Advanced Methods to Monitor and Control the Crystallisation Environment

A special issue of Crystals (ISSN 2073-4352). This special issue belongs to the section "Biomolecular Crystals".

Deadline for manuscript submissions: closed (1 August 2020) | Viewed by 54936

Special Issue Editors


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Guest Editor
Instituto de Química, Universidad Nacional Autónoma de México, Mexico City 04510, Mexico
Interests: protein crystals; biocrystals; crystal growth; protein crystallography; crystal chemistry; biomineralization; biomimetics; biological macromolecules
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UCIBIO, Faculdade de Ciências e Tecnologia, NOVA University of Lisbon, Department of Chemistry, 2829-516 Caparica, Portugal
Interests: structural biology; protein crystallization; macromolecular interactions; integrative methods; science outreach

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Guest Editor
UCIBIO, Faculdade de Ciências e Tecnologia, NOVA University of Lisbon, Department of Chemistry, 2829-516 Caparica, Portugal
Interests: structural biology; protein crystallization; metalloenzymes and enzymatic mechanisms; molybdoenzymes; science outreach and teaching

Special Issue Information

Dear Colleagues,

Currently, there are powerful experimental techniques for the 3D structure determination of biological macromolecules (proteins, nucleic acids, polysaccharides and their macromolecular complexes). Particularly, X-ray crystallography is one of the most important techniques in this field. This technique can reach quasi-atomic resolution in the most favorable cases. For this approach, the size and complexity of the system are not a priori limitations and this only requires high-quality single crystals.

This Special Issue on “Crystallization Under Special and Physical Environments” will not only include fundamentals for understanding the physical or chemical aspects of the crystallization process, but will also include advanced techniques for controlling the size and orientation through the utilization of electric and magnetic fields and other special environments (hydrogels, organogels, lipid cubic phases, etc.). The third part will include the crystallization of inorganic and organic compounds and proteins grown in special biological conditions, where the use of microorganisms produce crystals inside specialized cells (idioblast) that contains biforine cells that form crystals. Finally, the new trends in crystallography using techniques recently coined as the serial crystallography of macromolecular complexes (Free-lectron Lasers, XFEL) will be shortly discussed in terms of preparing nanocrystals for injection into the XFEL facilities.

Prof. Dr. Abel Moreno
Dr. Ana Luísa Carvalho
Prof. Dr. Maria João Romão
Guest Editors

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Published Papers (12 papers)

