A Strategy for Electron Tomographic Data Collection and Crystallographic Reconstruction of Biological Bundles |
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Authors: | Michael B Sherman Joanita Jakana Shujun Sun Paul Matsudaira Wah Chiu Michael F Schmid |
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Institution: | aNational Center for Macromolecular Imaging and Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas, 77030;bWhitehead Institute for Biomedical Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, 02142;cNational Center for Macromolecular Imaging and Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas, 77030 |
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Abstract: | Structures of highly ordered biological bundles have unique features which call for special experimental and computational methods in electron cryomicroscopy. They can be considered as three-dimensional quasi-crystals and reconstructed using a crystallographic approach. However, they are neither “infinitely” large with respect to the borders of the bundle, nor are they a single unit cell in thickness along the viewing direction. Also, because of their shape, bundles do not generally have a preferred azimuthal orientation, which poses challenges for orientation estimation and refinement. We developed a strategy for recording and processing electron cryomicroscopic images that differs from classical two-dimensional crystalline reconstruction techniques. These developments allowed us to merge data from tomographic tilt series of ice-embedded acrosomal bundles. The goal is to determine accurately amplitudes and phases at the diffraction maxima in terms ofhklindices, and compute a three-dimensional map from the diffraction data. |
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Keywords: | acrosomal bundles actin scruin electron crystallography tomography electron cryomicroscopy tilt series imaging profile fitting |
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