Introduction to Muscle Contraction

Introduction to Muscle Contraction, Part 2

Movement of the lever arm

Studies of the cross-bridge movement were undertaken by time resolved studies of contracting frog muscle using low angle x-ray fibe diffraction (Huxley et al. 1981; Irving et al. 1992). While these results are fully consistent with the swinging cross-bridge theory, the complexity of the system and low resolution of the method precludes an unambiguous interpretation. Therefore a simpler model system has been studied - decorated actin - an actin filament with a myosin cross-bridge bound to each actin. While the structure of decorated actin in the rigor state (no bound nucleotide) has been extensively studied (Moore et al. 1970) (Milligan and Flicker 1987) corresponding studies in the presence of ATP are difficult since the binding of ATP leads to rapid dissociation of the cross-bridges from actin.

Time-resolved electron micrograph studies in fact show no bulk change of the cross bridge orientation on binding ATP before dissociation takes place(Pollard et al. 1993) whereby a reorientation of the lever arm would not have been detected at the resolution attainable. High resolution electron micrographs of actin decorated with smooth muscle myosin, however, show a 30-35° rotation of the lever arm on binding ADP (Jontes et al. 1995; Whittaker et al. 1995). Although the main movement of the lever arm would be expected to be associated with phosphate release since this is a step associated with a large change in free energy, some fraction of the movement could arise from ADP binding and release. Moreover, this movement should be recoverable on adding ADP to actomyosin, which indeed it is. Although the effect has only been found in smooth muscle myosins it is generally important in providing the first direct demonstration of a nucleotide-induced lever-arm swing. More recent work using a spin label attached to a light chain also supports this result (Gollub et al. 1996). Moreover, x-ray diffraction studies of muscle fibres loaded with exogenous smooth muscle cross-bridges show the predicted changes in the fibre diffraction pattern resulting from the lever arm swing on binding ADP (Poole et al. 1997).

Purified myosin cross bridges (S1) can be attached to a substrate and used to transport actin filaments in vitro in the presence of ATP. A study by Spudich et al (Uyeda et al. 1996) shows that the speed of actin transport in motility assays is proportional to the length of the lever arm. Moreover, the fulcrum appears to lie near the broken helix (gg690-710) which contains the especially reactive thiols (SH1, SH2) of myosin. A similar result has been obtained by Manstein and coworkers using a synthetic lever arm made from repeating α-actinin repeats in place of the light chain binding region (Anson et al. 1996).Mutagenesis studies also indicate the importance of the SH1-SH2 region in controlling movement of the lever arm (Patterson and Spudich 1996). A ggG699A mutation, between the SH1 and SH2 groups, slows myosin transport of actin 100-fold (Kinose et al. 1996).

Specific fluorescent markers attached to the regulatory light chain show a small angular movement on contraction (Allen et al. 1995; Irving et al. 1995), whereas the lever arm hypothesis expects about 50° rotation. However, if only a fraction (ca. 10-20%) of the cross bridges in active muscle take part in contraction at any one time, the magnitude of this apparent rotation can be proportionally scaled up towards the anticipated value.

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