Electron Diffraction of Platinum Labelled Bacteriorhodopsin

1980 ◽  
Vol 38 ◽  
pp. 34-35
Author(s):  
M. E. Dumont ◽  
J. W. Wiggins ◽  
S. B. Hayward
Keyword(s):  
Amino Acids ◽  
High Resolution ◽  
Electron Diffraction ◽  
Structural Information ◽  
Heavy Atom ◽  
Halobacterium Halobium ◽  
Heavy Atoms ◽  
Isomorphous Replacement ◽  
Atom Localization ◽  
Resolution Structure

We are using electron diffraction to characterize a platinum-containing derivative of bacteriorhodopsin, the light-driven proton pump from Halobacterium halobium. This has been undertaken with the dual aims of: 1)using the method of multiple heavy atom isomorphous replacement to obtain high resolution structural information about the protein, and 2)locating heavy atom labelled amino acids in the structure in order to correlate the recently determined sequence with the structural map. A necessary first step in such studies is the location of the heavy atoms in the low resolution structure. This report focusses on ways of dealing with the inherent statistical uncertainties encountered in this heavy atom localization.

Isomorphous replacement in electron diffraction of wet crystals of rat hemoglobin

1983 ◽  
Vol 41 ◽  
pp. 446-447
Author(s):  
William F. Tivol ◽  
Murray Vernon King ◽  
D. F. Parsons
Keyword(s):  
Electron Diffraction ◽  
Heavy Atom ◽  
Isomorphous Substitution ◽  
X Ray Diffraction ◽  
Salt Solutions ◽  
X Ray ◽  
Isomorphous Replacement ◽  
Structure Factors ◽  
Diffraction Structure ◽  
The Mean

Feasibility of isomorphous substitution in electron diffraction is supported by a calculation of the mean alteration of the electron-diffraction structure factors for hemoglobin crystals caused by substituting two mercury atoms per molecule, following Green, Ingram & Perutz, but with allowance for the proportionality of f to Z3/4 for electron diffraction. This yields a mean net change in F of 12.5%, as contrasted with 22.8% for x-ray diffraction.Use of the hydration chamber in electron diffraction opens prospects for examining many proteins that yield only very thin crystals not suitable for x-ray diffraction. Examination in the wet state avoids treatments that could cause translocation of the heavy-atom labels or distortion of the crystal. Combined with low-fluence techniques, it enables study of the protein in a state as close to native as possible.We have undertaken a study of crystals of rat hemoglobin by electron diffraction in the wet state. Rat hemoglobin offers a certain advantage for hydration-chamber work over other hemoglobins in that it can be crystallized from distilled water instead of salt solutions.

A Base Specific Single Heavy Atom Stain for Electron Microscopy

1972 ◽  
Vol 30 ◽  
pp. 184-185
Author(s):  
J. P. Langmore ◽  
N. R. Cozzarelli ◽  
A. V. Crewe
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
High Resolution ◽  
Elastic Scattering ◽  
Heavy Atom ◽  
High Vacuum ◽  
Heavy Atoms ◽  
Large Movement ◽  
Base Sequencing ◽  
Scanning Electron

A system has been developed to allow highly specific derivatization of the thymine bases of DNA with mercurial compounds wich should be visible in the high resolution scanning electron microscope. Three problems must be completely solved before this staining system will be useful for base sequencing by electron microscopy: 1) the staining must be shown to be highly specific for one base, 2) the stained DNA must remain intact in a high vacuum on a thin support film suitable for microscopy, 3) the arrangement of heavy atoms on the DNA must be determined by the elastic scattering of electrons in the microscope without loss or large movement of heavy atoms.

The crystal structure of myoglobin. VI. Seal myoglobin

1960 ◽  
Vol 258 (1293) ◽  
pp. 181-201 ◽  
Keyword(s):  
Tertiary Structure ◽  
Heavy Atom ◽  
Three Dimensional ◽  
Sperm Whale ◽  
Fourier Synthesis ◽  
Heavy Atoms ◽  
Isomorphous Replacement ◽  
Unit Cells ◽  
The Common ◽  
Sperm Whale Myoglobin

