scholarly journals John Thomas Finch. 28 February 1930—5 December 2017

2018 ◽  
Vol 66 ◽  
pp. 183-199
Author(s):  
R. A. Crowther ◽  
K. C. Holmes
Keyword(s):  
Molecular Structure ◽  
Electron Microscopy ◽  
Electron Microscope ◽  
Three Dimensional ◽  
Important Place ◽  
Diverse Range ◽  
X Ray ◽  
X Ray Crystallography ◽  
Embedded Particles ◽  
Team Player

John Finch was a gifted experimentalist who used X-ray crystallography and electron microscopy to elucidate the structures of important biological assemblies, particularly viruses and chromatin. When he started research in the 1950s, little was known about the structure of viruses, and the methods for investigating them were fairly limited. His early work on crystals of viruses was important in establishing their symmetry, and later with the electron microscope he mapped out the molecular structure of many virus coats. His observations on negatively stained preparations demonstrated that images of particles prepared in this way represented projections of fully stained embedded particles, not merely one-sided footprints. This was very relevant to the development of methods for making three-dimensional maps of specimens from electron micrographs. Later, besides further studies of viruses, he worked on many other systems, including chromatin, nucleosomes and tRNA. John was very much a team player and held an important place as the key experimentalist in many influential collaborations, investigating a diverse range of biological specimens.

Microtubules, sickle cell hemoglobin fibers, and various crystal structures

1983 ◽  
Vol 41 ◽  
pp. 422-425
Author(s):  
Stuart J. Edelstein ◽  
Richard H. Crepeau ◽  
Christopher W. Akey ◽  
Thomas A. Ceska
Keyword(s):  
Molecular Structure ◽  
Electron Microscopy ◽  
Cytochrome Oxidase ◽  
Sickle Cell ◽  
Three Dimensional ◽  
Beef Heart ◽  
Structural Studies ◽  
Image Reconstruction Techniques ◽  
X Ray ◽  
X Ray Crystallography

Structural studies involving electron microscopy and image reconstruction techniques are now approaching levels of resolution that had previously been the domain of x-ray crystallography. Studies on purple membrane are a prime example. Although few other systems have reached comparable levels of resolution, progress is being made on several fronts. Citing work from this laboratory, I will illustrate three ways we are using electron microscopy to approach the border with x-ray crystallography, in studies on: three-dimensional crystals (cytochrome oxidase from P. aeruginosa, catalase, and beef heart F1-ATPase); arrays of proteins which do not form three-dimensional crystals (tubulin sheets and microtubules); and the supra-molecular structure of a protein of known atomic structure (sickle cell hemoglobin).A. Electron microscopy as an adjunct to X-ray crystallography for threedimensional crystalsOur applications in this area have primarily consisted of embedding and sectioning techniques with crystals that are too small or otherwise unsuitable for x-ray crystallography, such as occur for cytochrome oxidase from P. aeruginosa or beef heart F1-ATPase.

scholarly journals The Resolution in X-ray Crystallography and Single-Particle Cryogenic Electron Microscopy

Crystals ◽  
2020 ◽  
Vol 10 (7) ◽  
pp. 580
Author(s):  
Victor R.A. Dubach ◽  
Albert Guskov
Keyword(s):  
Electron Microscopy ◽  
Single Particle ◽  
Signal To Noise Ratio ◽  
Three Dimensional ◽  
Particle Analysis ◽  
Biological Macromolecules ◽  
Single Particle Analysis ◽  
X Ray ◽  
X Ray Crystallography ◽  
The Fourier Transform

X-ray crystallography and single-particle analysis cryogenic electron microscopy are essential techniques for uncovering the three-dimensional structures of biological macromolecules. Both techniques rely on the Fourier transform to calculate experimental maps. However, one of the crucial parameters, resolution, is rather broadly defined. Here, the methods to determine the resolution in X-ray crystallography and single-particle analysis are summarized. In X-ray crystallography, it is becoming increasingly more common to include reflections discarded previously by traditionally used standards, allowing for the inclusion of incomplete and anisotropic reflections into the refinement process. In general, the resolution is the smallest lattice spacing given by Bragg’s law for a particular set of X-ray diffraction intensities; however, typically the resolution is truncated by the user during the data processing based on certain parameters and later it is used during refinement. However, at which resolution to perform such a truncation is not always clear and this makes it very confusing for the novices entering the structural biology field. Furthermore, it is argued that the effective resolution should be also reported as it is a more descriptive measure accounting for anisotropy and incompleteness of the data. In single particle cryo-EM, the situation is not much better, as multiple ways exist to determine the resolution, such as Fourier shell correlation, spectral signal-to-noise ratio and the Fourier neighbor correlation. The most widely accepted is the Fourier shell correlation using a threshold of 0.143 to define the resolution (so-called “gold-standard”), although it is still debated whether this is the correct threshold. Besides, the resolution obtained from the Fourier shell correlation is an estimate of varying resolution across the density map. In reality, the interpretability of the map is more important than the numerical value of the resolution.

