Direct observation of single kinesin molecules moving along microtubules

Nature. 1996 Apr 4;380(6573):451-3. doi: 10.1038/380451a0.

Abstract

Kinesin is a two-headed motor protein that powers organelle transport along microtubules. Many ATP molecules are hydrolysed by kinesin for each diffusional encounter with the microtubule. Here we report the development of a new assay in which the processive movement of individual fluorescently labelled kinesin molecules along a microtubule can be visualized directly; this observation is achieved by low-background total internal reflection fluorescence microscopy in the absence of attachment of the motor to a cargo (for example, an organelle or bead). The average distance travelled after a binding encounter with a microtubule is 600 nm, which reflects a approximately 1% probability of detachment per mechanical cycle. Surprisingly, processive movement could still be observed at salt concentrations as high as 0.3 M NaCl. Truncated kinesin molecules having only a single motor domain do not show detectable processive movement, which is consistent with a model in which kinesin's two force-generating heads operate by a hand-over-hand mechanism.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Drosophila
  • Escherichia coli
  • Humans
  • Kinesins / genetics
  • Kinesins / metabolism*
  • Microscopy, Fluorescence / methods
  • Microtubules / metabolism*
  • Molecular Sequence Data
  • Osmolar Concentration
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism

Substances

  • Peptide Fragments
  • Recombinant Fusion Proteins
  • Kinesins