Effects of tropomyosin internal deletions on thin filament function.
Striated muscle tropomyosin spans seven actin monomers and contains seven quasi-repeating regions with loose sequence similarity. Each region contains a hypothesized actin binding motif. To examine the functions of these regions, full-length tropomyosin was compared with tropomyosin internal deletion mutants spanning either five or four actins. Actin-troponin-tropomyosin filaments lacking tropomyosin regions 2-3 exhibited calcium-sensitive regulation in in vitro motility and myosin S1 ATP hydrolysis experiments, similar to filaments with full-length tropomyosin. In contrast, filaments lacking tropomyosin regions 3-4 were inhibitory to these myosin functions. Deletion of regions 2-4, 3-5, or 4-6 had little effect on tropomyosin binding to actin in the presence of troponin or troponin-Ca(2+), or in the absence of troponin. However, all of these mutants inhibited myosin cycling. Deletion of the quasi-repeating regions diminished the prominent effect of myosin S1 on tropomyosin-actin binding. Interruption of this cooperative, myosin-tropomyosin interaction was least severe for the mutant lacking regions 2-3 and therefore correlated with inhibition of myosin cycling. Regions 3, 4, and 5 each contributed about 1.5 kcal/mol to this process, whereas regions 2 and 6 contributed much less. We suggest that a myosin-induced conformational change in actin facilitates the azimuthal repositioning of tropomyosin which is an essential part of regulation.
Landis
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Department of Internal Medicine, University of Iowa, Iowa City, Iowa 52242, USA.
Back
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Homsher
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Tobacman
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D000199:Actins / Q000378:metabolism; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D002118:Calcium; D009068:Movement; D018482:Muscle, Skeletal / Q000502:physiology*; D009210:Myofibrils / Q000502:physiology*; D015879:Myosin Subfragments / Q000378:metabolism; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017384:Sequence Deletion; D013816:Thermodynamics; D014335:Tropomyosin / Q000235:genetics / Q000378:metabolism*; D014336:Troponin / Q000378:metabolism
D000199:Actins; D015879:Myosin Subfragments; D010446:Peptide Fragments; D014335:Tropomyosin; D014336:Troponin; D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium