scholarly journals A fully automatic method yielding initial models from high-resolution electron cryo-microscopy maps

2018 ◽  
Author(s):  
Thomas C. Terwilliger ◽  
Paul D. Adams ◽  
Pavel V. Afonine ◽  
Oleg V. Sobolev
Keyword(s):  
High Resolution ◽  
Automatic Method ◽  
Resolution Electron ◽  
Fully Automatic ◽  
Rna Complexes ◽  
Median Fraction

AbstractA fully automated procedure for optimization and interpretation of reconstructions from cryo-EM is developed and applied to 476 datasets with resolution of 4.5 Å or better, including reconstructions of 47 ribosomes and 32 other protein-RNA complexes. The median fraction of residues in the deposited structures reproduced automatically was 71% for reconstructions determined at resolutions of 3 Å or better and 47% for those at lower resolution.

scholarly journals A fully automatic method yielding initial models from high-resolution cryo-electron microscopy maps

2018 ◽  
Vol 15 (11) ◽  
pp. 905-908 ◽  
Author(s):  
Thomas C. Terwilliger ◽  
Paul D. Adams ◽  
Pavel V. Afonine ◽  
Oleg V. Sobolev
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Automatic Method ◽  
Cryo Electron Microscopy ◽  
Fully Automatic

Light Optical Diffraction Studies of Macromolecular Ultrastructure

1970 ◽  
Vol 28 ◽  
pp. 258-259
Author(s):  
Glen B. Haydon
Keyword(s):  
High Resolution ◽  
Diffraction Analysis ◽  
Diffraction Pattern ◽  
Electron Micrographs ◽  
Optical Diffraction ◽  
Electron Micrograph ◽  
Its Analysis ◽  
Resolution Electron ◽  
Diffraction Patterns

Analysis of light optical diffraction patterns produced by electron micrographs can easily lead to much nonsense. Such diffraction patterns are referred to as optical transforms and are compared with transforms produced by a variety of mathematical manipulations. In the use of light optical diffraction patterns to study periodicities in macromolecular ultrastructures, a number of potential pitfalls have been rediscovered. The limitations apply to the formation of the electron micrograph as well as its analysis.(1) The high resolution electron micrograph is itself a complex diffraction pattern resulting from the specimen, its stain, and its supporting substrate. Cowley and Moodie (Proc. Phys. Soc. B, LXX 497, 1957) demonstrated changing image patterns with changes in focus. Similar defocus images have been subjected to further light optical diffraction analysis.

Structural analysis of the surface-layer protein of spirillum serpens by high-resolution electron microscopy

1983 ◽  
Vol 41 ◽  
pp. 738-739
Author(s):  
W. H. Wu ◽  
R. M. Glaeser
Keyword(s):  
Electron Microscopy ◽  
Surface Layer ◽  
Structural Analysis ◽  
High Resolution ◽  
Low Dose ◽  
High Resolution Electron Microscopy ◽  
Optical Diffraction ◽  
Surface Layer Protein ◽  
Morphological Unit ◽  
Resolution Electron

Spirillum serpens possesses a surface layer protein which exhibits a regular hexagonal packing of the morphological subunits. A morphological model of the structure of the protein has been proposed at a resolution of about 25 Å, in which the morphological unit might be described as having the appearance of a flared-out, hollow cylinder with six ÅspokesÅ at the flared end. In order to understand the detailed association of the macromolecules, it is necessary to do a high resolution structural analysis. Large, single layered arrays of the surface layer protein have been obtained for this purpose by means of extensive heating in high CaCl2, a procedure derived from that of Buckmire and Murray. Low dose, low temperature electron microscopy has been applied to the large arrays.As a first step, the samples were negatively stained with neutralized phosphotungstic acid, and the specimens were imaged at 40,000 magnification by use of a high resolution cold stage on a JE0L 100B. Low dose images were recorded with exposures of 7-9 electrons/Å2. The micrographs obtained (Fig. 1) were examined by use of optical diffraction (Fig. 2) to tell what areas were especially well ordered.

