A biochemical study of flagellar dynein from starfish spermatozoa: protein components of the arm structure.

Half of the adenosine triphosphatase (dynein) activity of starfish sperm tail axonemes was extracted with 0.6 m KCl-10 mm Tris · HCl (pH 7.8)-0.1 mm EDTA-0.5 mm dithiothreitol (KCl-EDTA), while with 1 mm Tris · HCl (pH 7.8)-0.1 mm EDTA-0.5 mm dithiothreitol (Tris-EDTA) around 90% of the activity was extracted. The main adenosine triphosphatase (ATPase) in the KCl-EDTA extract had a sedimentation coefficient of 20S and that in the Tris-EDTA extract had a sedimentation coefficient of 12S. The effects of divalent cations, pH, and an SH-blocking reagent and the K_m for ATP were different for the activities of the two forms of dynein ATPase. These two forms of dynein can interconvert to some extent when the ionic strength of the medium is changed. In a medium suitable for recombination of dynein to outer doublet microtubules (recombination buffer, 20 mm Tris-HCl (pH 7.6)-2 mm MgCl₂-0.5 mm dithiothreitol), the 20S ATPase converted to a 24S ATPase. Recombination of the ATPase activity from the KCl-EDTA extract was almost complete while that from the Tris-EDTA extract was around 50%. Outer arms disappeared preferentially by the treatment with KCl-EDTA, and the extracted arms could be reconstituted in the recombination buffer. In the case of the Tris-EDTA extraction, both the outer and inner arms disappeared and the reconstitution of the arms could not be confirmed. From the above results it can be considered that the 20 or 24S dynein represented the arm structure. The 20 or 24S ATPase fraction contained two large polypeptide chains as main components having electrophoretic mobilities in the presence of sodium dodecylsulfate similar to those of Tetrahymena ciliary dyneins and of sea urchin sperm flagellar dyneins. The relationship between these chains and dynein subunits is discussed.