scholarly journals Serial Femtosecond X-Ray Diffraction of HIV-1 Gag MA-IP6 Microcrystals at Ambient Temperature

2019 ◽  
Vol 20 (7) ◽  
pp. 1675 ◽  
Author(s):  
Halil I Ciftci ◽  
Raymond G Sierra ◽  
Chun Yoon ◽  
Zhen Su ◽  
Hiroshi Tateishi ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
High Resolution ◽  
Ambient Temperature ◽  
X Rays ◽  
Assembly Process ◽  
Gag Protein ◽  
Crystal Forms ◽  
X Ray ◽  
X Ray Crystallography ◽  
Hiv 1

The Human immunodeficiency virus-1 (HIV-1) matrix (MA) domain is involved in the highly regulated assembly process of the virus particles that occur at the host cell’s plasma membrane. High-resolution structures of the MA domain determined using cryo X-ray crystallography have provided initial insights into the possible steps in the viral assembly process. However, these structural studies have relied on large and frozen crystals in order to reduce radiation damage caused by the intense X-rays. Here, we report the first X-ray free-electron laser (XFEL) study of the HIV-1 MA domain’s interaction with inositol hexaphosphate (IP6), a phospholipid headgroup mimic. We also describe the purification, characterization and microcrystallization of two MA crystal forms obtained in the presence of IP6. In addition, we describe the capabilities of serial femtosecond X-ray crystallography (SFX) using an XFEL to elucidate the diffraction data of MA-IP6 complex microcrystals in liquid suspension at ambient temperature. Two different microcrystal forms of the MA-IP6 complex both diffracted to beyond 3.5 Å resolution, demonstrating the feasibility of using SFX to study the complexes of MA domain of HIV-1 Gag polyprotein with IP6 at near-physiological temperatures. Further optimization of the experimental and data analysis procedures will lead to better understanding of the MA domain of HIV-1 Gag and IP6 interaction at high resolution and will provide basis for optimization of the lead compounds for efficient inhibition of the Gag protein recruitment to the plasma membrane prior to virion formation.

scholarly journals Serial femtosecond X-ray diffraction of HIV-1 Gag MA-IP6 microcrystals at ambient temperature

2019 ◽  
Author(s):  
Halil I Ciftci ◽  
Raymond G Sierra ◽  
Chun Hong Yoon ◽  
Zhen Su ◽  
Hiroshi Tateishi ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
High Resolution ◽  
Ambient Temperature ◽  
X Rays ◽  
Assembly Process ◽  
Gag Protein ◽  
Crystal Forms ◽  
X Ray ◽  
X Ray Crystallography ◽  
Hiv 1

AbstractThe Human immunodeficiency virus-1 (HIV-1) matrix (MA) domain is involved in the highly regulated assembly process of the virus particles that occur at the host cell’s plasma membrane. High-resolution structures of the MA domain determined using cryo X- ray crystallography have provided initial insights into the possible steps in the viral assembly process. However, these structural studies have relied on large and frozen crystals in order to reduce radiation damage caused by the intense X-rays. Here, we report the first XFEL study of the HIV-1 MA domain’s interaction with inositol hexaphosphate (IP6), a phospholipid headgroup mimic. We also describe the purification, characterization and microcrystallization of two MA crystal forms obtained in the presence of IP6. In addition, we describe the capabilities of serial femtosecond X-ray crystallography (SFX) using X-ray free-electron laser (XFEL) to elucidate the diffraction data of MA-IP6 complex microcrystals in liquid suspension at ambient temperature. Two different microcrystal forms of MA-IP6 complex both diffracted to beyond 3.5 Å resolution, demonstrating the feasibility of using SFX to study the complexes of MA domain of HIV-1 Gag polyprotein with IP6 at near-physiological temperatures. Further optimization of the experimental and data analysis procedures will lead to better understanding of the MA domain of HIV-1 Gag and IP6 interaction at high resolution and provide basis for optimization of the lead compounds for efficient inhibition of the Gag protein recruitment to the plasma membrane prior to virion formation.

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.

