We have previously measured the process of displacement generation by a single head of muscle myosin (S1) using scanning probe nanometry. Given that the myosin head was rigidly attached to a fairly large scanning probe, it was assumed to stably interact with an underlying actin filament without diffusing away as would be the case in muscle. The myosin head has been shown to step back and forth stochastically along an actin filament with actin monomer repeats of 5.5 nm and to produce a net movement in the forward direction. The myosin head underwent 5 forward steps to produce a maximum displacement of 30 nm per ATP at low load (<1 pN). Here, we measured the steps over a wide range of forces up to 4 pN. The size of the steps (∼5.5 nm) did not change as the load increased whereas the number of steps per displacement and the stepping rate both decreased. The rate of the 5.5-nm steps at various force levels produced a force-velocity curve of individual actomyosin motors. The force-velocity curve from the individual myosin heads was comparable to that reported in muscle, suggesting that the fundamental mechanical properties in muscle are basically due to the intrinsic stochastic nature of individual actomyosin motors. In order to explain multiple stochastic steps, we propose a model arguing that the thermally-driven step of a myosin head is biased in the forward direction by a potential slope along the actin helical pitch resulting from steric compatibility between the binding sites of actin and a myosin head. Furthermore, computer simulations show that multiple cooperating heads undergoing stochastic steps generate a long (>60 nm) sliding distance per ATP between actin and myosin filaments, i.e., the movement is loosely coupled to the ATPase cycle as observed in muscle.
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Recent progress in single-molecule detection techniques is remarkable. These techniques have allowed the accurate determination of myosin-head-induced displacements and how mechanical cycles are coupled to ATP hydrolysis, by measuring individual mechanical events and chemical events of actomyosin directly at the single-molecule level. Here we review our recent work in which we have made detailed measurements of myosin step size and mechanochemical coupling, and propose a model of the myosin motor.
Introduction. Myosin is a biomolecular motor that is responsible for various types of cellular movement including muscle contraction and organelle transport. Movement is caused by the sliding movement of myosin molecules along actin filaments, fuelled by the chemical energy from adenosine tri-phosphate (ATP) hydrolysis. The fundamental problem of how this motor converts the chemical energy into the mechanical energy has yet to be elucidated. There are two major classes of models in terms of the relationship between the chemical reactions and the mechanical events. One class is the deterministic and mechanistic models, such as the lever armswinging model, where the movement of myosin along an actin filament is caused by a power stroke of the tail part of a myosin head acting as a lever arm, tightly coupled to the ATP hydrolysis. Fig. 1(a)): i) During the hydrolysis of a single ATP molecule the displacement of the myosin heads occurs in variable numbers (1-5) of 5.5 nm steps. The size of the individual steps corresponds to the distance between adjacent actin monomers in the actin filaments. This process is stochastic. ii) Steps also occasionally occur in the backward direction. Overall, the motion of myosin is preferentially directed to one end of the actin filament. iii) On applying a load to the myosin, the number of steps decreased and the dwell time Model describing the biased Brownian movement of myosinBy Seiji ESAKI, * ) Yoshiharu ISHII, ** ) and Toshio YANAGIDA * ), ** ), *** ), †) (Communicated by Fumio OOSAWA, M. J. A., Jan. 14, 2003) Abstract: A recent study with single molecule measurements has reported that myosin II, a molecular motor, generates stochastic and multiple steps during the hydrolysis of a single ATP molecule. In order to elucidate the mechanism for the motion of myosin, we traced the movements of individual molecules by simulating the Brownian movements along the potentials created by the interaction between a myosin molecule and an actin filament. We demonstrated that Brownian movement was biased to one direction as observed for myosins by either spatially tilting or temporally fluctuating the height of the potential. We incorporated the biased Brownian movement into an ATP hydrolysis reaction scheme and studied the effects of the load on the movement. The results could successfully explain the movements and mechanical properties of myosin. Thus, it was demonstrated that the movement of myosin is thermally driven and the random motion is biased by the energy released from the ATP hydrolysis.
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