Three-dimensional electron crystallography of protein microcrystals

We demonstrate that it is feasible to determine high-resolution protein structures by electron crystallography of three-dimensional crystals in an electron cryo-microscope (CryoEM). Lysozyme microcrystals were frozen on an electron microscopy grid, and electron diffraction data collected to 1.7 Å re...

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Bibliographic Details
Published in:eLife 2013-11, Vol.2, p.e01345-e01345
Main Authors: Shi, Dan, Nannenga, Brent L, Iadanza, Matthew G, Gonen, Tamir
Format: Article
Language:English
Subjects:
Biochemistry
Biophysics and Structural Biology
Crystallography
Crystallography, X-Ray - methods
Crystals
Data collection
Datasets
Diffraction
electron cryomicroscopy
electron crystallography
Electron diffraction
Electron microscopy
Lysozyme
microcrystals
microED
protein structure
Proteins
Proteins - chemistry
Quality
Sensors
X-rays
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Summary:We demonstrate that it is feasible to determine high-resolution protein structures by electron crystallography of three-dimensional crystals in an electron cryo-microscope (CryoEM). Lysozyme microcrystals were frozen on an electron microscopy grid, and electron diffraction data collected to 1.7 Å resolution. We developed a data collection protocol to collect a full-tilt series in electron diffraction to atomic resolution. A single tilt series contains up to 90 individual diffraction patterns collected from a single crystal with tilt angle increment of 0.1-1° and a total accumulated electron dose less than 10 electrons per angstrom squared. We indexed the data from three crystals and used them for structure determination of lysozyme by molecular replacement followed by crystallographic refinement to 2.9 Å resolution. This proof of principle paves the way for the implementation of a new technique, which we name 'MicroED', that may have wide applicability in structural biology. DOI: http://dx.doi.org/10.7554/eLife.01345.001.
ISSN:2050-084X
2050-084X
DOI:10.7554/elife.01345