Fluorescent Actin Filaments Move on Myosin Fixed to a Glass Surface

• Published in: Proceedings of the National Academy of Sciences - PNAS 1986-09, Vol.83 (17), p.6272-6276
• Main Authors: Kron, Stephen J., Spudich, James A.
• Format: Article
• Language: English
• Subjects: Actins; Actins - physiology; Actomyosin - physiology; Adenosine triphosphatases; Adenosine Triphosphate - metabolism; Animals; Biochemistry; Biological and medical sciences; Cell membranes; Conformational dynamics in molecular biology; Dictyostelium; Fundamental and applied biological sciences. Psychology; Glass; Hydrogen-Ion Concentration; In Vitro Techniques; Microfilaments; Microscopy, Fluorescence; Microtubules; Molecular biophysics; Molecules; Muscle Contraction; Myosins - physiology; Phalloidine; Plant cells; Rabbits; Rhodamines; Skeletal muscle; Velocity
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.83.17.6272

Single actin filaments stabilized with fluorescent phalloidin exhibit ATP-dependent movement on myosin filaments fixed to a surface. At pH 7.4 and 24 degrees C, the rates of movement average 3-4 μ m/s with skeletal muscle myosin and 1-2 μ m/s with Dictyostelium myosin. These rates are very similar to those measured in our previous myosin movement assays. The rates of movement are relatively independent of the type of actin used. The filament velocity shows a broad pH optimum between 7.0 and 9.0, and the concentration of ATP required for half-maximal velocity is 50 μ M. Evidence was obtained to suggest that movement of actin over myosin requires at most the number of heads in a single thick filament. This system provides a practical, quantitative myosin-movement assay with purified proteins.