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Journal Article

Cryo-electron microscopy of an extremely halophilic microbe: technical aspects

MPS-Authors
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Bollschweiler,  Daniel
Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society;

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Schaffer,  Miroslava
Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society;

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Lawrence,  C. Martin
Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society;

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Engelhardt,  Harald
Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society;

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Fulltext (public)

s00792-016-0912-0.pdf
(Publisher version), 2MB

Supplementary Material (public)

792_2016_912_MOESM1_ESM.pdf
(Supplementary material), 5MB

Citation

Bollschweiler, D., Schaffer, M., Lawrence, C. M., & Engelhardt, H. (2017). Cryo-electron microscopy of an extremely halophilic microbe: technical aspects. Extremophiles, 21(2), 393-398. doi:10.1007/s00792-016-0912-0.


Cite as: http://hdl.handle.net/11858/00-001M-0000-002D-E683-6
Abstract
Most halophilic Archaea of the class Halobacteriaceae depend on the presence of several molar sodium chloride for growth and cell integrity. This poses problems for structural studies, particularly for electron microscopy, where the high salt concentration results in diminished contrast. Since cryo-electron microscopy of intact cells provides new insights into the cellular and molecular organization under close-to-live conditions, we evaluated strategies and conditions to make halophilic microbes available for investigations in situ. Halobacterium salinarum, the test organism for this study, usually grows at 4.3 M NaCl. Adaptation to lower concentrations and subsequent NaCl reduction via dialysis led to still vital cells at 3 M salt. A comprehensive evaluation of vitrification parameters, thinning of frozen cells by focused-ion-beam micromachining, and cryo-electron microscopy revealed that structural studies under high salt conditions are possible in situ.