scholarly journals Sir Aaron Klug OM. 11 August 1926—20 November 2018

2020 ◽  
Vol 68 ◽  
pp. 273-296
Author(s):  
R. A. Crowther
Keyword(s):  
Electron Microscopy ◽  
Molecular Biology ◽  
Royal Society ◽  
Zinc Fingers ◽  
Three Dimensional ◽  
Transfer Rna ◽  
Biological Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Atomic Structures

Aaron Klug made outstanding contributions to the development of structural molecular biology. An early interest in viruses, stemming from work with Rosalind Franklin, prompted him to think deeply about extracting the information contained in electron micrographs. As a result, he proposed a method for making three-dimensional maps of biological specimens from the projected images given by micrographs. For this development and its application to complex molecular assemblies, he was awarded the 1982 Nobel Prize in Chemistry. The recent revolution in biological structure determination, whereby atomic structures can now be determined from micrographs of frozen hydrated specimens, derives from this initial breakthrough. With colleagues, Aaron applied X-ray crystallography and electron microscopy to determine the structures and thereby understand the functions of many biological assemblies, including viruses, transfer RNA, chromatin and zinc fingers. He also made important forays into the pathogenesis of Alzheimer's disease and related dementias. Aaron was director of the MRC Laboratory of Molecular Biology in Cambridge from 1986 to 1996 and President of the Royal Society from 1995 to 2000.

Bacteriophage PRD1 Capsid Structure: Iterative Combination of Threedimensional Electron Microscopy and X-Ray Crystallography

2000 ◽  
Vol 6 (S2) ◽  
pp. 284-285
Author(s):  
Carmen San Martin ◽  
Roger M. Burnett ◽  
Felix de Haas ◽  
Ralph Heinkel ◽  
Twan Rutten ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Three Dimensional ◽  
Cyclic Process ◽  
Particle Structure ◽  
X Ray ◽  
X Ray Crystallography ◽  
Bacteriophage Prd1 ◽  
Cryo Electron Microscopy ◽  
Moderate Resolution ◽  
Protein Rich

PRD1 is a ds-DNA bacteriophage from the Tectiviridae family with an unusual structural feature: the viral genome is enclosed by a protein-rich membrane, which is in turn enclosed by an external icosahedral protein shell (capsid). Three-dimensional reconstructions from cryo-electron microscopy (cryo-EM) images have revealed the structure of the PRD1 capsid at moderate resolution (28 Å), while X-ray crystallographic studies have recently provided a high resolution (1.85 Å) picture of the major coat protein, P3. We have now combined these results from different imaging methods to obtain a more detailed understanding of the virion organization. The combination has been made in a cyclic process: a preliminary fitting of the atomic structure of P3 to each one of its independent positions in the cryo-EM maps of the capsids provided initial models that could be used to improve the reconstructions; the refined maps then served as a base frame for an optimized fit. This process allows us to study the viral particle structure at “quasi-atomic” resolution.

Three-Dimensional Structure Of C3D From Low Temperature Scanning Transmission Electron Microscopy and X-Ray Crystallography

2000 ◽  
Vol 6 (S2) ◽  
pp. 294-295
Author(s):  
D.J. Martin ◽  
F.P. Ottensmeyer
Keyword(s):  
Electron Microscopy ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
X Rays ◽  
Complement Protein ◽  
X Ray ◽  
X Ray Crystallography ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Scanning Transmission

Macromolecular structure can be solved by x-ray crystallography to atomic resolution provided that the molecule can be crystallized, that the crystals diffract x-rays to high resolution, and that the phases of the diffracted x-rays can be determined. Though the resolution of single particle imaging by electron microscopy is lower than that of x-ray diffraction by crystals, electron microscopy can directly image a large molecular weight range of macromolecules in a non-crystalline environment, and provide the basis for the three-dimensional reconstruction of these structures. To investigate combining structural information from x-ray crystallography and electron microscopy for unknown structures, we have imaged a small protein of known structure (1), the 35 kDa human complement protein fragment C3d, in a scanning transmission electron microscope (STEM). The intention is to eventually combine the knowledge of electron densities and molecular boundaries from electron microscopy to assist in phase determination in x-ray crystallography.

