Using Optical Tweezers to Study the Fine Details of Myosin ATPase Mechanochemical Cycle

  • Christopher BattersEmail author
  • Claudia Veigel
Part of the Methods in Molecular Biology book series (MIMB, volume 778)


Optical tweezers offer the capability to directly observe nanometre displacements and apply piconewton forces to single proteins. This method has been applied to the study of many different biological systems. Optical tweezers have proven to be particularly useful in studying the fine details of the mechanisms of molecular motor proteins, and how their movement is coordinated with ATPase activity. This includes actin, microtubule, and also DNA- and RNA-based motor systems. Here, we provide the information necessary to reproduce the “three-bead geometry” widely applied to the study of actomyosin interactions, the “paradigm system” for motors that only interact intermittently with their filament substrate, and discuss how single-molecule interactions can be detected, calibrated and analysed.

Key words

Myosin Actin Optical tweezers 


  1. 1.
    Ashkin, A., Dziedzic, J. M., Bjorkholm, J. E., and Chu, S. (1986) Observation of a single-beam gradient force optical trap for dielectric particles. Opt. Lett. 11, 288–290.PubMedCrossRefGoogle Scholar
  2. 2.
    Ashkin, A. (1992) Forces of a single-beam gradient laser trap on a dielectric sphere in the ray optics regime. Biophys. J. 61, 569–582.PubMedCrossRefGoogle Scholar
  3. 3.
    Ashkin, A. (1997) Optical trapping and manipulation of neutral particles using lasers. Proc. Natl. Acad. Sci. USA 94, 4853–4860.PubMedCrossRefGoogle Scholar
  4. 4.
    Veigel, C., and Schmidt, C. F. (2011) Moving into the cell: Single molecule studies of molecular motors in complex environments. Nature Review Mol. Cell Biol. 12(3), 163–176.PubMedCrossRefGoogle Scholar
  5. 5.
    Veigel, C., Coluccio, L. M., Jontes, J. D., Sparrow, J. C., Milligan, R. A., and Molloy, J. E. (1999) The motor protein myosin-I produces its working stroke in two steps. Nature 398, 530–533.PubMedCrossRefGoogle Scholar
  6. 6.
    Veigel, C., Wang, F., Bartoo, M. L., Sellers, J. R., and Molloy, J. E. (2002) The gated gait of the processive molecular motor, myosin V. Nat. Cell Biol. 4, 59–65.PubMedCrossRefGoogle Scholar
  7. 7.
    Veigel, C., Molloy, J. E., Schmitz, S., and Kendrick-Jones, J. (2003) Load-dependent kinetics of force production by smooth muscle myosin measured with optical tweezers. Nat. Cell Biol. 5, 980–986.PubMedCrossRefGoogle Scholar
  8. 8.
    Veigel, C., Schmitz, S., Wang, F., and Sellers, J. R. (2005) Load-dependent kinetics of myosin-V can explain its high processivity. Nat. Cell Biol. 7, 861–869.PubMedCrossRefGoogle Scholar
  9. 9.
    Sellers, J. R., and Veigel, C. (2010) Direct observation of the myosin-Va power stroke and its reversal. Nat. Struct. Mol. Biol. 17, 590–595.PubMedCrossRefGoogle Scholar
  10. 10.
    Ghislain, L. P., and Webb, W. W. (1993) Scanning force microscope based on an optical trap. Opt. Lett. 18, 1678–1680.PubMedCrossRefGoogle Scholar
  11. 11.
    Finer, J. T., Simmons, R. M., and Spudich, J. A. (1994) Single myosin molecule mechanics-piconewton forces and nanometer steps. Nature 368, 113–119.PubMedCrossRefGoogle Scholar
  12. 12.
    Visscher, K., Gross, S. P., and Block, S. M. (1996) Construction of multiple-beam optical traps with nanometer-resolution position sensing. IEEE J. Sel. Top. Quantum Electron. 2, 1066–1076.CrossRefGoogle Scholar
  13. 13.
    Veigel, C., Bartoo, M. L., White, D. C. S., Sparrow, J. C., and Molloy, J. E. (1998) The stiffness of rabbit skeletal actomyosin cross-bridges determined with an optical tweezers transducer. Biophys. J. 75, 1424–1438.PubMedCrossRefGoogle Scholar
  14. 14.
    Pardee, J. D., and Spudich, J. A. (1982) Purification of muscle actin. Methods Enzymol. 85, 164–181.Google Scholar
  15. 15.
    Kron, S. J., and Spudich, J. A. (1986) Fluorescent actin-filaments move on myosin fixed to a glass surface. Proc. Natl. Acad. Sci. USA 83, 6272–6276.PubMedCrossRefGoogle Scholar
  16. 16.
    Kishino, A., and Yanagida, T. (1988) Force measurements by micromanioulation of a single actin filament by glass needles. Nature 334, 74–76.PubMedCrossRefGoogle Scholar
  17. 17.
    Kron, S. J., Toyoshima, Y. Y., Uyeda, T. Q. P., and Spudich, J. A. (1991) Assays for actin sliding movement over myosin-coated surfaces. Methods Enzymol. 196, 399–416.PubMedCrossRefGoogle Scholar
  18. 18.
    Svoboda, K., and Block, S. M. (1994) Biological applications of optical forces. Annu. Rev. Biophys. Biomol. Struct. 23, 247–285.PubMedCrossRefGoogle Scholar
  19. 19.
    Molloy, J. E., and Padgett, M. J. (2002) Lights, action: optical tweezers. Contemp. Phys. 43, 241–258.CrossRefGoogle Scholar
  20. 20.
    Molloy, J. E., Burns, J. E., Kendrick-Jones, J., Tregear, R. T., and White, D. C. S. (1995) Movement and force produced by a single myosin head. Nature 378, 209–212.PubMedCrossRefGoogle Scholar
  21. 21.
    Steffen, W., Smith, D., Simmons, R., and Sleep, J. (2001) Mapping the actin filament with myosin. Proc. Natl. Acad. Sci. USA 98, 14949–14954.PubMedCrossRefGoogle Scholar
  22. 22.
    Cremo, C. R., and Geeves, M. A. (1998) Interaction of actin and ADP with the head domain of smooth muscle myosin: Implications for strain-dependent ADP release in smooth muscle. Biochemistry 37, 1969–1978.PubMedCrossRefGoogle Scholar
  23. 23.
    Wells, J. A., and Yount, R. G. (1982) Chemical modification of myosin by active-site trapping of metal-nucleotides with thiol crosslinking reagents. Methods Enzymol. 85, 93–116.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Physiologisches Institute, Zelluläre PhysiologieLudwig-Maximilians-Universität MünchenMunichGermany

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