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J Struct Biol. 2014 Mar;185(3):418-26. doi: 10.1016/j.jsb.2013.12.010. Epub 2013 Dec 30.

ResLog plots as an empirical metric of the quality of cryo-EM reconstructions.

Author information

1
Department of Chemistry and Biochemistry, 95 Chieftan Way, Florida State University, Tallahassee, FL 32306-4380, United States; Institute of Molecular Biophysics, 91 Chieftan Way, Florida State University, Tallahassee, FL 32306-4380, United States. Electronic address: sstagg@mac.com.
2
Department of Physics, 77 Chieftan Way, Florida State University, Tallahassee, FL 32306-4350, United States.
3
Institute of Molecular Biophysics, 91 Chieftan Way, Florida State University, Tallahassee, FL 32306-4380, United States.
4
Department of Biochemistry & Molecular Biology, School of Medicine, Oregon Health & Science University, Portland, OR 97239-3098, United States.

Abstract

Compared to the field of X-ray crystallography, the field of single particle three-dimensional electron microscopy has few reliable metrics for assessing the quality of 3D reconstructions. New metrics are needed that can determine whether a given 3D reconstruction accurately reflects the structure of the particles from which it was derived or instead depicts a plausible though incorrect structure due to coarse misalignment of particles. Here an empirical procedure is presented for differentiating between a reconstruction with well-aligned particles and a reconstruction with grossly misclassified particles. For a given dataset, 3D reconstructions are computed from subsets of particles with decreasing numbers of particles contributing to the reconstruction. A plot of inverse resolution vs. the logarithm of the number of particles (a "ResLog" plot) provides metrics for the reliability of the reconstruction and the overall quality of the dataset and processing. Specifically, the y-intercept of a regression line provides a measure of the relative accuracy of the particle alignment and classification, and the slope is an indicator of the overall data quality including the imaging conditions and processing steps. ResLog plots can also be used to optimize conditions for data collection and reconstruction parameters. Although resolution estimates can vary by method of calculation, ResLog-derived parameters are consistent whether calculated by Fourier shell correlation or Fourier neighbor correlation, or a new coordinate-based metric that serves as a yardstick for structures where atomic coordinates are available. ResLog plots could become part of a standard set of parameters to be included in 3D reconstruction reports.

KEYWORDS:

Electron microscopy; Noise bias; Resolution; Structure validation

PMID:
24384117
PMCID:
PMC4001718
DOI:
10.1016/j.jsb.2013.12.010
[Indexed for MEDLINE]
Free PMC Article

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