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. 1994 Aug;67(2):766–781. doi: 10.1016/S0006-3495(94)80537-5

The force exerted by a single kinesin molecule against a viscous load.

A J Hunt 1, F Gittes 1, J Howard 1
PMCID: PMC1225420  PMID: 7948690

Abstract

Kinesin is a motor protein that uses the energy derived from the hydrolysis of ATP to power the transport of organelles along microtubules. To probe the mechanism of this chemical-to-mechanical energy transduction reaction, the movement of microtubules across glass surfaces coated with kinesin was perturbed by raising the viscosity of the buffer solution. When the viscosity of the solution used in the low density motility assay was increased approximately 100-fold through addition of polysaccharides and polypeptides, the longer microtubules, which experienced a larger drag force from the fluid, moved more slowly than the shorter ones. The speed of movement of a microtubule depended linearly on the drag force loading the motor. At the lowest kinesin density, where dilution experiments indicated that the movement was caused by a single kinesin molecule, extrapolation of the linear relationship yielded a maximum time-averaged drag force of 4.2 +/- 0.5 pN per motor (mean +/- experimental SE). The magnitude of the force argues against one type of "ratchet" model in which the motor is hypothesized to rectify the diffusion of the microtubule; at high viscosity, diffusion is too slow to account for the observed speeds. On the other hand, our data are consistent with models in which force is a consequence of strain developed in an elastic element within the motor; these models include a different "ratchet" model (of the type proposed by A. F. Huxley in 1957) as well as "power-stroke" models.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allen R. D., Weiss D. G., Hayden J. H., Brown D. T., Fujiwake H., Simpson M. Gliding movement of and bidirectional transport along single native microtubules from squid axoplasm: evidence for an active role of microtubules in cytoplasmic transport. J Cell Biol. 1985 May;100(5):1736–1752. doi: 10.1083/jcb.100.5.1736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Amos L., Klug A. Arrangement of subunits in flagellar microtubules. J Cell Sci. 1974 May;14(3):523–549. doi: 10.1242/jcs.14.3.523. [DOI] [PubMed] [Google Scholar]
  3. Beese L., Stubbs G., Cohen C. Microtubule structure at 18 A resolution. J Mol Biol. 1987 Mar 20;194(2):257–264. doi: 10.1016/0022-2836(87)90373-1. [DOI] [PubMed] [Google Scholar]
  4. Block S. M., Goldstein L. S., Schnapp B. J. Bead movement by single kinesin molecules studied with optical tweezers. Nature. 1990 Nov 22;348(6299):348–352. doi: 10.1038/348348a0. [DOI] [PubMed] [Google Scholar]
  5. Brady S. T. A novel brain ATPase with properties expected for the fast axonal transport motor. Nature. 1985 Sep 5;317(6032):73–75. doi: 10.1038/317073a0. [DOI] [PubMed] [Google Scholar]
  6. Córdova N. J., Ermentrout B., Oster G. F. Dynamics of single-motor molecules: the thermal ratchet model. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):339–343. doi: 10.1073/pnas.89.1.339. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Fenn W. O. The relation between the work performed and the energy liberated in muscular contraction. J Physiol. 1924 May 23;58(6):373–395. doi: 10.1113/jphysiol.1924.sp002141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Finer J. T., Simmons R. M., Spudich J. A. Single myosin molecule mechanics: piconewton forces and nanometre steps. Nature. 1994 Mar 10;368(6467):113–119. doi: 10.1038/368113a0. [DOI] [PubMed] [Google Scholar]
  9. Gilbert S. P., Johnson K. A. Expression, purification, and characterization of the Drosophila kinesin motor domain produced in Escherichia coli. Biochemistry. 1993 May 4;32(17):4677–4684. doi: 10.1021/bi00068a028. [DOI] [PubMed] [Google Scholar]
  10. Gittes F., Mickey B., Nettleton J., Howard J. Flexural rigidity of microtubules and actin filaments measured from thermal fluctuations in shape. J Cell Biol. 1993 Feb;120(4):923–934. doi: 10.1083/jcb.120.4.923. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. HUXLEY A. F. Muscle structure and theories of contraction. Prog Biophys Biophys Chem. 1957;7:255–318. [PubMed] [Google Scholar]
  12. Hackney D. D., Malik A. S., Wright K. W. Nucleotide-free kinesin hydrolyzes ATP with burst kinetics. J Biol Chem. 1989 Sep 25;264(27):15943–15948. [PubMed] [Google Scholar]
  13. Hall K., Cole D. G., Yeh Y., Scholey J. M., Baskin R. J. Force-velocity relationships in kinesin-driven motility. Nature. 1993 Jul 29;364(6436):457–459. doi: 10.1038/364457a0. [DOI] [PubMed] [Google Scholar]
