Fluorescence Resonance Energy Transfer in Acto-Myosin Complexes

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Myosin, the best known and most studied actin-binding protein, is a molecular motor protein that converts the chemical energy of ATP hydrolysis into mechanical work during muscle contraction, cytokinesis, and other motile functions of cells. At the molecular level muscle contraction is believed to result from the relative sliding of interdigitated thick (composed primarily of myosin) and thin (composed of actin and a host of calcium-regulatory proteins) filaments past one another. This is thought to be a cyclic process in which the hydrolysis of ATP mediates the interaction between actin and myosin (Eisenberg and Hill, 1985). The ATPase cycle of myosin consists of at least four distinct structural states. Myosin is initially bound to actin in the absence of nucleotide, or in a rigor complex (AM), at the end of the contractile cycle. Upon the addition of ATP (M.ATP), the affinity of myosin for actin is significantly reduced, causing dissociation of the acto-myosin complex. ATP is then hydrolyzed to form a second weakly bound state (M.ADP.Pi) which, like the M.ATP state, has a relatively low affinity for actin. During the process of releasing the products of ATP hydrolysis (ADP and Pi), a transition from “weak” to “strong” binding occurs, due to an increase in affinity for actin, and myosin generates force by changing the angle between the cross-bridge relative to the actin filament (i.e., the power stroke). Finally, myosin returns to the rigor state (AM), which is the highest-affinity state between actin and myosin. It is critically important to understand structurally how the binding of ATP to myosin greatly reduces its affinity for actin, and how the hydrolysis of ATP primes the cross-bridge (myosin head) for rebinding to actin in the strongly bound state and the subsequent power stroke, if chemomechanical coupling and the molecular mechanism of muscle contraction are ever to be completely understood.