A closer look at energy transduction in muscle

Abstract

Muscular force is the sum of unitary force interactions generated as filaments of myosins move forcibly along parallel filaments of actins, understanding that the free energy required comes from myosin-catalyzed ATP hydrolysis. Using results from conventional biochemistry, our own mutational studies, and diffraction images from others, we attempt, in molecular detail, an account of a unitary interaction, i.e., what happens after a traveling myosin head, bearing an ADP-P(i), reaches the next station of an actin filament in its path. We first construct a reasonable model of the myosin head and actin regions that meet to form the "weakly bound state". Separately, we consider Holmes' model of the rigor state [Holmes, K. C., Angert, I., Kull, F. J., Jahn, W. & Schröder, R. R. (2003) Nature 425, 423-427], supplemented with several heretofore missing residues, thus realizing the "strongly bound state." Comparing states suggests how influences initiated at the interface travel elsewhere in myosin to discharge various functions, including striking the actins. Overall, state change seems to occur by attachment of a hydrophobic triplet (Trp-546, Phe-547, and Pro-548) of myosin to an actin conduit with a hydrophobic guiding rail (Ile-341, Ile-345, Leu-349, and Phe-352) and the subsequent linear movement of the triplet along the rail.

Scheme 1.

Fig. 1.

Specialized “organs” of myosin. The N-terminal 25-kDa (residues 2–202), the upper 50-kDa (207–468), the lower 50-kDa (469–651), and the C-terminal 20-kDa (652–791) segments of the myosin motor domain heavy chain are colored yellowish green, red, ocher, and dark blue, respectively. The lever arm is not shown. Locations of actin-binding surface loops: 1, the second actin-binding loop; 2, the hydrophobic triplet; 3, the proline-rich loop; 4, the C terminus of loop 2; 5, the cardiomyopathy loop; and 6, loop 4. Mutated residues in these loops are indicated as balls. The crystal structure of the motor domain of Dictyostelium myosin with MgADP.VO₄⁻ is adapted from ref. . MgADP.VO₄⁻ is shown as balls in white (carbon), blue (nitrogen), red (oxygen), green (magnesium), and ocher (phosphorus).

Fig. 2.

Stereoview of the weakly bound AM.ADP.P_i complex. The same colors are used for segments of the myosin heavy chain as in Fig. 1, whereas the first actin and the second actin are colored cyan and dark green, respectively. The direction of the pointed end of the actin filament is upward. Myosin residues that are possibly involved in binding for actins (as balls): 1, Lys-652 and Lys-653 (blue); 2, Arg-530 (blue); 3, Lys-573 (blue); 4, Lys-576 and Lys-578 (blue), 5, Asp-374 (red), and 6, Trp-546 (purple), Phe-547 (dark blue), and Pro-548 (pink). The corresponding actin residues are also shown as balls. The cardiomyopathy loop is indicated as a red ribbon marked 7. Crystal structures of the myosin motor domain and monomeric rabbit skeletal actin are adapted from refs. and , respectively. Side chains of some residues (for example, Asp-374, Lys-652, and Lys-653 for myosin and Glu-4 for actin), important for actin binding, are missing in these structures.

Fig. 3.

Comparison between myosin heads in our weakly bound complex model (colors are same as those in Fig. 1) and in the rigor complex model (cyan) reported by Holmes et al. (12). Actins of two models are superimposed, and movements of the myosin head are shown by two curved arrows, a and b. Two actins are omitted, except for several residues (numbers and colors are the same as in Fig. 2) that bind myosin. The direction of the pointed end of the actin filament is upward (straight arrow).

Fig. 4.

The rigor complex of acto-S1. Structures shown in A and B are derived from the same structure reported by Holmes et al. (12). Segments of the myosin heavy chain and the first actin are colored the same as in Fig. 1 or 2, but the second actin is not shown. (A) The lower 50-kDa subdomain is omitted from the myosin structure to look at two attachment sites: a, Lys-652 and Lys-653 (blue balls) of myosin and Asp-24 and Asp-25 (red balls) of actin and b, the cardiomyopathy loop (red ribbon) of myosin and the dead-end surface (colored balls) surrounded by residues 26–30 and 337–338 of actin. The seven-stranded β-sheet is indicated by ribbons colored yellowish green (strand 1, 114–117; strand 2, 120–123; strand 4, 171–176), dark blue (strand 3, 678–684), and red (strand 5, 459–467; strand 6, 249–256; and strand 7, 262–269). The position of Lys-652 and Lys-653 is deduced by supplementing these residues from the crystal structure of the Dictyostelium myosin motor domain with MgADP.VO₄⁻ (10). The light purple ribbon indicates the superimposed Dictyostelium HW helix and the third strand of the β-sheet. A helix of residues 336–352 of the first actin is shown as a ribbon colored cyan. A big slanted arrow indicates the direction of the pointed end of the actin filament. (B) Only the lower 50-kDa subdomain and the C-terminal 20-kDa domain are shown to look at two specific attachment sites of the lower 50-kDa subdomain for the first actin: a, the hydrophobic triplet (purple for Trp-546, dark blue for Phe-547, and pink for Pro-548) of myosin and Ile-341 and Ile-345 (green) of actin and b, the proline-rich loop (pink for Pro-534 and Pro-535 and red for Glu-557) of myosin and Leu-349 (green) and Phe-352 (dark blue) of actin. A linear arrangement of Ile-341, Ile-345, Leu-349, and Phe-352 forms a hydrophobic conduit on actin. The proline-rich loop is supplemented to the helix of residues 515–545 from the Dictyostelium crystal structure (10). The orange ribbon indicates the superimposed Dictyostelium 515–545 helix. Three big arrows indicate a linear movement of the hydrophobic triplet along the conduit, a twisted rotation of a proximal part of the lower 50-kDa subdomain, and a rotation of the converter, respectively. Switch 1 (residues 237–246) and switch 2 (469–476) in the enzyme pocket are shown as ribbons colored red and yellow, respectively, and Arg-247 and Glu-470 are shown as balls colored blue and red, respectively.

Fig. 5.

A serial diagram depicting, step 1, the initial contact between a myosin head and two actins (first, cyan and second, dark green) of a filament; step 2, rotating the myosin head toward the first actin that follows the weak-to-strong transition; step 3, the rotation of the lower 50-kDa subdomain that results in P_i release and movement of the converter–lever arm system; step 4, rolling of the upper 50-kDa subdomain that accelerates the ADP release; and step 5, the release that frees the myosin head to move away toward the next station. The N-terminal 25-kDa domain, the upper 50-kDa subdomain, the lower 50-kDa subdomain, and the converter of myosin are colored yellowish green, red, ocher, and violet, respectively. The blue line in the myosin head indicates the C-terminal heavy-chain segment connecting between an actin-binding loop and the converter. The black spiral indicates a long α-helix that constitutes the core of the lever arm.