Electron microscopy of rabbit FC crystals

1983 ◽  
Vol 41 ◽  
pp. 730-731
Author(s):  
Robert A. Grant ◽  
Laura L. Degn ◽  
Wah Chiu ◽  
John Robinson
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
High Resolution Electron Microscopy ◽  
Molecular Replacement ◽  
X Ray ◽  
X Ray Crystallography ◽  
Resolution Electron ◽  
Replacement Technique ◽  
Hydrated Crystal ◽  
Molecular Structure Analysis

Proteolytic digestion of the immunoglobulin IgG with papain cleaves the molecule into an antigen binding fragment, Fab, and a compliment binding fragment, Fc. Structures of intact immunoglobulin, Fab and Fc from various sources have been solved by X-ray crystallography. Rabbit Fc can be crystallized as thin platelets suitable for high resolution electron microscopy. The structure of rabbit Fc can be expected to be similar to the known structure of human Fc, making it an ideal specimen for comparing the X-ray and electron crystallographic techniques and for the application of the molecular replacement technique to electron crystallography. Thin protein crystals embedded in ice diffract to high resolution. A low resolution image of a frozen, hydrated crystal can be expected to have a better contrast than a glucose embedded crystal due to the larger density difference between protein and ice compared to protein and glucose. For these reasons we are using an ice embedding technique to prepare the rabbit Fc crystals for molecular structure analysis by electron microscopy.

An Analysis of Dissociation of Molecules in a High Resolution Electron Microscope

1973 ◽  
Vol 31 ◽  
pp. 486-487
Author(s):  
Mihir Parikh
Keyword(s):  
Electron Microscope ◽  
High Resolution ◽  
Cross Sections ◽  
Resolving Power ◽  
Peak Intensity ◽  
Molecular Film ◽  
Resolution Electron ◽  
Dissociation Cross Section ◽  
High Resolution Electron Microscope ◽  
The Stability

It is well known that the resolution of bio-molecules in a high resolution electron microscope depends not just on the physical resolving power of the instrument, but also on the stability of these molecules under the electron beam. Experimentally, the damage to the bio-molecules is commo ly monitored by the decrease in the intensity of the diffraction pattern, or more quantitatively by the decrease in the peaks of an energy loss spectrum. In the latter case the exposure, EC, to decrease the peak intensity from IO to I’O can be related to the molecular dissociation cross-section, σD, by EC = ℓn(IO /I’O) /ℓD. Qu ntitative data on damage cross-sections are just being reported, However, the microscopist needs to know the explicit dependence of damage on: (1) the molecular properties, (2) the density and characteristics of the molecular film and that of the support film, if any, (3) the temperature of the molecular film and (4) certain characteristics of the electron microscope used

The High Resolution Appearance of Biological Substrates

1969 ◽  
Vol 27 ◽  
pp. 416-417
Author(s):  
Glen B. Haydon
Keyword(s):  
High Resolution ◽  
Phase Image ◽  
Biological Structure ◽  
Electron Microscopic ◽  
Thin Sections ◽  
Resolution Electron ◽  
Discrete Structures ◽  
Large Phase ◽  
Biological Substrates ◽  
High Resolution Electron Micrographs

High resolution electron microscopic study of negatively stained macromolecules and thin sections of tissue embedded in a variety of media are difficult to interpret because of the superimposed phase image granularity. Although all of the information concerning the biological structure of interest may be present in a defocused electron micrograph, the high contrast of large phase image granules produced by the substrate makes it impossible to distinguish the phase ‘points’ from discrete structures of the same dimensions. Theory predicts the findings; however, it does not allow an appreciation of the actual appearance of the image under various conditions. Therefore, though perhaps trivial, training of the cheapest computer produced by mass labor has been undertaken in order to learn to appreciate the factors which affect the appearance of the background in high resolution electron micrographs.

High resolution electron microscopy of lanthanum phosphate crystals

1978 ◽  
Vol 36 (1) ◽  
pp. 274-275
Author(s):  
J. C. Wheatley ◽  
J. M. Cowley
Keyword(s):  
Electron Microscopy ◽  
Catalytic Activity ◽  
High Resolution ◽  
Petroleum Industry ◽  
High Resolution Electron Microscopy ◽  
Lanthanum Phosphate ◽  
Good Catalyst ◽  
Resolution Electron ◽  
Good Catalytic Activity ◽  
Physical And Chemical

Rare-earth phosphates are of particular interest because of their catalytic properties associated with the hydrolysis of many aromatic chlorides in the petroleum industry. Lanthanum phosphates (LaPO4) which have been doped with small amounts of copper have shown increased catalytic activity (1). However the physical and chemical characteristics of the samples leading to good catalytic activity are not known.Many catalysts are amorphous and thus do not easily lend themselves to methods of investigation which would include electron microscopy. However, the LaPO4, crystals are quite suitable samples for high resolution techniques.The samples used were obtained from William L. Kehl of Gulf Research and Development Company. The electron microscopy was carried out on a JEOL JEM-100B which had been modified for high resolution microscopy (2). Standard high resolution techniques were employed. Three different sample types were observed: 669A-1-5-7 (poor catalyst), H-L-2 (good catalyst) and 27-011 (good catalyst).

