Nanocrystalline protein domains via salting-out

2021 ◽  
Vol 77 (11) ◽  
Author(s):  
Daniel G. Greene ◽  
Shannon Modla ◽  
Stanley I. Sandler ◽  
Norman J. Wagner ◽  
Abraham M. Lenhoff
Keyword(s):  
Generation X ◽  
Protein Structure Determination ◽  
Biological Macromolecules ◽  
Bulk Crystals ◽  
Salting Out ◽  
X Ray ◽  
Mesoscale Structure ◽  
Protein Gels ◽  
Amorphous Structures ◽  
The Rich

Protein salting-out is a well established phenomenon that in many cases leads to amorphous structures and protein gels, which are usually not considered to be useful for protein structure determination. Here, microstructural measurements of several different salted-out protein dense phases are reported, including of lysozyme, ribonuclease A and an IgG1, showing that salted-out protein gels unexpectedly contain highly ordered protein nanostructures that assemble hierarchically to create the gel. The nanocrystalline domains are approximately 10–100 nm in size, are shown to have structures commensurate with those of bulk crystals and grow on time scales in the order of an hour to a day. Beyond revealing the rich, hierarchical nanoscale to mesoscale structure of protein gels, the nanocrystals that these phases contain are candidates for structural biology on next-generation X-ray free-electron lasers, which may enable the study of biological macromolecules that are difficult or impossible to crystallize in bulk.

scholarly journals Electrospray sample injection for single-particle imaging with x-ray lasers

2019 ◽  
Vol 5 (5) ◽  
pp. eaav8801 ◽  
Author(s):  
Johan Bielecki ◽  
Max F. Hantke ◽  
Benedikt J. Daurer ◽  
Hemanth K. N. Reddy ◽  
Dirk Hasse ◽  
...  
Keyword(s):  
Generation X ◽  
Room Temperature ◽  
Protein Structure Determination ◽  
High Temporal Resolution ◽  
X Ray ◽  
Free Sample ◽  
Large Particles ◽  
Particle Imaging ◽  
Single Particle Imaging ◽  
Initial Droplet Size

The possibility of imaging single proteins constitutes an exciting challenge for x-ray lasers. Despite encouraging results on large particles, imaging small particles has proven to be difficult for two reasons: not quite high enough pulse intensity from currently available x-ray lasers and, as we demonstrate here, contamination of the aerosolized molecules by nonvolatile contaminants in the solution. The amount of contamination on the sample depends on the initial droplet size during aerosolization. Here, we show that, with our electrospray injector, we can decrease the size of aerosol droplets and demonstrate virtually contaminant-free sample delivery of organelles, small virions, and proteins. The results presented here, together with the increased performance of next-generation x-ray lasers, constitute an important stepping stone toward the ultimate goal of protein structure determination from imaging at room temperature and high temporal resolution.

scholarly journals Electrospray sample injection for single-particle imaging with X-ray lasers

2018 ◽  
Author(s):  
Johan Bielecki ◽  
Max F. Hantke ◽  
Benedikt J. Daurer ◽  
Hemanth K. N. Reddy ◽  
Dirk Hasse ◽  
...  
Keyword(s):  
Generation X ◽  
Room Temperature ◽  
Protein Structure Determination ◽  
High Temporal Resolution ◽  
X Ray ◽  
Free Sample ◽  
Large Particles ◽  
Particle Imaging ◽  
Single Particle Imaging ◽  
Initial Droplet Size

The possibility of imaging single proteins constitutes an exciting challenge for X-ray lasers. Despite encouraging results on large particles, imaging small particles has proven to be difficult for two reasons: not quite high enough pulse intensity from currently available X-ray lasers and, as we demonstrate here, contamination of the aerosolised molecules by non-volatile contaminants in the solution. The amount of contamination on the sample depends on the initial droplet-size during aerosolisation. Here we show that with our electrospray injector we can decrease the size of aerosol droplets and demonstrate virtually contaminant-free sample delivery of organelles, small virions, and proteins. The results presented here, together with the increased performance of next generation X-ray lasers, constitute an important stepping stone towards the ultimate goal of protein structure determination from imaging at room temperature and high temporal resolution.

