Comparison of slam-freezing and high-pressure freezing effects on the DNA cholesteric liquid crystalline structure |
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Authors: | A. Leforestier,K. Richter,F. Livolant,& J. Dubochet |
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Affiliation: | Laboratoire de Physique des Solides, Bât. 510, UniversitéParis-Sud, F-91405 Orsay, France; Laboratoire d'Analyse Ultrastructurale, Bât. de Biologie, Universitéde Lausanne-Dorigny, CH-1015 Lausanne, Switzerland |
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Abstract: | Using in parallel electron microscopy of ultrathin frozen-hydrated sections and freeze-fracture replicas, we compare the ultrastructural consequences of two freezing techniques: slam-freezing at liquid helium temperature and high-pressure freezing, on a model system, the DNA cholesteric liquid crystalline phase. Both freezing techniques are able to vitrify DNA liquid crystalline solutions containing up to 85% water, but induce structural rearrangements of the molecular organization. The cholesteric structure is preserved by the slam-freezing method despite the formation of periodic distortions induced by the mechanical compressive stress. In contrast, high-pressure freezing does not preserve the structure of the liquid crystal: the long-range cholesteric stratification disappears, and the local continuous twist between molecules is modified. These results show that vitrification, though necessary, may not be a sufficient token of preservation of the native state of hydrated materials. We discuss the possible origins of the molecular rearrangements that have time to occur in the specimens as a result of the low freezing rate permitted by the high-pressure freezing process. |
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Keywords: | Cryoelectron microscopy cryoultramicrotomy DNA liquid crystals freeze-fracture high-pressure freezing slam-freezing. |
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