Conformational Changes in Reconstituted Skeletal Muscle Thin Filaments Observed by Fluorescence Spectroscopy

  • Masao Miki
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 592)


The cyclic interaction of myosin and actin coupled ATP hydrolysis generates the mechanical force of muscle contraction. During this process, the system passes through several steps. One of these is thought to be identical to the stable rigor complex formed by myosin and actin in the absence of ATP. This cyclic interaction is regulated by changes in tropomyosin (Tm) and troponin (Tn) located on the actin filament in response alterations in intracellular Ca2+ concentration (Ebashi et al., 1969). Tm contains seven quasi-equivalent regions, each of which has a pair of putative actin-binding motifs. Tn comprises three different subunits, TnC, TnI, and TnT. TnI alone inhibits actomyosin ATPase activity which is removed on adding TnC, irrespective of Ca2+ concentration. TnT is required for full Ca2+-regulation of the ATPase activity of a reconstituted system (Ohtsuki et al., 1986). The globular part of the Tn complex (TnC, TnI and the C-terminal region of TnT) is located on residues 150–180 of Tm (White et al., 1987), and the elongated part, composed of the N-terminal region, covers an extensive region of the C-terminal half of Tm. The binding of Ca2+ to TnC induces a series of conformational changes in the other components of the thin filament. This allows the effective association of myosin with actin, thus producing force. Although numerous studies have characterized the interaction between these thin filament proteins, the molecular mechanism whereby the Ca2+-trigger is propagated from TnC to the rest of the thin filament is still not well understood.


Fluorescence Resonance Energy Transfer Thin Filament Actomyosin ATPase Donor Probe Myosin Subfragment 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

