Kinesin's step dissected with single-motor FRET
- Department of Physics and Astronomy and Laser Centre, VU University, de Boelelaan 1081, 1081 HV, Amsterdam, The Netherlands
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Edited by Steven M. Block, Stanford University, Stanford, CA, and approved August 31, 2009
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↵1S.V. and Z.L. contributed equally to this work. (received for review May 15, 2009)
Abstract
The motor protein Kinesin-1 drives intracellular transport along microtubules, with each of its two motor domains taking 16-nm steps in a hand-over-hand fashion. The way in which a single-motor domain moves during a step is unknown. Here, we use Förster resonance energy transfer (FRET) between fluorescent labels on both motor domains of a single kinesin. This approach allows us to resolve the relative distance between the motor domains and their relative orientation, on the submillisecond timescale, during processive stepping. We observe transitions between high and low FRET values for certain kinesin constructs, depending on the location of the labels. These results reveal that, during a step, a kinesin motor domain dwells in a well-defined intermediate position for ≈3 ms.
Footnotes
- 2To whom correspondence should be addressed. E-mail: erwinp{at}nat.vu.nl
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Author contributions: E.J.G.P. designed research; S.V. and Z.L. performed research; S.V. and Z.L. analyzed data; and S.V., Z.L., and E.J.G.P. wrote the paper.
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The authors declare no conflict of interest.
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This article is a PNAS Direct Submission.
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This article contains supporting information online at www.pnas.org/cgi/content/full/0905177106/DCSupplemental.