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Research

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11 pages, 2190 KiB  
Article
Vapour Diffusion Sitting Drop Method to Induce Nucleation of Calcium Phosphate on Exfoliated Graphene and Graphene Oxide Flakes
by Francisco Javier Acebedo-Martínez, Raquel Fernández-Penas, Cristóbal Verdugo-Escamilla, Duane Choquesillo-Lazarte and Jaime Gómez-Morales
Crystals 2021, 11(7), 767; https://doi.org/10.3390/cryst11070767 - 30 Jun 2021
Cited by 1 | Viewed by 2056
Abstract
The preparation of graphene/apatite and graphene oxide/apatite hybrid nanocomposites has recently attracted great attention in the biomaterial community. The sitting drop vapor diffusion technique has been assessed as a preparative method for such nanocomposites in this work. The technique has been employed to [...] Read more.
The preparation of graphene/apatite and graphene oxide/apatite hybrid nanocomposites has recently attracted great attention in the biomaterial community. The sitting drop vapor diffusion technique has been assessed as a preparative method for such nanocomposites in this work. The technique has been employed to induce heterogeneous nucleation and growth of calcium phosphate in the presence of exfoliated graphene and commercial graphene oxide flakes, both labeled with L-Alanine. Exfoliated multilayered graphene flakes were produced by sonication-assisted liquid-phase exfoliation of graphite. In both composites, the apatite nanocrystals displayed similar size and shape, but different labile and B-type carbonation contributions. Graphene and graphene oxide flakes also influenced the carbonation degree of the apatite, which was almost half that measured for the apatite blank, as well as the aggregation state of their composites. In this regard, those composites with graphene oxide formed larger aggregates because of their wider size distribution, with a high-volume percentage of nanosheets (of about 4 nm length). Overall, the method is very useful to prepare small amounts of nanocomposite with high reproducibility. Full article
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12 pages, 4872 KiB  
Article
The Influence of Silicateins on the Shape and Crystalline Habit of Silica Carbonate Biomorphs of Alkaline Earth Metals (Ca, Ba, Sr)
by Nuria Sánchez-Puig, Mayra Cuéllar-Cruz, Selene R. Islas, Juana V. Tapia-Vieyra, Roberto A. Arreguín-Espinosa and Abel Moreno
Crystals 2021, 11(4), 438; https://doi.org/10.3390/cryst11040438 - 17 Apr 2021
Cited by 5 | Viewed by 2614
Abstract
This contribution presents the effect of two ortholog enzymes from marine sponges called silicateins on the formation of silica carbonate biomorphs of alkaline metals (Ca, Ba, Sr). In vivo, these enzymes participate in the polymerization of silica. Silicateins from Tethya aurantia and Suberitis [...] Read more.
This contribution presents the effect of two ortholog enzymes from marine sponges called silicateins on the formation of silica carbonate biomorphs of alkaline metals (Ca, Ba, Sr). In vivo, these enzymes participate in the polymerization of silica. Silicateins from Tethya aurantia and Suberitis domuncula were produced recombinantly and presented different degrees of activity, as evidenced by their ability to cleave silyl ether-like bonds in a model compound. Biomorphs are typically inorganic structures that show characteristic shapes resembling those of biological structures such as helices, leaves, flowers, disks or spheres. Irrespective of the concentration or the enzyme used, the presence of silicateins inhibited the formation of classic morphologies of biomorphs, albeit to different extents. Thus, not only the silica condensation activity of the enzyme but also its ability to bind silica compounds is implicated in the inhibition process. The largest effect was observed for the strontium and barium biomorphs, leading to the formation of spheres similar to those observed in diatoms and Radiolaria rather than the classical non-symmetrical forms. Characterization of the samples using Raman spectroscopy showed that silicatein did not affect the crystalline structure of the alkaline earth metal carbonate but did modify the crystalline habit. Full article
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19 pages, 13136 KiB  
Article
Tools to Ease the Choice and Design of Protein Crystallisation Experiments
by Nicholas Rosa, Marko Ristic, Luke Thorburn, Gabriel J. Abrahams, Bevan Marshall, Christopher J. Watkins, Alex Kruger, Alex Khassapov and Janet Newman
Crystals 2020, 10(2), 95; https://doi.org/10.3390/cryst10020095 - 7 Feb 2020
Cited by 6 | Viewed by 5726
Abstract
The process of macromolecular crystallisation almost always begins by setting up crystallisation trials using commercial or other premade screens, followed by cycles of optimisation where the crystallisation cocktails are focused towards a particular small region of chemical space. The screening process is relatively [...] Read more.
The process of macromolecular crystallisation almost always begins by setting up crystallisation trials using commercial or other premade screens, followed by cycles of optimisation where the crystallisation cocktails are focused towards a particular small region of chemical space. The screening process is relatively straightforward, but still requires an understanding of the plethora of commercially available screens. Optimisation is complicated by requiring both the design and preparation of the appropriate secondary screens. Software has been developed in the C3 lab to aid the process of choosing initial screens, to analyse the results of the initial trials, and to design and describe how to prepare optimisation screens. Full article
