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A universal pathway for kinesin stepping

Nature Structural & Molecular Biology volume 18, pages 1020–1027 (2011)Cite this article

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Abstract

Kinesin-1 is an ATP-driven, processive motor that transports cargo along microtubules in a tightly regulated stepping cycle. Efficient gating mechanisms ensure that the sequence of kinetic events proceeds in the proper order, generating a large number of successive reaction cycles. To study gating, we created two mutant constructs with extended neck-linkers and measured their properties using single-molecule optical trapping and ensemble fluorescence techniques. Owing to a reduction in the inter-head tension, the constructs access an otherwise rarely populated conformational state in which both motor heads remain bound to the microtubule. ATP-dependent, processive backstepping and futile hydrolysis were observed under moderate hindering loads. On the basis of measurements, we formulated a comprehensive model for kinesin motion that incorporates reaction pathways for both forward and backward stepping. In addition to inter-head tension, we found that neck-linker orientation is also responsible for ensuring gating in kinesin.

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Figure 1: Single-molecule records and backstepping velocity for Kin6AA.
Figure 2: Bidirectionality of Kin6AA as a function of load and ATP concentration.
Figure 3: Model for stepping by kinesin dimers, showing forward-stepping, backward-stepping and futile-hydrolysis pathways.
Figure 4: Fluorescence data for Kin6AA and KinWT with TMR probes attached to both neck-linkers.
Figure 5: Binding of 2′dmT to Kin6AA.

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Acknowledgements

S.M.B. acknowledges support from grant GM51453, and S.S.R. acknowledges support from grant AR048565 from the US National Institutes of Health.

Author information

Authors and Affiliations

  1. Department of Biology, Stanford University, Stanford, California, USA

    Bason E Clancy & Steven M Block

  2. Department of Biology, Columbia University, New York, New York, USA

    William M Behnke-Parks

  3. Department of Physics, Stanford University, Stanford, California, USA

    Johan O L Andreasson

  4. Department of Neurology, Columbia University, New York, New York, USA

    Steven S Rosenfeld

  5. Department of Applied Physics, Stanford University, Stanford, California, USA

    Steven M Block

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  1. Bason E Clancy
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Contributions

B.E.C., W.M.B.-P. and J.O.L.A. designed and carried out experiments, collected and analyzed data, and cowrote the paper. B.E.C. and J.O.L.A. carried out modeling and fits. S.M.B. and S.S.R. helped to design experiments and analyze results, and cowrote the paper.

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Correspondence to Steven M Block.

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Clancy, B., Behnke-Parks, W., Andreasson, J. et al. A universal pathway for kinesin stepping. Nat Struct Mol Biol 18, 1020–1027 (2011). https://doi.org/10.1038/nsmb.2104

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