Abstract
The purpose of this paper is to deduce whether the maximum force, steplike movement, and rate of ATP consumption of kinesin, as measured in buffer, are sufficient for the task of fast transport of vesicles in cells. Our results show that moving a 200-nm vesicle in viscoelastic COS7 cytoplasm, with the same steps as observed for kinesin-driven beads in buffer, required a maximum force of 16 pN and work per step of 1 +/- 0.7 ATP, if the drag force was assumed to decrease to zero between steps. In buffer, kinesin can develop a force of 6-7 pN while consuming 1 ATP/step, comparable to the required values. As an alternative to assuming that the force vanishes between steps, the measured COS7 viscoelasticity was extrapolated to zero frequency by a numerical fit. The force required to move the bead then exceeded 75 pN at all times and peaked briefly to 92 pN, well beyond the measured capabilities of a single kinesin in buffer. The work per step increased to 7 +/- 5 ATP, greatly exceeding the energy available to a single motor.
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- Alberty R. A., Goldberg R. N. Standard thermodynamic formation properties for the adenosine 5'-triphosphate series. Biochemistry. 1992 Nov 3;31(43):10610–10615. doi: 10.1021/bi00158a025. [DOI] [PubMed] [Google Scholar]
- Bausch A. R., Möller W., Sackmann E. Measurement of local viscoelasticity and forces in living cells by magnetic tweezers. Biophys J. 1999 Jan;76(1 Pt 1):573–579. doi: 10.1016/S0006-3495(99)77225-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Coppin C. M., Finer J. T., Spudich J. A., Vale R. D. Detection of sub-8-nm movements of kinesin by high-resolution optical-trap microscopy. Proc Natl Acad Sci U S A. 1996 Mar 5;93(5):1913–1917. doi: 10.1073/pnas.93.5.1913. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Coppin C. M., Pierce D. W., Hsu L., Vale R. D. The load dependence of kinesin's mechanical cycle. Proc Natl Acad Sci U S A. 1997 Aug 5;94(16):8539–8544. doi: 10.1073/pnas.94.16.8539. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Coy D. L., Wagenbach M., Howard J. Kinesin takes one 8-nm step for each ATP that it hydrolyzes. J Biol Chem. 1999 Feb 5;274(6):3667–3671. doi: 10.1074/jbc.274.6.3667. [DOI] [PubMed] [Google Scholar]
- Crevel I., Carter N., Schliwa M., Cross R. Coupled chemical and mechanical reaction steps in a processive Neurospora kinesin. EMBO J. 1999 Nov 1;18(21):5863–5872. doi: 10.1093/emboj/18.21.5863. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Evans E. Probing the relation between force--lifetime--and chemistry in single molecular bonds. Annu Rev Biophys Biomol Struct. 2001;30:105–128. doi: 10.1146/annurev.biophys.30.1.105. [DOI] [PubMed] [Google Scholar]
- Hunt A. J., Gittes F., Howard J. The force exerted by a single kinesin molecule against a viscous load. Biophys J. 1994 Aug;67(2):766–781. doi: 10.1016/S0006-3495(94)80537-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaether C., Skehel P., Dotti C. G. Axonal membrane proteins are transported in distinct carriers: a two-color video microscopy study in cultured hippocampal neurons. Mol Biol Cell. 2000 Apr;11(4):1213–1224. doi: 10.1091/mbc.11.4.1213. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kojima H., Muto E., Higuchi H., Yanagida T. Mechanics of single kinesin molecules measured by optical trapping nanometry. Biophys J. 1997 Oct;73(4):2012–2022. doi: 10.1016/S0006-3495(97)78231-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Meyhöfer E., Howard J. The force generated by a single kinesin molecule against an elastic load. Proc Natl Acad Sci U S A. 1995 Jan 17;92(2):574–578. doi: 10.1073/pnas.92.2.574. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nishiyama M., Muto E., Inoue Y., Yanagida T., Higuchi H. Substeps within the 8-nm step of the ATPase cycle of single kinesin molecules. Nat Cell Biol. 2001 Apr;3(4):425–428. doi: 10.1038/35070116. [DOI] [PubMed] [Google Scholar]
- Sato M., Wong T. Z., Brown D. T., Allen R. D. Rheological properties of living cytoplasm: a preliminary investigation of squid axoplasm (Loligo pealei). Cell Motil. 1984;4(1):7–23. doi: 10.1002/cm.970040103. [DOI] [PubMed] [Google Scholar]
- Schnitzer M. J., Block S. M. Kinesin hydrolyses one ATP per 8-nm step. Nature. 1997 Jul 24;388(6640):386–390. doi: 10.1038/41111. [DOI] [PubMed] [Google Scholar]
- Schnitzer M. J., Visscher K., Block S. M. Force production by single kinesin motors. Nat Cell Biol. 2000 Oct;2(10):718–723. doi: 10.1038/35036345. [DOI] [PubMed] [Google Scholar]
- Svoboda K., Block S. M. Force and velocity measured for single kinesin molecules. Cell. 1994 Jun 3;77(5):773–784. doi: 10.1016/0092-8674(94)90060-4. [DOI] [PubMed] [Google Scholar]
- Svoboda K., Schmidt C. F., Schnapp B. J., Block S. M. Direct observation of kinesin stepping by optical trapping interferometry. Nature. 1993 Oct 21;365(6448):721–727. doi: 10.1038/365721a0. [DOI] [PubMed] [Google Scholar]
- Valberg P. A., Feldman H. A. Magnetic particle motions within living cells. Measurement of cytoplasmic viscosity and motile activity. Biophys J. 1987 Oct;52(4):551–561. doi: 10.1016/S0006-3495(87)83244-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Visscher K., Schnitzer M. J., Block S. M. Single kinesin molecules studied with a molecular force clamp. Nature. 1999 Jul 8;400(6740):184–189. doi: 10.1038/22146. [DOI] [PubMed] [Google Scholar]
- Yamada S., Wirtz D., Kuo S. C. Mechanics of living cells measured by laser tracking microrheology. Biophys J. 2000 Apr;78(4):1736–1747. doi: 10.1016/S0006-3495(00)76725-7. [DOI] [PMC free article] [PubMed] [Google Scholar]