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New Strategy for Crystallographic Reconstruction of Biological Bundle-Like Structures

Published online by Cambridge University Press:  02 July 2020

M.B. Sherman ,
J. Jakana ,
S. Sunt ,
p. Matsudaira ,
W. Chiu  and
M.F. Schmid
M.B. Sherman
Affiliation:
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX77030;
J. Jakana
Affiliation:
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX77030;
S. Sunt
Affiliation:
The Whitehead Institute for Biomedical Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, 02142
p. Matsudaira
Affiliation:
The Whitehead Institute for Biomedical Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, 02142
W. Chiu
Affiliation:
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX77030;
M.F. Schmid
Affiliation:
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, One Baylor Plaza, Houston, TX77030;
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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

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References

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6. This research has been supported by NIH grant RR02250.Google Scholar