Kinesin Takes One 8-nm Step for Each ATP That It Hydrolyzes*
- From the Department of Physiology and Biophysics, University of Washington, Seattle, Washington 98195-7290
Abstract
Conventional kinesin is a motor protein that moves stepwise along microtubules carrying membrane-bound organelles toward the periphery of cells. The steps are of amplitude 8.1 nm, the distance between adjacent tubulin binding sites, and are powered by the hydrolysis of ATP. We have asked: how many steps does kinesin take for each molecule of ATP that it hydrolyzes? To answer this question, the motility and ATP hydrolysis of recombinant, heterotetrameric and homodimeric conventional Drosophila kinesins adsorbed to 200-nm-diameter casein-coated silica beads were assayed under identical, single-molecule conditions. Division of the speed by the maximum microtubule-activated ATPase rate gave a stoichiometry of 1.08 ± 0.09 steps for each ATP hydrolyzed at 1 mmATP. Therefore, under low loads in which the drag force ≪ 1 pN, coupling between the chemical and mechanical cycles of kinesin is tight, consistent with conventional power stroke models. Our results rule out models that require two or more ATPs/step, such as some thermal ratchet models, or that propose multiple steps powered by single ATPs.
Footnotes
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↵* This work was supported by National Institutes of Health Grant AR40593 and by a grant from the Human Frontier Science Program (to J. H.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) AF053733 and AF055298.
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↵‡ Supported by National Institutes of Health Molecular Biophysics Training Grant GM08268 and by the Achievement Reward for College Scientists Foundation.
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↵§ To whom correspondence should be addressed: Dept. of Physiology & Biophysics, University of Washington, Box 357290, Seattle, WA 98195-7290. Fax: 206-685-0619; E-mail:johoward{at}u.washington.edu.
- Abbreviations:
- AMP-PNP
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adenosine 5′-(β,γ-imino)triphosphate
- GMP-CPP
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guanylyl-(α,β)-methylene-diphosphonate
- PIPES
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1,4-piperazinediethanesulfonic acid.
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- Received August 27, 1998.
- Revision received November 5, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
