Reinforced protein crystals (original) (raw)
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Growth of Ultrastable Protein–Silica Composite Crystals
Crystal Growth & Design, 2013
Protein crystals were obtained in a wide range of silica gel concentrations, 2.0−22.0% (v/v), using the counter-diffusion technique. The protein crystal lattice incorporates silica fibers during their growth, making the crystal appear optically translucent while maintaining the diffraction quality. The effect of the silica fibers on the nucleation and growth morphology is discussed, and the amount of incorporated silica matrix is quantified. The practical implications of the presence of a high hygroscope phase on the crystal properties are discusse, and the improvement of the mechanical properties and stability of the crystals is shown. These reinforced protein crystals, able to include large amounts of silica, open a new range of possibilities for the characterization of protein crystals and the application in the biotechnological industry.
Protein crystal quality studies using rod-shaped crystals
Journal of Crystal Growth, 1996
Lysozyme single crystals were grown into X-ray capillaries to a size larger than the capillary diameter thus filling it. The two ends of the same crystal grow at different rates, the difference being at least one order of magnitude. These rod-shaped crystals allow ideal diffraction experiments to test crystal quality as a function of the growth rate. In situ X-ray diffraction experiments were carded out using the capillary where the crystal grew. Oscillation pictures yield different values of the maximum resolution level, ranging from 2.5 to 1.2 A for the opposite ends of the crystal suggesting a large influence of growth rate on protein crystal quality. * Corresponding
Growth and Characterization of High-quality Protein Crystals for X-ray Crystallography
Annals of the New York Academy of Sciences, 2009
Tetragonal hen egg white lysozyme is grown by the batch method in solution and gel media to study the influence of high magnetic fields on the quality of macromolecular crystals. The crystallographic quality of crystals grown in the absence and in the presence of 7-and 10-T fields are analyzed in terms of mosaicity and high-resolution X-ray imaging methods. Crystals grown by the batch method from solution showed a remarkable enhancement of the crystallographic quality, although the overall crystal quality was higher for gel-grown crystals than solution-grown crystals. The observed improvement in crystal quality can be attributed to the suppression of convective transport during the crystal growth process and the control of the nucleation kinetics by the use of a magnetic force.
Generation of Size-Controlled, Submicrometer Protein Crystals
Chemistry of Materials, 2005
Cross-linked protein crystals represent a new class of micro-and mesoporous materials with properties potentially useful in technologies as diverse as chiral separations and drug delivery. These applications require the generation of protein crystals in formats and sizes quite distinct from the traditional submillimeter single crystals favored by structural biologists. Here we report a high-yield method for forming monodisperse protein crystals with tunable dimensions ranging from 250 nm up to tens of micrometers. X-ray powder diffraction of these crystallites confirms that the hen egg white lysozyme crystals are identical in structure to that of the bulk tetragonal lysozyme (a) 79.05 Å; c) 37.96 Å).
Crystal Growth & Design, 2016
Major discoveries in structural biology depend on obtaining well-diffracting macromolecular crystals. This necessity has motivated many fundamental studies on protein crystallization using lysozyme as a model system. In the present contribution, we report the unprecedented observation of lysozyme crystals that stop dissolving under undersaturated conditions imposed to sub-microliter crystallization drops at mild temperatures. Subsequent growth of the same crystals is apparently undisturbed after the drops are cooled below the saturation temperature. The succession of heating/cooling cycles only partially recovers crystal dissolution while crystal growth becomes gradually slower. Ultimately, increasing and decreasing the temperature between 10 and 37 °C has no visible effect on the size of the crystals. We ascribe this phenomenon to the partial denaturation of the soluble protein in the drop as evidenced by the decreasing glycoside hydrolase activity of lysozyme over the incubation time. The disturbances in the phase transition processes are explained as the result of the changed chemical potential due to different folding states. In a time when the high-hanging fruits in structural biology have to be picked, the present findings call attention to interfacial phenomena as an important, though often imperceptible, aspect that affects protein stability and justifies further optimization of current crystallization methods.
Growth of shaped single crystals of proteins
Journal of Crystal Growth, 1996
We present a procedure for obtaining protein single crystals that fill the capillary tubes in which they grow. The implementation was typical of the gel acupuncture method and the four different proteins are used as examples: lysozyme (HEW), thaumatin I, ferritin and insulin. Rod-and prismatic-shaped protein single crystals of these four proteins were grown inside capillary tubes of 0.2, 0.3, 0.5 mm in diameter and, for the case of lysozyme, up to 1.2 mm in diameter. The maximum length measured along the long axes of the rod crystals was 1.6 mm again for lysozyme crystals. It was observed that, once the capillary tube was filled, the crystal continues to grow by diffusion of the precipitating agent throughout the porous network formed by the protein crystal structure. We also discuss the possibility of growing these cylinders of crystalline proteins by the addition of protein solution to the mother liquor through the upper end of the glass capillary while the precipitating agent diffuses through the protein crystal itself. X-ray diffraction patterns confirm the single crystal character of the protein rods.
High Resolution Imaging as a Characterization Tool for Biological Crystals
Annals of the New York Academy of Sciences, 2004
A BSTRACT : Biomolecular crystals consist of large unit cells that form a rather flexible medium that is able to accommodate a certain degree of lattice distortion, leading to several interesting issues ranging from structural to physical properties. Several techniques, from X-ray diffraction to microscopy, have been adapted to study the structural and physical properties of biomolecular crystals systematically. The use of synchrotron-based monochromatic X-ray diffraction topography, with triple axis diffractometry and rocking curve measurements, to characterize biomolecular crystals is reviewed. Recent X-ray diffraction images from gel and solution grown lysozyme crystals are presented. Defect structures in these crystals are discussed, together with reciprocal space mapping, and compared with results obtained from crystals grown in a low gravity environment.
Acta Crystallographica Section D Biological Crystallography, 2005
A microscope for quantitative analysis of the birefringence properties of samples is introduced. The microscope is used to measure variations in the slow optical axis position (SOAP) across hen egg-white lysozyme, glucose isomerase and ®bronectin crystals. By comparing these variations with indicators of diffraction quality, it is shown that the optical properties of a protein crystal provide a non-invasive method of determining crystal diffraction quality before any X-ray data collection is attempted.
ACS Omega, 2018
The well-known difficulty to obtain high-quality protein crystals has motivated researchers to come up with new methods or modifications of established crystallization methods to stimulate the growth of good diffracting crystals. In the present work, a new approach, using a protein thin film organized by external electric field (EEF) as a template for protein crystal growth, is introduced. This method increased nucleation of hen egg white lysozyme (HEWL) in comparison with the classical vapor diffusion method, besides improving crystal morphology and size. X-ray diffraction analyses indicated improvements in crystal quality. When HEWL was crystallized at pH 6.2, in which this protein presents biological activity, the control crystal presented a poorly ordered crystalline structure and a low resolution cutoff at 3.42 Å, whereas the crystal grown with the EEF protein film revealed a high-resolution limit at 1.67 Å. These results suggest that protein films organized by EEF may improve protein crystals and their data quality.
Nucleation and Crystallization of Lysozyme: Role of Substrate Surface Chemistry and Topography
In an effort to better understand, direct and control the crystallization of molecular and macromolecular compounds, an approach using colloidal templates as substrates for heterogeneous nucleation was investigated. These templates combine both tuneable chemical functionalities and geometrical features, altering the crystal-substrate interactions. Colloidal templates were prepared from silica nanoparticles, where the surface chemistry was modified by silanization. Particle size varied from 30 to 700 nm, with silanols, NH 2 , CF 3 , phenyl, or dodecyl as surface functional groups. Here, we report on the template assisted crystallization of chicken egg white lysozyme (CEWL). Nucleation was dramatically affected by the surface chemistry and topography of the templates. Using 220 nm particles, hydrophobic templates generally produced fewer, larger crystals, while a larger number of small crystals were obtained on hydrophilic templates. The use of different particle sizes also affected the crystal size, the optimal for nucleation being 432 nm. Classical Nucleation Theory (CNT) can interpret surface chemistry effects but does not support the effect of particle size. This paper reports that the combined use of both geometrical and chemical interactions results in an increased ability to control the nucleation and growth of protein crystals.