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          Institute: MPI für Biochemie     Collection: Structural Biology (W. Baumeister)     Display Documents



ID: 318781.0, MPI für Biochemie / Structural Biology (W. Baumeister)
Automated electron microscope tomography of frozen-hydrated chromatin - the irregular three-dimensional zigzag architecture persists in compact, isolated fibers
Authors:Horowitz, R. A.; Koster, A. J.; Walz, J.; Woodcock, C. L.
Date of Publication (YYYY-MM-DD):1997-12
Title of Journal:Journal of Structural Biology
Volume:120
Issue / Number:3
Start Page:353
End Page:362
Audience:Not Specified
Abstract / Description:The potential of electron microscope tomography as a tool for obtaining three-dimensional (3D) information about large macromolecular assemblies is greatly extended by automation of data collection. With the implementation of automated control of tilting, focusing, and digital image recording described here, tilt series of frozen-hydrated specimens can be collected with the requisite low dose. Long chromatin fibers were prepared in 90 mM monovalent ions to maintain a fully compact conformation, and after vitrification were completely contained within the ice layer. Tilt series of this material were recorded at 5 degrees tilt increments between +60 degrees and -60 degrees, with a cumulative dose of similar to 35 e(-)/Angstrom(2) for the series. This extremely low dose data was successfully aligned, then reconstructed by weighted backprojection. The underlying architecture of the fibers is an irregular 3D zigzag of interconnected nucleosomes, with the linker DNA between successive nucleosomes in a largely extended conformation, The visualization of this structural motif within long, frozen-hydrated chromatin fibers at relatively high salt extends our previous studies on small fragments at low ionic strength and is in agreement with the observation of this architecture in chromatin fibers in situ in sectioned nuclei. (C) 1997 Academic Press. [References: 31]
Free Keywords:Cryoelectron microscopy; 3-dimensional structure; Ultrastructure; Organization; Resolution; Nucleosome; Particle.; Biochemistry & biophysics.
External Publication Status:published
Document Type:Article
Communicated by:Anton Hillebrand
Affiliations:MPI für Biochemie/Structural Biology (W. Baumeister)/
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