Structural transitions in myosin and the origin of contractile force in muscle

Force generation in actively contracting muscle very likely involves a structural transition in each cycling cross-bridge while it is attached to actin. Two different types of force-generating mechanisms are discussed in this paper: (1) the force-generating event results from a change in the effective angle of the myosin head (S-1 subunit) and (2) a conformational change in the α-helical S-2 region of the crossbridge occurs when this portion of the bridge is released from the thick-filament surface. This process causes shortening in S-2 and produces force. Evidence supporting mechanism 2 has been obtained from recent studies of actively contracting muscle fibers and of isolated myosin rod subfragments using an enzyme- (chymotrypsin) probe technique to detect and pinpoint local melting within the α-helical structure. The kinetics of proteolysis and the site of cleavage were determined at various temperatures (5 to 40°C) by electrophoresis of digestion products on SDS gels. We found the cleavage sites to be localized in a restricted segment of S-2 spanning the LMM/HMM hinge region (64,000–90,000 Mr/polypeptide chain from the C-terminus of the myosin rod). The cleavage rate constant for activated muscle fibers in the presence of an ATP-regenerating system was about 100 times larger at each temperature than that for rigor or relaxed muscle fibers, and it showed a marked increase in magnitude with increasing temperature. Comparative plots of the apparent rate constant for cleavage within the S-2 hinge domain and the isometric force generated by active fibers vs MgATP concentration gave closely similar profiles, suggesting a close coupling between the conformational transitions within the S-2 hinge domain and contractile force when the cross-bridges undergo cycling.