Preliminary Three-Dimensional Model of Insect Lipoprotein HDLp by Using Electron Microscopy and X-ray Crystallography

2004 ◽  
Vol 10 (S02) ◽  
pp. 1514-1515 ◽  
Author(s):  
Karine M Valentijn ◽  
Roman Koning ◽  
Yvonne Derks ◽  
Jan M Van Doorn ◽  
Theo P Van der Krift ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Three Dimensional ◽  
Dimensional Model ◽  
Three Dimensional Model ◽  
X Ray ◽  
X Ray Crystallography

Extended abstract of a paper presented at Microscopy and Microanalysis 2004 in Savannah, Georgia, USA, August 1–5, 2004.

Methodology for determining the three-dimensional crystal structure of myosin Sl from electron microscopy of orthogonal thin sections

1986 ◽  
Vol 44 ◽  
pp. 26-29
Author(s):  
T. S. Baker ◽  
D. A. Winkelmann
Keyword(s):  
Crystal Structure ◽  
Electron Microscopy ◽  
Single Unit ◽  
Three Dimensional ◽  
Detailed Knowledge ◽  
Thin Sections ◽  
X Ray ◽  
X Ray Crystallography ◽  
Density Map ◽  
Dimensional Crystal

Detailed knowledge of the structure of myosin is essential for understanding the mechanism of muscle contraction. The discovery of conditions for crystallizing the head of myosin (Sl = subfragment 1) has led to systematic studies of the SI structure by x-ray crystallography and electron microscopy. We describe the method used to determine the structure of the myosin Sl molecule by electron microscopy (Fig. 3). This method involved independently reconstructing the three-dimensional density of several thin sections obtained from oriented crystals and then combining these reconstructions to produce a final, averaged density map of a single unit cell.

Image Processing and Lattice Determination for Three-Dimensional Nanocrystals

2011 ◽  
Vol 17 (6) ◽  
pp. 879-885 ◽  
Author(s):  
Linhua Jiang ◽  
Dilyana Georgieva ◽  
Igor Nederlof ◽  
Zunfeng Liu ◽  
Jan Pieter Abrahams
Keyword(s):  
Molecular Structure ◽  
Three Dimensional ◽  
X Ray ◽  
X Ray Crystallography ◽  
Hard Materials ◽  
Cryo Electron Microscopy ◽  
Diffraction Patterns ◽  
Radiation Hard ◽  
Orientation Parameters

AbstractThree-dimensional nanocrystals can be studied by electron diffraction using transmission cryo-electron microscopy. For molecular structure determination of proteins, such nanosized crystalline samples are out of reach for traditional single-crystal X-ray crystallography. For the study of materials that are not sensitive to the electron beam, software has been developed for determining the crystal lattice and orientation parameters. These methods require radiation-hard materials that survive careful orienting of the crystals and measuring diffraction of one and the same crystal from different, but known directions. However, as such methods can only deal with well-oriented crystalline samples, a problem exists for three-dimensional (3D) crystals of proteins and other radiation sensitive materials that do not survive careful rotational alignment in the electron microscope. Here, we discuss our newly released software AMP that can deal with nonoriented diffraction patterns, and we discuss the progress of our new preprocessing program that uses autocorrelation patterns of diffraction images for lattice determination and indexing of 3D nanocrystals.

3-D reconstruction of molecular shape from microcrystal thin sections

1983 ◽  
Vol 41 ◽  
pp. 748-749
Author(s):  
S. Cusack ◽  
J.-C. Jésior
Keyword(s):  
Three Dimensional ◽  
Molecular Shape ◽  
Biological Molecules ◽  
Fourier Space ◽  
Single Particles ◽  
Thin Sections ◽  
Standard Methods ◽  
X Ray ◽  
X Ray Crystallography ◽  
Dimensional Reconstruction

Three-dimensional reconstruction techniques using electron microscopy have been principally developed for application to 2-D arrays (i.e. monolayers) of biological molecules and symmetrical single particles (e.g. helical viruses). However many biological molecules that crystallise form multilayered microcrystals which are unsuitable for study by either the standard methods of 3-D reconstruction or, because of their size, by X-ray crystallography. The grid sectioning technique enables a number of different projections of such microcrystals to be obtained in well defined directions (e.g. parallel to crystal axes) and poses the problem of how best these projections can be used to reconstruct the packing and shape of the molecules forming the microcrystal.Given sufficient projections there may be enough information to do a crystallographic reconstruction in Fourier space. We however have considered the situation where only a limited number of projections are available, as for example in the case of catalase platelets where three orthogonal and two diagonal projections have been obtained (Fig. 1).

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.

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