  14. Harrison B. C., Marchese-Ragona S. P., Gilbert S. P., Cheng N., Steven A. C., Johnson K. A. Decoration of the microtubule surface by one kinesin head per tubulin heterodimer. Nature. 1993 Mar 4;362(6415):73–75. doi: 10.1038/362073a0. [DOI] [PubMed] [Google Scholar]
  15. Howard J., Hudspeth A. J., Vale R. D. Movement of microtubules by single kinesin molecules. Nature. 1989 Nov 9;342(6246):154–158. doi: 10.1038/342154a0. [DOI] [PubMed] [Google Scholar]
  16. Howard J., Hunt A. J., Baek S. Assay of microtubule movement driven by single kinesin molecules. Methods Cell Biol. 1993;39:137–147. doi: 10.1016/s0091-679x(08)60167-3. [DOI] [PubMed] [Google Scholar]
  17. Hunt A. J., Howard J. Kinesin swivels to permit microtubule movement in any direction. Proc Natl Acad Sci U S A. 1993 Dec 15;90(24):11653–11657. doi: 10.1073/pnas.90.24.11653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Huxley A. F. Muscular contraction. J Physiol. 1974 Nov;243(1):1–43. [PMC free article] [PubMed] [Google Scholar]
  19. Hyman A., Drechsel D., Kellogg D., Salser S., Sawin K., Steffen P., Wordeman L., Mitchison T. Preparation of modified tubulins. Methods Enzymol. 1991;196:478–485. doi: 10.1016/0076-6879(91)96041-o. [DOI] [PubMed] [Google Scholar]
  20. Ishijima A., Harada Y., Kojima H., Funatsu T., Higuchi H., Yanagida T. Single-molecule analysis of the actomyosin motor using nano-manipulation. Biochem Biophys Res Commun. 1994 Mar 15;199(2):1057–1063. doi: 10.1006/bbrc.1994.1336. [DOI] [PubMed] [Google Scholar]
  21. Kron S. J., Spudich J. A. Fluorescent actin filaments move on myosin fixed to a glass surface. Proc Natl Acad Sci U S A. 1986 Sep;83(17):6272–6276. doi: 10.1073/pnas.83.17.6272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Leibler S., Huse D. A. Porters versus rowers: a unified stochastic model of motor proteins. J Cell Biol. 1993 Jun;121(6):1357–1368. doi: 10.1083/jcb.121.6.1357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Miller R. H., Lasek R. J. Cross-bridges mediate anterograde and retrograde vesicle transport along microtubules in squid axoplasm. J Cell Biol. 1985 Dec;101(6):2181–2193. doi: 10.1083/jcb.101.6.2181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Oiwa K., Chaen S., Kamitsubo E., Shimmen T., Sugi H. Steady-state force-velocity relation in the ATP-dependent sliding movement of myosin-coated beads on actin cables in vitro studied with a centrifuge microscope. Proc Natl Acad Sci U S A. 1990 Oct;87(20):7893–7897. doi: 10.1073/pnas.87.20.7893. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pate E., Cooke R. Simulation of stochastic processes in motile crossbridge systems. J Muscle Res Cell Motil. 1991 Aug;12(4):376–393. doi: 10.1007/BF01738593. [DOI] [PubMed] [Google Scholar]
  26. Ray S., Meyhöfer E., Milligan R. A., Howard J. Kinesin follows the microtubule's protofilament axis. J Cell Biol. 1993 Jun;121(5):1083–1093. doi: 10.1083/jcb.121.5.1083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Rayment I., Rypniewski W. R., Schmidt-Bäse K., Smith R., Tomchick D. R., Benning M. M., Winkelmann D. A., Wesenberg G., Holden H. M. Three-dimensional structure of myosin subfragment-1: a molecular motor. Science. 1993 Jul 2;261(5117):50–58. doi: 10.1126/science.8316857. [DOI] [PubMed] [Google Scholar]
  28. Romberg L., Vale R. D. Chemomechanical cycle of kinesin differs from that of myosin. Nature. 1993 Jan 14;361(6408):168–170. doi: 10.1038/361168a0. [DOI] [PubMed] [Google Scholar]
  29. Scholey J. M., Heuser J., Yang J. T., Goldstein L. S. Identification of globular mechanochemical heads of kinesin. Nature. 1989 Mar 23;338(6213):355–357. doi: 10.1038/338355a0. [DOI] [PubMed] [Google Scholar]
  30. Sheetz M. P., Block S. M., Spudich J. A. Myosin movement in vitro: a quantitative assay using oriented actin cables from Nitella. Methods Enzymol. 1986;134:531–544. doi: 10.1016/0076-6879(86)34118-1. [DOI] [PubMed] [Google Scholar]
  31. Sheetz M. P., Spudich J. A. Movement of myosin-coated fluorescent beads on actin cables in vitro. Nature. 1983 May 5;303(5912):31–35. doi: 10.1038/303031a0. [DOI] [PubMed] [Google Scholar]
  32. Sweet R. M., Wright H. T., Janin J., Chothia C. H., Blow D. M. Crystal structure of the complex of porcine trypsin with soybean trypsin inhibitor (Kunitz) at 2.6-A resolution. Biochemistry. 1974 Sep 24;13(20):4212–4228. doi: 10.1021/bi00717a024. [DOI] [PubMed] [Google Scholar]
  33. Vale R. D., Oosawa F. Protein motors and Maxwell's demons: does mechanochemical transduction involve a thermal ratchet? Adv Biophys. 1990;26:97–134. doi: 10.1016/0065-227x(90)90009-i. [DOI] [PubMed] [Google Scholar]

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