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Filament formation by purified Physarum myosin
Abstract:
Physarum myosin can be separated from actomyosin by ultracentrifugation, and purified by gel filtration. Unlike actomyosin, myosin is soluble in 0.05 M KCl in the pH range of 6-7. However, in the absence of actin, the slime mold myosin can be precipitated in 0.05 M KCl by the addition of millimolar concentrations of CaCl(2). The precipitates consist of aggregated, short bipolar filaments. Magnesium has a similar effect, but results in the precipitation of more loosely packed aggregates. The length of the compact filaments is 0.45 mum; thus, predominantly tail-to-tail, but also some head-to-tail, interactions occur under these conditions. Since the size and shape of these thick filaments are close to those seen in fixed and sectioned ameboid cells and in platelets, all of these filaments are probably composed of myosins.
Insights
Slime mold myosin, purified via ultracentrifugation and gel filtration, forms short bipolar filaments in low salt conditions with calcium or magnesium. These myosin filaments resemble those found in cells.
Area of Science:
- Biochemistry
- Cell Biology
- Biophysics
Background:
- Myosin is a crucial protein for cellular motility.
- Understanding myosin's self-assembly properties is key to cell mechanics.
Purpose of the Study:
- To characterize the self-assembly behavior of Physarum myosin.
- To investigate the formation of myosin filaments in vitro.
Main Methods:
- Ultracentrifugation to separate myosin from actomyosin.
- Gel filtration for myosin purification.
- Induction of myosin precipitation using CaCl(2) and MgCl(2).
Main Results:
- Physarum myosin is soluble in 0.05 M KCl (pH 6-7) but precipitates with millimolar CaCl(2) or MgCl(2).
- Precipitates form short bipolar filaments (0.45 μm length) with tail-to-tail and head-to-tail interactions.
- Filament morphology resembles in-cell myosin structures.
Conclusions:
- Physarum myosin can self-assemble into bipolar filaments under specific ionic conditions.
- These in vitro formed filaments share structural similarities with cellular myosin structures.
- This suggests a conserved mechanism for myosin filament formation across different cell types.