Observation of Mechanical Process for Myosin-Ii by a Dark-Field Microscopy Combined with an Optical Trap

Abstract

Myosin-II molecules are self-assembled and form a muscle. Individual myosin heads in muscle does not seem to interrupt each motion and fully exhibit their function. Myosin heads are connected to the backbone filament via S2-coiled-coil (∼ 60 nm) with non-linear elasticity. Motor domain undergoes Brownian motion (probably > 10 nm fluctuation), but small working stroke (∼ 10 nm) by the single myosin head is fully converted to macroscopic sliding motion of muscle. Thus, flexibility enabling myosin head's Brownian motion reduces molecular friction in muscle, but how small working stroke within the thermal noise can be fully converted to macroscopic motion is still unclear. To answer the question, we should visualize the Brownian process and transition from pre-working state to post-working state. Detection for the process has succeeded using micro-needle method (Kitamura et al., 1999), but detail analysis at modulated conditions was hard due to the difficulty for the method. Here we used dark-field microscopy combined with an optical trap and observed interaction between myosin II S-1 and actin. S1 was tagged with small optically-trapped particle (60 nm gold or 200 nm polystyrene bead), which enabled us to quantify non-processive steps and processive steps concerning a few myosin heads with rapid response time. We will discuss these results and trials of mechanical modulation to the myosin head at sub-milli second time scale to examine strain-dependent force generation like myosin-VI (Iwaki et al., 2009).