scholarly journals A public database of macromolecular diffraction experiments

2016 ◽  
Vol 72 (11) ◽  
pp. 1181-1193 ◽  
Author(s):  
Marek Grabowski ◽  
Karol M. Langner ◽  
Marcin Cymborowski ◽  
Przemyslaw J. Porebski ◽  
Piotr Sroka ◽  
...  
Keyword(s):  
Experimental Data ◽  
Diffraction Data ◽  
Distributed Storage ◽  
Protein Structure Determination ◽  
Data Bank ◽  
Scientific Integrity ◽  
Biological Macromolecules ◽  
Data Sets ◽  
X Ray ◽  
Unrestricted Access

The low reproducibility of published experimental results in many scientific disciplines has recently garnered negative attention in scientific journals and the general media. Public transparency, including the availability of `raw' experimental data, will help to address growing concerns regarding scientific integrity. Macromolecular X-ray crystallography has led the way in requiring the public dissemination of atomic coordinates and a wealth of experimental data, making the field one of the most reproducible in the biological sciences. However, there remains no mandate for public disclosure of the original diffraction data. The Integrated Resource for Reproducibility in Macromolecular Crystallography (IRRMC) has been developed to archive raw data from diffraction experiments and, equally importantly, to provide related metadata. Currently, the database of our resource contains data from 2920 macromolecular diffraction experiments (5767 data sets), accounting for around 3% of all depositions in the Protein Data Bank (PDB), with their corresponding partially curated metadata. IRRMC utilizes distributed storage implemented using a federated architecture of many independent storage servers, which provides both scalability and sustainability. The resource, which is accessibleviathe web portal at http://www.proteindiffraction.org, can be searched using various criteria. All data are available for unrestricted access and download. The resource serves as a proof of concept and demonstrates the feasibility of archiving raw diffraction data and associated metadata from X-ray crystallographic studies of biological macromolecules. The goal is to expand this resource and include data sets that failed to yield X-ray structures in order to facilitate collaborative efforts that will improve protein structure-determination methods and to ensure the availability of `orphan' data left behind for various reasons by individual investigators and/or extinct structural genomics projects.

Effect of Annealing temperature on the Structural and Magnetic Properties of TbxY3-xFe5O12(x = 0.0, 1.0, 2.0) Thin Films Prepared by Sol-Gel Process

2012 ◽  
Vol 501 ◽  
pp. 236-241 ◽  
Author(s):  
Ftema W. Aldbea ◽  
Noor Bahyah Ibrahim ◽  
Mustafa Hj. Abdullah ◽  
Ramadan E. Shaiboub
Keyword(s):  
Thin Films ◽  
Magnetic Properties ◽  
Room Temperature ◽  
Annealing Temperature ◽  
Sol Gel ◽  
Vibrating Sample Magnetometer ◽  
X Ray ◽  
Garnet Phase ◽  
Sol Gel Process ◽  
Amorphous Structures

Thin films nanoparticles TbxY3-xFe5O12 (x=0.0, 1.0, 2.0) were prepared by the sol-gel process followed by annealing process at various annealing temperatures of 700° C, 800° C and 900° C in air for 2 h. The results obtained from X-ray diffractometer (XRD) show that the films annealed below 900°C exhibit peaks of garnet mixed with small amounts of YFeO3 and Fe2O3. Pure garnet phase has been detected in the films annealed at 900°C. Before annealing the films show amorphous structures. The particles sizes measurement using the field emission scanning electron microscope (FE-SEM) showed that the particles sizes increased as the annealing temperature increased. The magnetic properties were measured at room temperature using the vibrating sample magnetometer (VSM). The saturation magnetization (Ms) of the films also increased with the annealing temperature. However, different behavior of coercivity (Hc) has been observed as the annealing temperature was increased.

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.