Three-dimensional crystallization of membrane proteins

1988 ◽  
Vol 21 (4) ◽  
pp. 429-477 ◽  
Author(s):  
W. Kühlbrandt
Keyword(s):  
High Resolution ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Protein Complexes ◽  
Three Dimensional ◽  
Biological Macromolecules ◽  
X Ray ◽  
X Ray Crystallography ◽  
Membrane Protein Complexes ◽  
Essential Prerequisite

As recently as 10 years ago, the prospect of solving the structure of any membrane protein by X-ray crystallography seemed remote. Since then, the threedimensional (3-D) structures of two membrane protein complexes, the bacterial photosynthetic reaction centres of Rhodopseudomonas viridis (Deisenhofer et al. 1984, 1985) and of Rhodobacter sphaeroides (Allen et al. 1986, 1987 a, 6; Chang et al. 1986) have been determined at high resolution. This astonishing progress would not have been possible without the pioneering work of Michel and Garavito who first succeeded in growing 3-D crystals of the membrane proteins bacteriorhodopsin (Michel & Oesterhelt, 1980) and matrix porin (Garavito & Rosenbusch, 1980). X-ray crystallography is still the only routine method for determining the 3-D structures of biological macromolecules at high resolution and well-ordered 3-D crystals of sufficient size are the essential prerequisite.

Cryogenic high-resolution X-ray spectrometers for SR-XRF and microanalysis

1998 ◽  
Vol 5 (3) ◽  
pp. 515-517 ◽  
Author(s):  
M. Frank ◽  
C. A. Mears ◽  
S. E. Labov ◽  
L. J. Hiller ◽  
J. B. le Grand ◽  
...  
Keyword(s):  
High Resolution ◽  
Energy Resolution ◽  
Superconducting Tunnel Junction ◽  
Semiconductor Detectors ◽  
X Rays ◽  
X Ray ◽  
Sr Xrf ◽  
Demonstration Experiment ◽  
Near Future ◽  
Stj Detector

Experimental results are presented obtained with a cryogenically cooled high-resolution X-ray spectrometer based on a 141 × 141 µm Nb-Al-Al2O3-Al-Nb superconducting tunnel junction (STJ) detector in an SR-XRF demonstration experiment. STJ detectors can operate at count rates approaching those of semiconductor detectors while still providing a significantly better energy resolution for soft X-rays. By measuring fluorescence X-rays from samples containing transition metals and low-Z elements, an FWHM energy resolution of 6–15 eV for X-rays in the energy range 180–1100 eV has been obtained. The results show that, in the near future, STJ detectors may prove very useful in XRF and microanalysis applications.

Applications of High-Resolution Powder X-Ray Diffraction

2007 ◽  
Vol 130 ◽  
pp. 7-14 ◽  
Author(s):  
Andrew N. Fitch
Keyword(s):  
High Resolution ◽  
Synchrotron Radiation ◽  
Powder Diffraction ◽  
X Rays ◽  
X Ray Diffraction ◽  
Crystalline Materials ◽  
X Ray ◽  
Structural Characterisation ◽  
Pdf Analysis

The highly-collimated, intense X-rays produced by a synchrotron radiation source can be harnessed to build high-resolution powder diffraction instruments with a wide variety of applications. The general advantages of using synchrotron radiation for powder diffraction are discussed and illustrated with reference to the structural characterisation of crystalline materials, atomic PDF analysis, in-situ and high-throughput studies where the structure is evolving between successive scans, and the measurement of residual strain in engineering components.

scholarly journals Structure of a 14-3-3 Coordinated Hexamer of the Plant Plasma Membrane H+-ATPase by Combining X-Ray Crystallography and Electron Cryomicroscopy

2007 ◽  
Vol 25 (3) ◽  
pp. 427-440 ◽  
Author(s):  
Christian Ottmann ◽  
Sergio Marco ◽  
Nina Jaspert ◽  
Caroline Marcon ◽  
Nicolas Schauer ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Electron Cryomicroscopy ◽  
X Ray ◽  
X Ray Crystallography ◽  
Plant Plasma Membrane

HIGH-RESOLUTION PIXE USING BRAGG’S SPECTROMETER

1991 ◽  
Vol 01 (03) ◽  
pp. 251-258 ◽  
Author(s):  
M. TERASAWA
Keyword(s):  
High Resolution ◽  
Heavy Ions ◽  
X Rays ◽  
Light Elements ◽  
Element Analysis ◽  
X Ray ◽  
Peak Intensity ◽  
Moderate Energy ◽  
Practical Analysis ◽  
Wavelength Selectivity