Possible applications of fraunhofer holography in high resolution electron microscopy

1978 ◽  
Vol 36 (1) ◽  
pp. 222-223 ◽  
Author(s):  
N. Bonnet ◽  
M. Troyon ◽  
P. Gallion
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Transfer Function ◽  
Radiation Damage ◽  
High Resolution Electron Microscopy ◽  
Far Field ◽  
Field Region ◽  
Crystalline Materials ◽  
Resolution Electron ◽  
The Ideal

Two main problems in high resolution electron microscopy are first, the existence of gaps in the transfer function, and then the difficulty to find complex amplitude of the diffracted wawe from registered intensity. The solution of this second problem is in most cases only intended by the realization of several micrographs in different conditions (defocusing distance, illuminating angle, complementary objective apertures…) which can lead to severe problems of contamination or radiation damage for certain specimens.Fraunhofer holography can in principle solve both problems stated above (1,2). The microscope objective is strongly defocused (far-field region) so that the two diffracted beams do not interfere. The ideal transfer function after reconstruction is then unity and the twin image do not overlap on the reconstructed one.We show some applications of the method and results of preliminary tests.Possible application to the study of cavitiesSmall voids (or gas-filled bubbles) created by irradiation in crystalline materials can be observed near the Scherzer focus, but it is then difficult to extract other informations than the approximated size.

Electron crystallographic determination of the 3-D structure of staurolite at atomic resolution

1991 ◽  
Vol 49 ◽  
pp. 540-541
Author(s):  
Kenneth H. Downing ◽  
Hu Meisheng ◽  
Hans-Rudolf Went ◽  
Michael A. O'Keefe
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
High Resolution Electron Microscopy ◽  
Atomic Resolution ◽  
Dimensional Structure ◽  
Structure Information ◽  
Resolution Electron ◽  
Scattering Effects ◽  
High Resolution Images ◽  
Effects Of Radiation

With current advances in electron microscope design, high resolution electron microscopy has become routine, and point resolutions of better than 2Å have been obtained in images of many inorganic crystals. Although this resolution is sufficient to resolve interatomic spacings, interpretation generally requires comparison of experimental images with calculations. Since the images are two-dimensional representations of projections of the full three-dimensional structure, information is invariably lost in the overlapping images of atoms at various heights. The technique of electron crystallography, in which information from several views of a crystal is combined, has been developed to obtain three-dimensional information on proteins. The resolution in images of proteins is severely limited by effects of radiation damage. In principle, atomic-resolution, 3D reconstructions should be obtainable from specimens that are resistant to damage. The most serious problem would appear to be in obtaining high-resolution images from areas that are thin enough that dynamical scattering effects can be ignored.

Alternative approaches to ultra-high resolution imaging

1991 ◽  
Vol 49 ◽  
pp. 650-651
Author(s):  
J.M. Cowley
Keyword(s):  
High Resolution ◽  
High Voltage ◽  
High Resolution Electron Microscopy ◽  
Crystal Defects ◽  
High Resolution Imaging ◽  
General Applicability ◽  
Resolution Electron ◽  
Radiation Resistant ◽  
Resolution Imaging ◽  
Alternative Approaches

By extrapolation of past experience, it would seem that the future of ultra-high resolution electron microscopy rests with the advances of electron optical engineering that are improving the instrumental stability of high voltage microscopes to achieve the theoretical resolutions of 1Å or better at 1MeV or higher energies. While these high voltage instruments will undoubtedly produce valuable results on chosen specimens, their general applicability has been questioned on the basis of the excessive radiation damage effects which may significantly modify the detailed structures of crystal defects within even the most radiation resistant materials in a period of a few seconds. Other considerations such as those of cost and convenience of use add to the inducement to consider seriously the possibilities for alternative approaches to the achievement of comparable resolutions.

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