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J Struct Biol. 2015 Nov;192(2):209-15. doi: 10.1016/j.jsb.2015.06.014. Epub 2015 Jun 15.

Description and comparison of algorithms for correcting anisotropic magnification in cryo-EM images.

Author information

1
Molecular Structure and Function Program, The Hospital for Sick Children, Canada; Department of Medical Biophysics, University of Toronto, Canada.
2
Department of Computer Science, University of Toronto, Canada.
3
Molecular Structure and Function Program, The Hospital for Sick Children, Canada.
4
Molecular Structure and Function Program, The Hospital for Sick Children, Canada; Department of Medical Biophysics, University of Toronto, Canada; Department of Biochemistry, University of Toronto, Canada. Electronic address: john.rubinstein@utoronto.ca.

Abstract

Single particle electron cryomicroscopy (cryo-EM) allows for structures of proteins and protein complexes to be determined from images of non-crystalline specimens. Cryo-EM data analysis requires electron microscope images of randomly oriented ice-embedded protein particles to be rotated and translated to allow for coherent averaging when calculating three-dimensional (3D) structures. Rotation of 2D images is usually done with the assumption that the magnification of the electron microscope is the same in all directions. However, due to electron optical aberrations, this condition is not met with some electron microscopes when used with the settings necessary for cryo-EM with a direct detector device (DDD) camera. Correction of images by linear interpolation in real space has allowed high-resolution structures to be calculated from cryo-EM images for symmetric particles. Here we describe and compare a simple real space method, a simple Fourier space method, and a somewhat more sophisticated Fourier space method to correct images for a measured anisotropy in magnification. Further, anisotropic magnification causes contrast transfer function (CTF) parameters estimated from image power spectra to have an apparent systematic astigmatism. To address this problem we develop an approach to adjust CTF parameters measured from distorted images so that they can be used with corrected images. The effect of anisotropic magnification on CTF parameters provides a simple way of detecting magnification anisotropy in cryo-EM datasets.

KEYWORDS:

Anisotropic magnification; Contrast transfer function; Correction; Cryo-EM; Differential magnification; Direct detector device; Direct electron detector; Distortion

PMID:
26087140
DOI:
10.1016/j.jsb.2015.06.014
[Indexed for MEDLINE]

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