scholarly journals Ribosome structures to near-atomic resolution from thirty thousand cryo-EM particles

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Xiao-chen Bai ◽  
Israel S Fernandez ◽  
Greg McMullan ◽  
Sjors HW Scheres
Keyword(s):  
High Resolution ◽  
Structural Biology ◽  
Atomic Resolution ◽  
Particle Analysis ◽  
Full Potential ◽  
Single Particle Analysis ◽  
Noise Performance ◽  
Signal To Noise ◽  
Fundamental Limits ◽  
New Generation

Although electron cryo-microscopy (cryo-EM) single-particle analysis has become an important tool for structural biology of large and flexible macro-molecular assemblies, the technique has not yet reached its full potential. Besides fundamental limits imposed by radiation damage, poor detectors and beam-induced sample movement have been shown to degrade attainable resolutions. A new generation of direct electron detectors may ameliorate both effects. Apart from exhibiting improved signal-to-noise performance, these cameras are also fast enough to follow particle movements during electron irradiation. Here, we assess the potentials of this technology for cryo-EM structure determination. Using a newly developed statistical movie processing approach to compensate for beam-induced movement, we show that ribosome reconstructions with unprecedented resolutions may be calculated from almost two orders of magnitude fewer particles than used previously. Therefore, this methodology may expand the scope of high-resolution cryo-EM to a broad range of biological specimens.

scholarly journals On the explicit use of experimental images in high resolution cryo-EM refinement

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 665
Author(s):  
Jacqueline Cherfils ◽  
Jorge Navaza
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Structural Biology ◽  
Single Particle ◽  
Atomic Resolution ◽  
Full Potential ◽  
X Ray ◽  
X Ray Crystallography ◽  
Atomic Models

Single particle cryogenic electron microscopy (cryo-EM) is transforming structural biology by enabling the analysis of difficult macromolecular specimens, such as membrane proteins or large complexes with flexible elements, at near atomic resolution with an accuracy close to that of X-ray crystallography. As the technique continues to improve, it is important to assess and exploit its full potential to produce the most possible reliable atomic models. Here we propose to use the experimental images as the data for refinement and validation, instead of the reconstructed maps as currently used. This procedure, which is in spirit quite similar to that used in X-ray crystallography where the data include experimental phases, should contribute to improve the quality of the cryo-EM atomic models.

scholarly journals On the explicit use of experimental images in high resolution cryo-EM refinement

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 665
Author(s):  
Jacqueline Cherfils ◽  
Jorge Navaza
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Membrane Proteins ◽  
Structural Biology ◽  
Single Particle ◽  
Atomic Resolution ◽  
Full Potential ◽  
X Ray ◽  
X Ray Crystallography

Single particle cryogenic electron microscopy (cryo-EM) is transforming structural biology by enabling the analysis of difficult macromolecular specimens, such as membrane proteins or large complexes with flexible elements, at near atomic resolution with an accuracy close to that of X-ray crystallography. As the technique continues to improve, it is important to assess and exploit its full potential to produce the most possible reliable atomic models. Here we propose to use the experimental images as the data for refinement and validation, instead of the reconstructed maps as currently used. This procedure, which is in spirit quite similar to that used in X-ray crystallography where the data include experimental phases, should contribute to improve the quality of the models.

A3Near-atomic Resolution Single Particle Analysis with the Volta Phase Plate

Microscopy ◽  
2015 ◽  
Vol 64 (suppl 1) ◽  
pp. i9.2-i9
Author(s):  
Radostin Danev ◽  
Maryam Khoshouei ◽  
Wolfgang Baumeister
Keyword(s):  
Single Particle ◽  
Atomic Resolution ◽  
Particle Analysis ◽  
Phase Plate ◽  
Single Particle Analysis ◽  
Resolution Single

scholarly journals High-resolution Single Particle Analysis from Electron Cryo-microscopy Images Using SPHIRE

10.3791/55448 ◽  
2017 ◽  
Author(s):  
Toshio Moriya ◽  
Michael Saur ◽  
Markus Stabrin ◽  
Felipe Merino ◽  
Horatiu Voicu ◽  
...  
Keyword(s):  
High Resolution ◽  
Single Particle ◽  
Particle Analysis ◽  
Single Particle Analysis ◽  
Resolution Single ◽  
Microscopy Images

The stressosome: molecular architecture of a signalling hub

2010 ◽  
Vol 38 (4) ◽  
pp. 928-933 ◽  
Author(s):  
Jon Marles-Wright ◽  
Richard J. Lewis
Keyword(s):  
Uv Light ◽  
Atomic Resolution ◽  
Particle Analysis ◽  
Macromolecular Structure ◽  
Single Particle Analysis ◽  
Molecular Architecture ◽  
Signal Transduction System ◽  
Live Bacteria ◽  
The Individual ◽  
Signalling Cascade

The stressosome co-ordinates the response of Bacillus subtilis to the imposition of a variety of physical and environmental insults. These stresses include fluctuations in salt concentration, the presence of ethanol, changes in pH and even the level of UV light. Despite the obvious and significant differences between these quite different physicochemical stimuli, the result is the same: the stressosome is phosphorylated by a key kinase to initiate the σB cascade. The phosphorylation of the stressosome initiates a signal transduction system that up-regulates the expression of stress-responsive genes so that the Bacillus can survive the imposition of stress. Hence the stressosome acts as a hub, receiving manifold different stimuli to effect a single outcome. Using single-particle analysis of cryo-electron micrographs, we have been able to reconstruct a series of molecular envelopes of the stressosome. These maps have been interpreted at near-atomic resolution with crystal structures of the individual components of the stressosome to provide the first visualization of this unique signalling hub. The macromolecular structure adopted by the stressosome provides the signalling cascade with the potential for co-operative behaviour, which we have also measured in live bacteria. These experiments are consistent with the tuning of the response of B. subtilis to stress relative to the magnitude of the insult.

scholarly journals EMAN2.1 - A New Generation of Software for Validated Single Particle Analysis and Single Particle Tomography

2014 ◽  
Vol 20 (S3) ◽  
pp. 832-833 ◽  
Author(s):  
SC Murray ◽  
JG Galaz-Montoya ◽  
G Tang ◽  
JF Flanagan ◽  
SJ Ludtke
Keyword(s):  
Single Particle ◽  
Particle Analysis ◽  
Single Particle Analysis ◽  
New Generation

scholarly journals Survey of the analysis of continuous conformational variability of biological macromolecules by electron microscopy

2019 ◽  
Vol 75 (1) ◽  
pp. 19-32 ◽  
Author(s):  
C. O. S. Sorzano ◽  
A. Jiménez ◽  
J. Mota ◽  
J. L. Vilas ◽  
D. Maluenda ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Image Processing ◽  
High Resolution ◽  
Conformational Changes ◽  
Three Dimensional ◽  
Particle Analysis ◽  
Biological Macromolecules ◽  
Single Particle Analysis ◽  
Conformational Variability ◽  
Established Technique

Single-particle analysis by electron microscopy is a well established technique for analyzing the three-dimensional structures of biological macromolecules. Besides its ability to produce high-resolution structures, it also provides insights into the dynamic behavior of the structures by elucidating their conformational variability. Here, the different image-processing methods currently available to study continuous conformational changes are reviewed.

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