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Ca2+-sensitivity of actomyosin ATPase purified from Physarum polycephalum
Abstract:
A contractile protein closely resembling natural actomyosin (myosin B) of rabbit skeletal muscle was extracted from plasmodia of the slime mold, Physarum polycephalum, by protecting the SH-groups with beta-mercaptoethanol or dithiothreitol. Superprecipitation of the protein induced by Mg2+-ATP at low ionic strength was observed only in the presence of very low concentrations of free Ca2+ ions, and the Mg2+-ATPase [EC 3.6.1.3] reaction was activated 2- to 6-fold by 1 muM of free Ca2+ ions. Crude myosin and actin fractions were separated by centrifuging plasmodium myosin B in the presence of Mg2+-PPi at high ionic strength. The crude myosin showed both EDTA- and Ca2+-activated ATPase activities. The Mg2+-ATPase activity of crude myosin from plasmodia was markedly activated by the addition of pure F-actin from rabbit skeletal muscle. Addition of the F-action-regulatory protein complex prepared from rabbit skeletal muscle as well as the actin fraction of plasmodium caused the same degree of activation as the addition of pure F-actin only in the presence of very low concentrations of Ca2+ ion
Insights
Researchers extracted a contractile protein similar to actomyosin from slime mold. This protein
Area of Science:
- Biochemistry
- Cell Biology
- Biophysics
Background:
- Actomyosin is a key contractile protein complex in muscle.
- Slime molds possess unique cellular structures and motility mechanisms.
Purpose of the Study:
- To isolate and characterize contractile proteins from Physarum polycephalum.
- To investigate the calcium ion (Ca2+) dependency of the Mg2+-ATPase activity of Physarum actomyosin.
Main Methods:
- Extraction of contractile proteins from Physarum polycephalum using reducing agents.
- Assay of Mg2+-ATPase activity and superprecipitation.
- Separation of myosin and actin fractions via ultracentrifugation.
- Investigating the effects of Ca2+ and regulatory proteins on ATPase activity.
Main Results:
- A contractile protein resembling rabbit actomyosin was successfully extracted from Physarum polycephalum.
- The Mg2+-ATPase activity and superprecipitation of the extracted protein were found to be dependent on low concentrations of free Ca2+ ions, with a 2- to 6-fold activation observed.
- Crude myosin from Physarum exhibited both EDTA- and Ca2+-activated ATPase activities and its Mg2+-ATPase activity was significantly enhanced by the addition of F-actin.
Conclusions:
- Physarum polycephalum contains a contractile protein system analogous to muscle actomyosin.
- Calcium ions play a crucial regulatory role in the Mg2+-ATPase activity of this slime mold contractile system.
- The interaction between Physarum actin and myosin is regulated by calcium, similar to vertebrate muscle systems.