Reduction of charging in protein electron cryomicroscopy |
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Authors: | Brink Gross Tittmann Sherman & Chiu |
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Affiliation: | National Center for Macromolecular Imaging, Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, U.S.A.,;Institute of Cell Biology, ETH-Hönggerberg, 8093 Zürich, Switzerland |
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Abstract: | Charging causes a loss of resolution in electron cryomicroscopy with biological specimens prepared without a continuous carbon support film. Thin conductive films were deposited onto catalase crystals prepared across holes using ion-beam sputtering and thermal evaporation and evaluated for the effectiveness of charge reduction. Deposits applied by ion-beam sputtering reduced charging but concurrently resulted in structural damage. Coatings applied by thermal evaporation also reduced charging, and preserved the specimen structure beyond 5 Å resolution as judged from electron diffraction patterns and images of glucose-embedded catalase crystals tilted to 45° in the microscope. This study demonstrates for the first time the feasibility of obtaining high-resolution data from unstained, unsupported protein crystals with a conductive surface coating. |
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Keywords: | Charging electron diffraction pattern electron gun-equipped evaporator high resolution ion beam coater spot-scan imaging surface deposit |
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