11.6. References

  1. Bacchiocchi, C., and Lehrer, S. S., 2001, Ca2+-induced movement of tropomyosin in skeletal muscle thin filaments observed by multi-site FRET. Biophys. J. 82:1524–1536.Google Scholar
  2. Borovikov, Y. S., Nowak, E., Khoroshev, M. I., and Dabrowska, R., 1993, The effect of Ca2+ on the conformation of tropomyosin and actin in regulated actin filaments with or without bound myosin subfragment 1. Biochim. Biophys. Acta. 1163:280–286.PubMedGoogle Scholar
  3. Ebashi, S., Endo, M., and Ohtsuki, I., 1969, Control of muscle contraction, Q. Rev. Biophys. 2:351–384.PubMedGoogle Scholar
  4. Geeves, M. A., and Lehrer, S. S., 1994, Dynamics of the muscle thin filament regulatory switch: the size of the cooperative unit. Biophys. J. 67:273–282.PubMedGoogle Scholar
  5. Greene, L. E., Williams, D. L., Jr., and Eisenberg, E., 1987, Regulation of actomyosin ATPase activity by troponin-tropomyosin: Effect of the binding of the myosin subfragment 1 (S-1), ATP complex. Proc. Natl. Acad. Sci. USA 84:3102–3106.PubMedCrossRefGoogle Scholar
  6. Hai, H., Sano, K., Maeda, K., Maéda, Y., and Miki, M., 2002, Ca2+-induced conformational change of reconstituted skeletal muscle thin filaments with an internal deletion mutant d234-tropomyosin observed by fluorescence energy transfer spectroscopy: Structural evidence for three states of thin filament. J. Biochem. 131:407–418.PubMedGoogle Scholar
  7. Haselgrove, J., 1972, X-ray evidence for a conformational change in the actin containing filaments of vertebrate striated muscle. Cold Spring Harb. Symp. Quant. Biol. 37:341–352.Google Scholar
  8. Holmes, K. C., 1995, The actomyosin interaction and its control by tropomyosin. Biophys. J. 68:2s–7s.PubMedGoogle Scholar
  9. Huxley, H. E., 1972, Structural changes in the actin-and myosin-containing filaments during contraction. Cold Spring Harb. Symp. Quant. Biol. 37:361–376.Google Scholar
  10. Kimura, C., Maeda, K., Hai, H., and Miki, M., 2002b, Ca2+-and S1-induced movement of troponin T on mutant thin filaments reconstituted with functionally deficient mutant tropomyosin. J. Biochem. 132:345–352.PubMedGoogle Scholar
  11. Kimura, C., Maeda, K., Maéda, Y., and Miki, M., 2002a, Ca2+-and S1-induced movement of troponin T on reconstituted skeletal muscle thin filaments observed by fluorescence energy transfer spectroscopy. J. Biochem. 132:93–102.PubMedGoogle Scholar
  12. Kobayashi, T., Kobayashi, M., and Collins, J. H., 2001, Ca2+-dependent, myosin subfragment 1-induced proximity changes between actin and the inhibitory region of troponin I. Biochim. Biophys. Acta. 1549:148–154.PubMedGoogle Scholar
  13. Kress, M., Huxley, H. E., Faruqi, A. R., and Hendrix, J., 1986, Structural changes during activation of frog muscle studied by time-resolved x-ray diffraction, J. Mol. Biol. 188:325–342.PubMedCrossRefGoogle Scholar
  14. Landis, C. A., Bobkova, A., Homsher, E., and Tobacman, L. S., 1997, The active state of the thin filament is destabilized by an internal deletion in tropomyosin. J. Biol. Chem. 272:14051–14056.PubMedCrossRefGoogle Scholar
  15. Lin, T-I., and Dowben, R. M., 1983, Studies on the spatial arrangement of muscle thin filament proteins using fluorescence energy transfer. J. Biol. Chem. 258:5142–5150.PubMedGoogle Scholar
  16. Lorenz, M., Popp, D., and Holmes, K. C., 1993, Refinement of the F-actin model against x-ray fiber diffraction data by the use of a directed mutation algorithm. J. Mol. Biol. 234:826–836.PubMedCrossRefGoogle Scholar
  17. McKillop, D. F., and Geeves, M. A., 1993, Regulation of the interaction between actin and myosin subfragment 1: evidence for three states of the thin filament. Biophys. J. 65:693–701.PubMedGoogle Scholar
  18. Maytum, R., Geeves, M. A., and Lehrer, S. S., 2002, A modulatory role for the troponin T tail domain in thin filament regulation. J. Biol. Chem. 277:29774–29780.PubMedCrossRefGoogle Scholar
  19. Miki, M., 1990, Resonance energy transfer between points in a reconstituted skeletal muscle thin filament: A conformational change of the thin filament in response to a change in Ca2+ concentration. Eur. J. Biochem. 187:155–162.PubMedCrossRefGoogle Scholar