K, L, and M X-rays in the wavelengths between 6Å and 130Å generated by the bombardment of 200 keV protons and other heavy ions were measured by means of a wavelength dispersive Bragg’s spectrometer. The X-ray peak intensity was fairly high in general, while the background was very low. The technique was favorably applied to a practical analysis of several light elements (Be, B, C, N, O, and F). Use of moderate-energy heavy ions considering the wavelength selectivity in X-ray generation was effective for the element analysis. The high-resolution spectrometry in the analytical application of ion-induced X-ray generation was found to be useful for the study of fine electronic structure, e.g. satellite and hypersatellite X-ray study, and of the chemical state of materials.

scholarly journals Mechanism of IPPase shown by high resolution Neutron and X-ray crystallography

2014 ◽  
Vol 70 (a1) ◽  
pp. C1211-C1211
Author(s):  
Joseph Ng ◽  
Ronny Hughes ◽  
Michelle Morris ◽  
Leighton Coates ◽  
Matthew Blakeley ◽  
...  
Keyword(s):  
High Resolution ◽  
Active Site ◽  
Inorganic Pyrophosphatase ◽  
Inorganic Pyrophosphate ◽  
X Ray ◽  
X Ray Crystallography ◽  
Cellular Processes ◽  
Crystallographic Structures ◽  
Hydrolysis Of ◽  
Low Energy Conformations

Soluble inorganic pyrophosphatase (IPPase) catalyzes the hydrolysis of inorganic pyrophosphate (PPi) to form orthophosphate (Pi). The action of this enzyme shifts the overall equilibrium in favor of synthesis during a number of ATP-dependent cellular processes such as in the polymerization of nucleic acids, production of coenzymes and proteins and sulfate assimilation pathways. Two Neutron crystallographic (2.10-2.50Å) and five high-resolution X-ray (0.99Å-1.92Å) structures of the archaeal IPPase from Thermococcus thioreducens have been determined under both cryo and room temperatures. The structures determined include the recombinant IPPase bound to Mg+2, Ca+2, Br-, SO2-2 or PO4-2 involving those with non-hydrolyzed and hydrolyzed pyrophosphate complexes. All the crystallographic structures provide snapshots of the active site corresponding to different stages of the hydrolysis of inorganic pyrophosphate. As a result, a structure-based model of IPPase catalysis is devised showing the enzyme's low-energy conformations, hydration states, movements and nucleophile generation within the active site.

scholarly journals Micrometer-resolution imaging using MÖNCH: towards G2-less grating interferometry

2016 ◽  
Vol 23 (6) ◽  
pp. 1462-1473 ◽  
Author(s):  
Sebastian Cartier ◽  
Matias Kagias ◽  
Anna Bergamaschi ◽  
Zhentian Wang ◽  
Roberto Dinapoli ◽  
...  
Keyword(s):  
High Resolution ◽  
Silicon Detector ◽  
Limiting Factor ◽  
X Rays ◽  
X Ray ◽  
Charge Cloud ◽  
High Resolution Images ◽  
Resolution Imaging ◽  
Hybrid Silicon ◽  
Small Pixel

MÖNCH is a 25 µm-pitch charge-integrating detector aimed at exploring the limits of current hybrid silicon detector technology. The small pixel size makes it ideal for high-resolution imaging. With an electronic noise of about 110 eV r.m.s., it opens new perspectives for many synchrotron applications where currently the detector is the limiting factor,e.g.inelastic X-ray scattering, Laue diffraction and soft X-ray or high-resolution color imaging. Due to the small pixel pitch, the charge cloud generated by absorbed X-rays is shared between neighboring pixels for most of the photons. Therefore, at low photon fluxes, interpolation algorithms can be applied to determine the absorption position of each photon with a resolution of the order of 1 µm. In this work, the characterization results of one of the MÖNCH prototypes are presented under low-flux conditions. A custom interpolation algorithm is described and applied to the data to obtain high-resolution images. Images obtained in grating interferometry experiments without the use of the absorption grating G2are shown and discussed. Perspectives for the future developments of the MÖNCH detector are also presented.

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