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Resolving the molecular structure of microtubules under physiological conditions with scanning force microscopy

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Abstract

We have imaged microtubules, essential structural elements of the cytoskeleton in eukaryotic cells, in physiological conditions by scanning force microscopy. We have achieved molecular resolution without the use of cross-linking and chemical fixation methods. With tip forces below 0.3 nN, protofilaments with ~6 nm separation could be clearly distinguished. Lattice defects in the microtubule wall were directly visible, including point defects and protofilament separations. Higher tip forces destroyed the top half of the microtubules, revealing the inner surface of the substrate-attached protofilaments. Monomers could be resolved on these inner surfaces.

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Fig. 1A, B
Fig. 2A, B
Fig. 3
Fig. 4A–C
Fig. 5A–D

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Abbreviations

APTS:

(3-aminopropyl)triethoxysilane

DETA:

N 1-[3-(trimethoxysilyl)propyl]diethylenetriamine

EM:

electron microscopy

MT:

microtubule

SFM:

scanning force microscopy

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Acknowledgements

We thank Ken Downing for sharing his 3D MT model, Per Bullough for help with image analysis, Fred MacKintosh for helpful discussions, Johanna van Nes and René Koops for help with exploratory experiments, and financial support of the Dutch Foundation for Fundamental Research of Matter (FOM) and ALW/FOM project no. 01FB28/2.

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Author notes

  1. Pedro J. de Pablo

    Present address: Departamento de Física de la Materia Condensada C-III, Universidad Autónoma de Madrid, 28049 , Madrid, Spain

Authors and Affiliations

  1. Section Physics of Complex Systems, Department of Physics and Astronomy, Faculty of Sciences, Vrije Universiteit Amsterdam, De Boelelaan 1081, 1081 HV , Amsterdam, The Netherlands

    Iwan A. T. Schaap, Pedro J. de Pablo & Christoph F. Schmidt

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  1. Iwan A. T. Schaap
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  2. Pedro J. de Pablo
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  3. Christoph F. Schmidt
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Correspondence to Iwan A. T. Schaap.

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Schaap, I.A.T., de Pablo, P.J. & Schmidt, C.F. Resolving the molecular structure of microtubules under physiological conditions with scanning force microscopy. Eur Biophys J 33, 462–467 (2004). https://doi.org/10.1007/s00249-003-0386-8

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  • DOI: https://doi.org/10.1007/s00249-003-0386-8

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