Myoglobin from the common seal ( Phoca vitulina ) when crystallized from ammonium sulphate forms monoclinic crystals with space group the unit cell, a = 57·9Å, b = 29·6Å, c = 106·4Å, β = 102°15', contains four molecules. The method of isomorphous replacement has been used in an investigation of the centrosymmetric b -axis projection in which it has been possible to determine signs for nearly all the h0l reflexions having spacings greater than 4Å. Three independent heavy-atom derivatives were employed and the signs so determined have been used to compute a map of the electron density projected on the (010) plane. This projection has been interpreted in terms of the molecule of sperm-whale myoglobin, as deduced by Bodo, Dintzis, Kendrew & Wyckoff (1959) from a three-dimensional Fourier synthesis to 6Å resolution. The results of the interpretation show that the two myoglobin molecules are very similar in form (tertiary structure) in spite of the differences in their amino-acid composition. The relative orientation of the two unit cells with respect to the myoglobin molecule is given and a comparison is made of the positions of the heavy atoms in each molecule.

scholarly journals How electrostatic networks modulate specificity and stability of collagen

2018 ◽  
Vol 115 (24) ◽  
pp. 6207-6212 ◽  
Author(s):  
Hongning Zheng ◽  
Cheng Lu ◽  
Jun Lan ◽  
Shilong Fan ◽  
Vikas Nanda ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Hydrogen Bonding ◽  
High Resolution ◽  
Predictive Model ◽  
Self Assembly ◽  
Structural Information ◽  
Atomistic Simulations ◽  
Specific Interactions ◽  
Collagen Stability ◽  
Resolution Structure

One-quarter of the 28 types of natural collagen exist as heterotrimers. The oligomerization state of collagen affects the structure and mechanics of the extracellular matrix, providing essential cues to modulate biological and pathological processes. A lack of high-resolution structural information limits our mechanistic understanding of collagen heterospecific self-assembly. Here, the 1.77-Å resolution structure of a synthetic heterotrimer demonstrates the balance of intermolecular electrostatics and hydrogen bonding that affects collagen stability and heterospecificity of assembly. Atomistic simulations and mutagenesis based on the solved structure are used to explore the contributions of specific interactions to energetics. A predictive model of collagen stability and specificity is developed for engineering novel collagen structures.

scholarly journals Features of the secondary structure of a protein molecule from powder diffraction data

2010 ◽  
Vol 66 (7) ◽  
pp. 756-761 ◽  
Author(s):  
Sebastian Basso ◽  
Céline Besnard ◽  
Jonathan P. Wright ◽  
Irene Margiolaki ◽  
Andrew Fitch ◽  
...  
Keyword(s):  
Secondary Structure ◽  
Powder Diffraction ◽  
De Novo ◽  
Structural Information ◽  
Heavy Atom ◽  
Protein Molecule ◽  
Lattice Parameter ◽  
Egg White Lysozyme ◽  
Replacement Method ◽  
Isomorphous Replacement

Protein powder diffraction is shown to be suitable for obtainingde novosolutions to the phase problem at low resolutionviaphasing methods such as the isomorphous replacement method. Two heavy-atom derivatives (a gadolinium derivative and a holmium derivative) of the tetragonal form of hen egg-white lysozyme were crystallized at room temperature. Using synchrotron radiation, high-quality powder patterns were collected in which pH-induced anisotropic lattice-parameter changes were exploited in order to reduce the challenging and powder-specific problem of overlapping reflections. The phasing power of two heavy-atom derivatives in a multiple isomorphous replacement analysis enabled molecular structural information to be obtained up to approximately 5.3 Å resolution. At such a resolution, features of the secondary structure of the lysozyme molecule can be accurately located using programs dedicated to that effect. In addition, the quoted resolution is sufficient to determine the correct hand of the heavy-atom substructure which leads to an electron-density map representing the protein molecule of proper chirality.

Effect of diffraction crystallography on HREM

2003 ◽  
Vol 218 (4) ◽  
Author(s):  
Fang-hua Li
Keyword(s):  
Image Processing ◽  
High Resolution ◽  
Electron Diffraction ◽  
Diffraction Data ◽  
Heavy Atom ◽  
Empirical Method ◽  
Crystal Defects ◽  
Electron Diffraction Data ◽  
Crystal Thickness ◽  
Resolution Electron

AbstractA simple image contrast theory in high-resolution electron microscopy (HREM) is introduced to demonstrate that below a certain critical crystal thickness the intensity of the Scherzer focus image is linear to the projected potential of an artificial crystal that is isomorphic to the examined one. It has become the theoretical base of electron crystallographic image processing techniques relying on the weak-phase-object approximation and kinematical diffraction. Two techniques of image processing are introduced. One of them aims at determining crystal structures by combining electron diffraction data and applying diffraction analysis methods. To reduce various kinds of electron diffraction intensity distortion the diffraction data are corrected by means of an empirical method set up by referring to the heavy atom method and Wilson statistic. The other one aims at revealing crystal defects at atomic resolution from the image taken with a medium-voltage field-emission high-resolution electron microscope. The dynamical effect is corrected by forcing the integral amplitudes of reflections in the diffractogram of image equal to the amplitudes of corresponding structure factors for the perfect crystal. The principle of the two techniques is briefly introduced, and examples of applications to crystal structure and defect determination are given.

