Charting and unzipping the surface layer of Corynebacterium glutamicum with the atomic force microscope

Authors

  • Simon Scheuring,

    1. M. E. Müller Institute for Microscopy at the Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
    2. Institut Curie, UMR-CNRS 168 and LRC-CEA 8, 11 rue Pierre et Marie Curie, 75231 Paris Cedex 05, France.
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  • Henning Stahlberg,

    1. M. E. Müller Institute for Microscopy at the Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
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  • Mohamed Chami,

    1. M. E. Müller Institute for Microscopy at the Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
    2. Institut Curie, UMR-CNRS 168 and LRC-CEA 8, 11 rue Pierre et Marie Curie, 75231 Paris Cedex 05, France.
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  • Christine Houssin,

    1. Laboratoire de Biologie Moléculaire des Cornyebactéries, URA 1354 CNRS, Univérsité de Paris-Sud, 91405 Orsay, France.
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  • Jean-Louis Rigaud,

    1. Institut Curie, UMR-CNRS 168 and LRC-CEA 8, 11 rue Pierre et Marie Curie, 75231 Paris Cedex 05, France.
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  • Andreas Engel

    Corresponding author
    1. M. E. Müller Institute for Microscopy at the Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
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Summary

Bacterial surface layers (S-layers) are extracellular protein networks that act as molecular sieves and protect a large variety of archaea and bacteria from hostile environments. Atomic force microscopy (AFM) was used to asses the S-layer of Coryne-bacterium glutamicum formed of PS2 proteins that assemble into hexameric complexes within a hexagonal lattice. Native and trypsin-treated S-layers were studied. Using the AFM stylus as a nanodissector, native arrays that adsorbed to mica as double layers were separated. All surfaces of native and protease-digested S-layers were imaged at better than 1 nm lateral resolution. Difference maps of the topographies of native and proteolysed samples revealed the location of the cleaved C-terminal fragment and the sidedness of the S-layer. Because the corrugation depths determined from images of both sides span the total thickness of the S-layer, a three-dimensional reconstruction of the S-layer could be calculated. Lattice defects visualized at 1 nm resolution revealed the molecular boundaries of PS2 proteins. The combination of AFM imaging and single molecule force spectroscopy allowed the mechanical properties of the Corynebacterium glutamicum S-layer to be examined. The results provide a basis for understanding the amazing stability of this protective bacterial surface coat.

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