Abstract
It has been shown that skeletal and smooth muscle myosin heads binding to actin results in the movement of smooth muscle tropomyosin, as revealed by a change in fluorescence resonance energy transfer between a fluorescence donor on tropomyosin and an acceptor on actin (Graceffa, P. (1999) Biochemistry 38, 11984-11992). In this work, tropomyosin movement was similarly monitored as a function of unphosphorylated and phosphorylated smooth muscle myosin double-headed fragment smHMM. In the absence of nucleotide and at low myosin head/actin ratios, only phosphorylated heads induced a change in energy transfer. In the presence of ADP, the effect of head phosphorylation was even more dramatic, in that at all levels of myosin head/actin, phosphorylation was necessary to affect energy transfer. It is proposed that the regulation of tropomyosin position on actin by phosphorylation of myosin heads plays a key role in the regulation of smooth muscle contraction. In contrast, actin-bound caldesmon was not moved by myosin heads at low head/actin ratios, as uncovered by fluorescence resonance energy transfer and disulfide cross-linking between caldesmon and actin. At higher head concentration caldesmon was dissociated from actin, consistent with the multiple binding model for the binding of caldesmon and myosin heads to actin (Chen, Y., and Chalovich, J. M. (1992) Biophys. J. 63, 1063-1070).
Highlights
Ca2ϩ to troponin, removes the inhibition of contraction
In our previous work, using fluorescence resonance energy transfer (FRET), we found that, smooth muscle tropomyosin moved upon the binding to actin of single-headed and double-headed fragments of skeletal muscle myosin and the single-headed fragment of smooth muscle myosin [23]
Since smS1 is constitutively active, in the present study we examined the movement of smooth muscle tropomyosin by the double-headed fragment of smooth muscle myosin whose activity is regulated by phosphorylation
Summary
Ca2ϩ to troponin, removes the inhibition of contraction (reviewed in Ref. 1). Interaction of each tropomyosin molecule with seven actin subunits and end-to-end tropomyosin interaction on the thin filament imparts cooperativity to the switching process. It has been shown that skeletal and smooth muscle myosin heads binding to actin results in the movement of smooth muscle tropomyosin, as revealed by a change in fluorescence resonance energy transfer between a fluorescence donor on tropomyosin and an acceptor on actin
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