Isolation and characterization of myosin from amoebae of Physarum polycephalum.
Myosin was isolated from amoebae of Physarum polycephalum and compared with myosin from plasmodia, another motile stage in the Physarum life cycle. Amoebal myosin contained heavy chains (Mr approximately 220,000), phosphorylatable light chains (Mr 18,000), and Ca2+-binding light chains (Mr 14,000) and possessed a two-headed long-tailed shape in electron micrographs after rotary shadow casting. In the presence of high salt concentrations, myosin ATPase activity increased in the following order: Mg-ATPase activity less than K-EDTA-ATPase activity less than Ca-ATPase activity. In the presence of low salt concentrations, Mg-ATPase activity was activated approximately 9-fold by skeletal muscle actin. This actin-activated ATPase activity was inhibited by micromolar levels of Ca2+. Amoebal myosin was indistinguishable from plasmodial myosin in ATPase activities and molecular shape. However, the heavy chain and phosphorylatable light chains of amoebal myosin could be distinguished from those of plasmodial myosin in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, peptide mapping, and immunological studies, suggesting that these are different gene products. Ca2+-binding light chains of amoebal and plasmodial myosins were found to be identical using similar criteria, supporting our hypothesis that the Ca2+-binding light chain plays a key role in the inhibition of actin-activated ATPase activity in Physarum myosins by micromolar levels of Ca2+.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002918:Chymotrypsin; D007700:Kinetics; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008970:Molecular Weight; D009218:Myosins / Q000302:isolation & purification* / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D010804:Physarum / Q000254:growth & development / Q000378:metabolism*
D046911:Macromolecular Substances; D010446:Peptide Fragments; D002918:Chymotrypsin; D000251:Adenosine Triphosphatases; D009218:Myosins; D002118:Calcium