Dynamics of actomyosin interactions in relation to the cross-bridge cycle

Abstract

Transient kinetic measurements of the actomyosin ATPase provided the basis of the Lymn-Taylor model for the cross-bridge cycle, which underpins current models of contraction. Following the determination of the structure of the myosin motor domain, it has been possible to introduce probes at defined sites and resolve the steps in more detail. Probes have been introduced in the Dicytostelium myosin II motor domain via three routes: (i) single tryptophan residues at strategic locations throughout the motor domain; (ii) green fluorescent protein fusions at the N and C termini; and (iii) labelled cysteine residues engineered across the actin-binding cleft. These studies are interpreted with reference to motor domain crystal structures and suggest that the tryptophan (W501) in the relay loop senses the lever arm position, which is controlled by the switch 2 open-to-closed transition at the active site. Actin has little effect on this process per se. A mechanism of product release is proposed in which actin has an indirect effect on the switch 2 and lever arm position to achieve mechanochemical coupling. Switch 1 closing appears to be a key step in the nucleotide-induced actin dissociation, while its opening is required for the subsequent activation of product release. This process has been probed with F239W and F242W substitutions in the switch 1 loop. The E706K mutation in skeletal myosin IIa is associated with a human myopathy. To simulate this disease we investigated the homologous mutation, E683K, in the Dictyostelium myosin motor domain.

Motor proteins and molecular motors: how to operate machines at the nanoscale

“…Bulk biochemical experiments have measured rates of chemical reactions associated with motor proteins activity [28, 32, 129–132]. The application of well-established chemical-kinetic methods that include stopped flow, isotope exchange, fluorescent labeling and temperature quenching, has shown that motor proteins catalytic activities involve a complex network of biochemical states and conformations [129,131], although for many myosins and kinesins motor proteins one or few dominating biochemical pathways related to ATP hydrolysis have been identified [28,32,64,129,130].…”

Myosinopathies: pathology and mechanisms

“…In vitro motility studies on myosin IIa isolated from single muscle fibers of patients carrying the E706K mutation clearly showed a marked reduction of speed [38]. Furthermore, studies on the motor function of E683K mutant myosin of Dictyostelium discoideum, which is equivalent to the human MyHC IIa E706K mutation, demonstrated a threefold reduction of the ATP hydrolysis step followed by the slower acto-myosin dissociation [87]. Consequently, these effects lead to a reduced velocity of contraction.…”

The structure of the rigor complex and its implications for the power stroke

“…As is discussed by Sweeney & Houdusse (2004), the commonly occurring post-rigor state is likely to be the form of myosin that rapidly releases from actin after the rebinding of ATP at the end of the power stroke (for further discussion see Zeng et al 2004). (Rayment et al 1993a) shown as a ribbon diagram in the orientation it would take on binding to actin viewed (a) from the pointed (À)-end of the actin filament and (b) at right angles to the actin filament.…”