Measurement and reduction of damage in frozen hydrated crystalline specimens

1978 ◽  
Vol 36 (2) ◽  
pp. 10-11
Author(s):  
Robert M. Glaeser ◽  
Steven B. Hayward
Keyword(s):  
High Resolution ◽  
Electron Diffraction ◽  
Diffraction Pattern ◽  
Electron Diffraction Pattern ◽  
Cell Membranes ◽  
Structural Information ◽  
Protein Crystals ◽  
Biological Macromolecules ◽  
Diffraction Intensity ◽  
Specimen Material

Highly ordered or crystalline biological macromolecules become severely damaged and disordered after a brief electron exposure, as may be seen by observing the fading and loss of the specimen's electron diffraction pattern. Loss of the diffraction pattern intensity has, in turn, a one-to-one relationship with a loss of the possibility to see structural information in the image. The actual electron exposure that results in a significant decrease in the diffraction intensity will depend first of all upon the resolution (Bragg spacing) involved, and in some cases upon the chemical make-up and composition of the specimen material. For high resolution features (in the range 3Å to 5Å resolution) of specimens such as protein crystals and cell membranes, the structure can become damaged and disordered after an exposure of about 1 electron/Å2 or less. Roughly speaking, this exposure is about 104 times lower than that which is required to produce a statistically defined image at high resolution.

Clusters: The New Markers

1982 ◽  
Vol 40 ◽  
pp. 364-365
Author(s):  
J. F. Hainfeld ◽  
J. S. Wall
Keyword(s):  
High Resolution ◽  
Image Enhancement ◽  
Beam Energy ◽  
Periodic Structures ◽  
Heavy Atom ◽  
High Dose ◽  
Protein Mass ◽  
Heavy Atoms ◽  
Atom Motion

For several years high resolution STEMS have easily visualized single heavy atoms. Attempts to mark biomolecules by covalently linking reactive heavy atom compounds have not been totally successful due to: 1) high dose - 100-3000 electrons/Å2 are necessary to see a single heavy atom, causing much damage to the biomolecule, 2) atom motion - beam energy is high enough to break bonds and form ions, thus allowing the atoms to hop away from their native positions, 3) background heavy atoms confuse the interpretation, 4) single heavy atoms on top of a protein mass have reduced contrast and are often not clearly visible. Although image enhancement procedures and averaging of periodic structures may help, some of the above problems usually remain.

Electron-diffraction patterns from frozen hydrate protein microcrystals: A new tool instructural studies

1994 ◽  
Vol 52 ◽  
pp. 102-103
Author(s):  
Christoph Burmester ◽  
Kenneth C. Holmes ◽  
Rasmus R. Schröder
Keyword(s):  
Electron Diffraction ◽  
Diffraction Data ◽  
Heavy Atom ◽  
Atomic Resolution ◽  
Protein Crystals ◽  
Electron Diffraction Data ◽  
Phase Information ◽  
Isomorphous Replacement ◽  
Diffraction Patterns

Electron crystallography of 2D protein crystals can yield models with atomic resolution by taking Fourier amplitudes from electron diffraction and phase information from processed images. Imaging at atomic resolution is more difficult than the recording of corresponding electron diffraction patterns. Therefore attempts have been made to recover phase information from diffraction data from 2-D and 3-D crystals by the method of isomorphous replacement using heavy atom labelled protein crystals. These experiments, however, have so far not produced usable phase information, partly because of the large experimental error in the spot intensities. Here we present electron diffraction data obtained from frozen hydrated 3-D protein crystals with an energy-filter microscope and a specially constructed Image Plate scanner which are of considerably better crystallographic quality (as evidenced in much smaller values for the crystallographic R-factors Rsym and Rmerge) than those reported before. The quality of this data shows that the method of isomorphous replacement could indeed be used for phase determination for diffraction data obtained from 3-D microcrystals by electron diffraction.

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