Polychromatic X-ray microdiffraction studies of mesoscale structure and dynamics

2005 ◽  
Vol 12 (2) ◽  
pp. 155-162 ◽  
Author(s):  
G. E. Ice ◽  
B. C. Larson ◽  
W. Yang ◽  
J. D. Budai ◽  
J. Z. Tischler ◽  
...  
Keyword(s):  
X Ray ◽  
Structure And Dynamics ◽  
Mesoscale Structure

A high-energy triple-axis X-ray diffractometer for the study of the structure of bulk crystals

2004 ◽  
Vol 37 (6) ◽  
pp. 901-910 ◽  
Author(s):  
C. Seitz ◽  
M. Weisser ◽  
M. Gomm ◽  
R. Hock ◽  
A. Magerl
Keyword(s):  
High Energy ◽  
X Rays ◽  
Bulk Crystals ◽  
X Ray Diffraction ◽  
X Ray ◽  
Peak Intensity ◽  
High Penetration ◽  
Massive Sample ◽  
Laue Geometry ◽  
The Ideal

A triple-axis diffractometer for high-energy X-ray diffraction is described. A 450 kV/4.5 kW stationary tungsten X-ray tube serves as the X-ray source. Normally, 220 reflections of thermally annealed Czochralski Si are employed for the monochromator and analyser. Their integrated reflectivity is about ten times higher than the ideal crystal value. With the same material as the sample, and working with the WKα line at 60 keV in symmetric Laue geometry for all axes, the full width at half-maximum (FWHM) values for the longitudinal and transversal resolution are 2.5 × 10−3and 1.1 × 10−4for ΔQ/Q, respectively, and the peak intensity for a non-dispersive setting is 3000 counts s−1. In particular, for a double-axis mode, an energy well above 100 keV from theBremsstrahlungspectrum can be used readily. High-energy X-rays are distinguished by a high penetration power and materials of several centimetre thickness can be analysed. The feasibility of performing experiments with massive sample environments is demonstrated.

scholarly journals The low-mass pre-main sequence population of Scorpius OB1

2018 ◽  
Vol 615 ◽  
pp. A148 ◽  
Author(s):  
Francesco Damiani
Keyword(s):  
Total Mass ◽  
Main Sequence ◽  
Near Infrared ◽  
Large Population ◽  
X Ray ◽  
Uv Excess ◽  
Low Mass ◽  
Ob Associations ◽  
The Rich ◽  
A New Technique

Context. The low-mass members of OB associations, expected to be a major component of their total population, are in most cases poorly studied because of the difficulty of selecting these faint stars in crowded sky regions. Our knowledge of many OB associations relies on only a relatively small number of massive members. Aims. We study here the Sco OB1 association, with the aim of a better characterization of its properties, such as global size and shape, member clusters and their morphology, age and formation history, and total mass. Methods. We use deep optical and near-infrared (NIR) photometry from the VPHAS+ and VVV surveys, over a wide area (2.6° × 2.6°), complemented by Spitzer infrared (IR) data, and Chandra and XMM-Newton X-ray data. A new technique is developed to find clusters of pre-main sequence M-type stars using suitable color-color diagrams, complementing existing selection techniques using narrow-band Hα photometry or NIR and ultraviolet (UV) excesses, and X-ray data. Results. We find a large population of approximately 4000 candidate low-mass Sco OB1 members whose spatial properties correlate well with those of Hα-emission, NIR-excess, UV-excess, and X-ray detected members, and unresolved X-ray emission. The low-mass population is spread among several interconnected subgroups: they coincide with the HII regions G345.45+1.50 and IC4628, and the rich clusters NGC 6231 and Trumpler 24, with an additional subcluster intermediate between these two. The total mass of Sco OB1 is estimated to be ~ 8500 M⊙. Indication of a sequence of star-formation events is found, from South (NGC 6231) to North (G345.45+1.50). We suggest that the diluted appearance of Trumpler 24 indicates that the cluster is now dissolving into the field, and that tidal stripping by NGC 6231 nearby contributes to the process.

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