Related Experiment Videos
A structural difference between filaments of phosphorylated and dephosphorylated Acanthamoeba myosin II revealed by
1Laboratory of Biochemistry and Metabolism, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Abstract:
The actin-activated Mg(2+)-ATPase activity of filamentous Acanthamoeba myosin II is regulated by the state of phosphorylation of three sites at the C terminus of each heavy chain. This phosphorylation at the tip of the tails of monomers in a bipolar filament abolishes the activity of sites some 90 nm distant in the globular heads. Previous studies with copolymeric filaments of phosphorylated and dephosphorylated monomers strongly indicated that the activity of each monomer in a filament is dependent on the level of phosphorylation of neighboring monomers in the filament. We report here electric birefringence measurements showing that, although the overall structures of phosphorylated and dephosphorylated filaments are very similar, large, Mg2+ concentration-dependent differences in internal motion and flexibility are observed. Filaments of dephosphorylated myosin II appear to be about 50-fold stiffer than filaments of phosphorylated myosin II at 4 mM Mg2+. These results are consistent with a model in which the stiffness of the putative hinge region within the rod-like tail of each monomer is determined by the phosphorylation state of the C-terminal tails of overlapping, neighboring monomers. The flexibility of the filaments appears to be directly related to their actin-activated Mg(2+)-ATPase activity.
Insights
Phosphorylation of Acanthamoeba myosin II filaments alters their flexibility and Mg(2+) ATPase activity. Dephosphorylated filaments are significantly stiffer, impacting overall motor function.
Area of Science:
- Molecular and Cellular Biology
- Biochemistry
- Biophysics
Background:
- Actin-activated Mg(2+)-ATPase activity of Acanthamoeba myosin II is regulated by C-terminal heavy chain phosphorylation.
- Phosphorylation of myosin II tails affects the activity of distant sites in the globular heads.
- Previous studies suggested monomer activity in filaments depends on neighboring monomer phosphorylation.
Purpose of the Study:
- To investigate the impact of phosphorylation on the physical properties and flexibility of Acanthamoeba myosin II filaments.
- To correlate filament flexibility with actin-activated Mg(2+)-ATPase activity.
Main Methods:
- Electric birefringence measurements were used to assess filament structure and dynamics.
- Mg(2+) concentration-dependent differences in internal motion and flexibility were analyzed.
- Filament stiffness was compared between phosphorylated and dephosphorylated myosin II.
Main Results:
- Phosphorylated and dephosphorylated myosin II filaments exhibit similar overall structures but differ significantly in internal motion and flexibility.
- Filaments of dephosphorylated myosin II are approximately 50-fold stiffer than phosphorylated filaments at 4 mM Mg(2+).
- Filament flexibility is directly correlated with actin-activated Mg(2+)-ATPase activity.
Conclusions:
- The phosphorylation state of C-terminal tails influences the stiffness of the hinge region in neighboring myosin II monomers within a filament.
- Myosin II filament flexibility is a key determinant of its actin-activated Mg(2+)-ATPase activity.
- These findings support a model where inter-monomer interactions, modulated by phosphorylation, regulate myosin II motor function.