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New Strategy for Crystallographic Reconstruction of Biological Bundle-Like Structures
Published online by Cambridge University Press: 02 July 2020
Extract
The acrosomal bundle is an intracellular quasi-crystalline organelle in the head of the sperm of Limulus polyphemus . It is a long (up to 60 μm) straight bundle ∼1000 Å in diameter1 consisting of two major proteins in a 1:1 stoichiometric ratio: actin (42 kDa) and scruin (102 kDa) with a minor calmodulin-like protein (14 kDa) presumably bound to scruin molecule. Previous helical reconstructions of single filaments in the bundle showed actin-scruin interactions. We recorded tilt series from single bundles in a 400 kV cryomicroscope and have developed a novel crystallographic technique to reconstruct a unit cell from the bundle as a whole to reveal interfilament interactions.
Acrosomal bundles were purified as described elsewhere and embedded in vitreous ice over holes on a holey carbon film. Tilt series images were collected in a JEOL 4000EX electron cryo-microscope at 400 kV using 10,000x microscope magnification. Total dose was ∼16-18 electrons/Å per series covering a tilt range of ± 60° with 5° angular increment.
- Type
- Electron Crystallography; the Electron Phase Problem
- Information
- Microscopy and Microanalysis , Volume 3 , Issue S2: Proceedings: Microscopy & Microanalysis '97, Microscopy Society of America 55th Annual Meeting, Microbeam Analysis Society 31st Annual Meeting, Histochemical Society 48th Annual Meeting, Cleveland, Ohio, August 10-14, 1997 , August 1997 , pp. 1035 - 1036
- Copyright
- Copyright © Microscopy Society of America 1997