dependent both on Mg2+ concentration and on the state of phosphorylation of three serine sites at the C-terminal end of the heavy chains. Previous electric birefringence experiments on minifilaments showed a large dependence of signal amplitude on the phosphorylation state and Mg2+ concentration, consistent with large changes in filament flexibility. These observations suggested that minifilament stiffness was important for function. We now report that the binding of nucleotides to dephosphorylated minifilaments at Mg2+ concentrations needed for optimal activity increases the flexibility by about 10-fold, as inferred from the birefringence signal amplitude increase. An increase in flexibility with nucleotide binding is not observed for dephosphorylated minifilaments at lower Mg2+ concentrations or for phosphorylated minifilaments at any Mg2+ concentrations examined. The relaxation times for minifilament rotations that are sensitive to the conformation myosin heads are also observed to depend on phosphorylation, Mg2+ concentration, and nucleotide binding. These latter experiments indicate that the actin-activated Mg2+ concentration, and nucleotide binding. These latter experiments indicate that the actin-activated Mg(2+)-ATPase activity of Acanthamoeba myosin II correlates with both changes in myosin head conformation and the ability of minifilaments to cycle between stiff and flexible conformations coupled to nucleotide binding and release.
Redowicz
M J
MJ
Laboratory of Cell Biology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Korn
E D
ED
Rau
D C
DC
D000048:Acanthamoeba / Q000378:metabolism*; D000227:Adenine Nucleotides / Q000494:pharmacology*; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D009218:Myosins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D015398:Signal Transduction
D000227:Adenine Nucleotides; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins