Essential "ankle" in the myosin lever arm.

C ellular motors are fascinating machines that function by undergoing successive conformational changes that require joints in their structure. Where these are located is particularly critical for molecular motors that produce force with relatively rigid lever arms, such as myosins (1). A long-standing paradox in myosin function may finally be understood from structural insights provided by Cohen and colleagues in PNAS (2). Although highly flexible joints are necessary to turn off their activity, these molecular motors have evolved to control such compliance in active heads to produce force under various strain. The structure of the light chain-binding domain (LCD) of scallop catch muscle myosin II (2) describes a unique hinge in this lever arm that could be essential for the regulation of the motor and might also help explain how strain can promote the attachment of the second head of the myosin molecule to the actin filament in isometric contraction (3) and the efficient increase of the resistance of active muscle to stretch (4). Myosin II forms the thick filaments of muscle that slide against the actincontaining thin filaments to allow muscle contraction. Nonmuscle myosins II are also critical for a number of functions in all eukaryotic cells, including cell migration and cytokinesis. To convert chemical energy into force production, these motors amplify the conformational changes of their motor domain, thanks to a lever arm composed of a converter subdomain followed by a C-terminal elongated region of the myosin head or LCD. The swing of the lever arm upon strong binding to actin is coupled with the release of the ATPase products and produces a stroke of approximately 10 nm (1) (Fig. 1A). The LCD of myosin II is a ternary complex composed of a long heavy chain α-helical segment (containing two special IQ motifs) stabilized by the recruitment of members of the calmodulin superfamily, namely an essential light chain (ELC) on IQ1 close to the converter and a distal regulatory light chain (RLC) on IQ2. The “WxW” motif in this IQ2 is characteristic and corresponds to the hook region where a rather acute bend occurs in the heavy chain. Myosins II are double-headed molecules, and the region that follows IQ2 corresponds to a dimeric heavy chain coiledcoil. Although the first part of the coiledcoil or S2 fragment allows the two myosin heads to be connected to the thick filament via an elongated rope, the remainder of the coiled-coil contains triggering sequences that allow assembly of the myosin molecules to form the helical thick (striated muscle) or side-polar filaments (smooth muscle). The ATPase and actin binding potential of these powerful motors need to be controlled in the cell, and failure to do so can promote the development of cancers (5). Although thin filament regulation is predominant in striated muscles, myosinlinked (thick filament) regulation was discovered in the 1970s with molluscan catch muscle (6), in which direct Ca binding on the ELC is required to switch on myosin activity. In smooth and nonmuscle myosin II, Ca activates the myosin light chain kinase that phosphorylates the RLC at position S19 (7). It is becoming clear that the sequence of these motors has evolved not only to optimize their motor activity but also to conserve the features necessary for the switch between inactivated (“off”) and activated (“on”) states of the motor. Regulatory sites within the LCD control the conformation of the hinges to promote intramolecular interactions between the two heads and the S2 region in the off state (dephosphorylated or Ca-free state) (8–11). Phosphorylated or Cabound LCD activates the motor because both heads become free and disordered and can interact with actin to produce force (12). As an extension of the converter subdomain, the LCD is a major component of the lever arm and as such must be relatively rigid. Because flexibility in the LCD is necessary for myosin regulation, it is critical to identify the hinges in the LCD, how they operate depending on the state of the motor, and how they are controlled. Thus, the study published in PNAS (2) is remarkable in that it reveals that variation can occur in the angle of the hook. This provides insights regarding how the distal “ankle” joint of the myosin motor head may operate. In addition to the compliance within the motor domain, three hinges have now been identified in the myosin II lever arm (Fig. 1). A pliant region between the converter and the ELC (13, 14) can operate in the primed ADP.Pi state of the motor and allows exploration of various lateral orientations of the heads that likely contribute to efficiency in rebinding to the actin filament. In states strongly bound to actin, the presence of the N-terminal and SH3 subdomains of the motor next to the ELC limits the compliance at this point for heads in which the lever arm has swung to a down position (Fig. 1A). The second joint in the LCD corresponds to Fig. 1. (A) Myosin powerstroke is illustrated with the motor domain (including SH3 domain in pink) bound to F-actin and with the lever arm (converter and LCD) in two positions: the primed (prepowerstroke, PPS) and down (Rigor) positions. Three hinges (1–3) are indicated. The newly identified “ankle” hinge involves the characteristic “hook” in the RLC binding region. The angle about the hook can be either obtuse (gray helix) or acute (red helix). (B) The unstrained myosin II dimer binds to F-actin using only one head. Strain applied to the dimer in combination with the structural change in the ankle region may promote binding of the second head to the actin filament. (C) The off state of myosin II promotes asymmetric interactions between the two heads (depicted using similar colors as in A) and the S2 coiled-coil region (gray) of the molecule.