  20. Miki, M., Hai, H., Saeki, K., Shitaka, Y., Sano, K-I., Maéda, Y., and Wakabayashi, T., 2004, Fluorescence energy transfer between points on actin and the C-terminal region of tropomyosin in skeletal muscle thin filaments. J. Biochem. 136:39–47.PubMedCrossRefGoogle Scholar
  21. Miki, M., and Iio, T., 1993, Kinetics of structural changes of reconstituted skeletal muscle thin filaments observed by fluorescence resonance energy transfer. J. Biol. Chem. 268:7101–7106.PubMedGoogle Scholar
  22. Miki, M., Kobayashi, T., Kimura, H., Hagiwara, A., Hai, H., and Maéda, Y., 1998a, Ca2+-induced distance change between points on actin and troponin in skeletal muscle thin filaments estimated by fluorescence resonance energy transfer spectroscopy. J. Biochem. 123:324–331.PubMedGoogle Scholar
  23. Miki, M., and Mihashi, K., 1979, Conformational change of the reconstituted thin filament — fluorescence energy transfer and fluorescence polarization measurements. Seibutsu-Butsuri 19:135–140 (in Japanese).Google Scholar
  24. Miki, M., Miura, T., Sano, K., Kimura, H., Kondo, H., Ishida, H., and Maéda, Y., 1998b, Fluorescence resonance energy transfer between points on tropomyosin and actin in skeletal muscle thin filaments: Does tropomyosin move? J. Biochem. 123:1104–1111.PubMedGoogle Scholar
  25. Monteiro, P. B., Lataro, C., Ferro, J. A., and Reinach, F. C., 1994, Functional a-tropomyosin produced in Escherichia coli. J. Biol. Chem. 269:10461–10466.PubMedGoogle Scholar
  26. Nagashima, H., and Asakura, S., 1982, Studies on co-operative properties of tropomyosin-actin and tropomyosin-troponin-actin complexes by the use of N-ethylmaleimide-treated and untreated species of myosin subfragment 1. J. Mol. Biol. 155:409–428.PubMedCrossRefGoogle Scholar
  27. Narita, A., Yasunaga, T., Ishikawa, T., Mayanagi, K., and Wakabayashi, T., 2001, Ca2+-induced switching of troponin and tropomyosin on actin filaments as revealed by electron cryo-microscopy. J. Mol. Biol. 308:241–261.PubMedCrossRefGoogle Scholar
  28. Ohtsuki, I., Maruyama, K., and Ebashi, S., 1986, Regulatory and cytoskeletal proteins of vertebrate skeletal muscle, Adv. Protein Chem. 38:1–67.PubMedCrossRefGoogle Scholar
  29. Resetar, A. M., Stephens, J. M., and Chalovich, J. M., 2002, Troponin-tropomyosin: An allosteric switch or a steric blocker? Biophys. J. 83:1039–1049.PubMedCrossRefGoogle Scholar
  30. Squire, J. M., and Morris, E. P., 1998, A new look at thin filament regulation in vertebrate skeletal muscle. FASEB J. 12:761–771.PubMedGoogle Scholar
  31. Shitaka, Y., Kimura, C., Iio, T., and Miki, M., 2004, Kinetics of the structural transition of muscle thin filaments observed by fluorescence resonance energy transfer. Biochemistry 46:10739–10747.CrossRefGoogle Scholar
  32. Shitaka, Y., Kimura, C., and Miki, M., 2005, The rates of switching movement of troponin-T between three states of skeletal muscle thin filaments determined by fluorescence resonance energy transfer. J. Biol. Chem. 280:2613–2619.PubMedCrossRefGoogle Scholar
  33. Stryer, L., 1978, Fluorescence energy transfer as a spectroscopic ruler, Annu. Rev. Biochem. 47:819–846.PubMedCrossRefGoogle Scholar
  34. Tao, T., Gong, B. J., and Leavis, P. C., 1990, Calcium-induced movement of troponin-I relative to actin in skeletal muscle thin filaments. Science 247:1339–1341.PubMedCrossRefGoogle Scholar
  35. Tao, T., Lamkin, M., and Lehrer, S. S., 1983, Excitation energy transfer studies of the proximity between tropomyosin and actin in reconstituted skeletal muscle thin filaments. Biochemistry 22:3059–3066.PubMedCrossRefGoogle Scholar
  36. Vibert, P., Craig, R., and Lehman, W., 1997, Steric-model for activation of muscle thin filaments. J. Mol. Biol. 266:8–14.PubMedCrossRefGoogle Scholar
  37. White, S. P., Cohen, C., and Phillips, G. N., Jr, 1987, Structure of co-crystals of tropomyosin and troponin. Nature 325:826–828.PubMedCrossRefGoogle Scholar

Copyright information

© Springer 2007

Authors and Affiliations

  • Masao Miki
    • 1
    • 2
  1. 1.Department of Applied Chemistry and BiotechnologyFukui UniversityFukuiJapan
  2. 2.Research and Education Program for Life ScienceFukui UniversityFukuiJapan

Personalised recommendations