Mg-ATPase activity and motility of native thick filaments isolated from the anterior byssus retractor muscle ofMytilus edulis

Abstract

A method for isolating native thick filaments from the anterior byssus retractor muscle (ABRM) ofMytilus edulis is described. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that the isolated thick filament preparation contained mainly paramyosin and myosin but almost no actin. Electron microscopy of negatively stained preparations showed that the isolated thick filaments were tapered at both ends and of various sizes, in the range 5–31 Μm in length and 51–94nm in width in the central region. Central bare zones were observed in the smaller filaments, but were not clearly seen in the larger filaments. Mg-ATPase activity of the isolated thick filaments was activated by skeletal muscle F-actin in a Ca²⁺-dependent manner. The maximal activity was about 20 nmol min⁻¹ mg⁻¹ thick filaments (20‡ C, pH7.0). Motility of the thick filaments attached to latex beads (diameter, 2 Μm) was also studied using the native actin cables of the freshwater alga,Chara. In the presence of Mg-ATP and Ca²⁺, the beads moved along the actin cables at a maximal velocity of about 1 Μm s⁻¹. In the absence of Ca²⁺, almost no movement was observed. These results show that the isolated thick filaments are structurally intact and retain the essential mechanochemical characteristics of the ABRM myosin.