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13 pages, 2377 KiB  
Article
On the Quality of Protein Crystals Grown under Diffusion Mass-transport Controlled Regime (I)
by José A. Gavira, Fermín Otálora, Luis A. González-Ramírez, Emilio Melero, Alexander E.S. van Driessche and Juan Manuel García-Ruíz
Crystals 2020, 10(2), 68; https://doi.org/10.3390/cryst10020068 - 25 Jan 2020
Cited by 10 | Viewed by 3182
Abstract
It has been previously shown that the diffraction quality of protein crystals strongly depends on mass transport during their growth. In fact, several studies support the idea that the higher the contribution of the diffusion during mass transport, the better the diffraction quality [...] Read more.
It has been previously shown that the diffraction quality of protein crystals strongly depends on mass transport during their growth. In fact, several studies support the idea that the higher the contribution of the diffusion during mass transport, the better the diffraction quality of the crystals. In this work, we have compared the crystal quality of two model (thaumatin and insulin) and two target (HBII and HBII-III) proteins grown by two different methods to reduce/eliminate convective mass transport: crystal growth in agarose gels and crystal growth in solution under microgravity. In both cases, we used identical counterdiffusion crystallization setups and the same data collection protocols. Additionally, critical parameters such as reactor geometry, stock batches of proteins and other chemicals, temperature, and duration of the experiments were carefully monitored. The diffraction datasets have been analyzed using a principal component analysis (PCA) to determine possible trends in quality indicators. The relevant indicators show that, for the purpose of structural crystallography, there are no obvious differences between crystals grown under reduced convective flow in space and convection-free conditions in agarose gel, indicating that the key factor contributing to crystal quality is the reduced convection environment and not how this reduced convection is achieved. This means that the possible detrimental effect on crystal quality due to the incorporation of gel fibers into the protein crystals is insignificant compared to the positive impact of an optimal convection-free environment provided by gels. Moreover, our results confirm that the counterdiffusion technique optimizes protein crystal quality and validates both environments in order to deliver high quality protein crystals, although other considerations, such as protein/gel interactions, must be considered when defining the optimal crystallization setup. Full article
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13 pages, 4634 KiB  
Article
Monitoring the Production of High Diffraction-Quality Crystals of Two Enzymes in Real Time Using In Situ Dynamic Light Scattering
by Raphaël de Wijn, Kévin Rollet, Sylvain Engilberge, Alastair G. McEwen, Oliver Hennig, Heike Betat, Mario Mörl, François Riobé, Olivier Maury, Eric Girard, Philippe Bénas, Bernard Lorber and Claude Sauter
Crystals 2020, 10(2), 65; https://doi.org/10.3390/cryst10020065 - 23 Jan 2020
Cited by 7 | Viewed by 4763
Abstract
The reproducible preparation of well-diffracting crystals is a prerequisite for every structural study based on crystallography. An instrument called XtalController has recently been designed that allows the monitoring of crystallization assays using dynamic light scattering and microscopy, and integrates piezo pumps to alter [...] Read more.
The reproducible preparation of well-diffracting crystals is a prerequisite for every structural study based on crystallography. An instrument called XtalController has recently been designed that allows the monitoring of crystallization assays using dynamic light scattering and microscopy, and integrates piezo pumps to alter the composition of the mother liquor during the experiment. We have applied this technology to study the crystallization of two enzymes, the CCA-adding enzyme of the psychrophilic bacterium Planococcus halocryophilus, and the lysozyme from hen egg white in the presence of a synthetic chemical nucleant. We were able to (i) detect early nucleation events and (ii) drive the crystallization system (through cycles of dissolution/crystallization) toward growth conditions yielding crystals with excellent diffraction properties. This technology opens a way to the rational production of samples for crystallography, ranging from nanocrystals for electron diffraction, microcrystals for serial or conventional X-ray diffraction, to larger crystals for neutron diffraction. Full article
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8 pages, 2676 KiB  
Article
X-ray Single-Crystal Structural Analysis of a Magnetically Oriented Monoclinic Microcrystal Suspension of α-Glycine
by Tatsuya Tanaka, Chiaki Tsuboi, Kazuaki Aburaya, Fumiko Kimura, Masataka Maeyama and Tsunehisa Kimura
Crystals 2019, 9(11), 561; https://doi.org/10.3390/cryst9110561 - 26 Oct 2019
Cited by 1 | Viewed by 3856
Abstract
We previously reported on a method for X-ray single-crystal structure determination from a powder sample via a magnetically oriented microcrystal suspension (MOMS). The method was successfully applied to orthorhombic microcrystals (L-alanine, P212121). In this study, we apply [...] Read more.
We previously reported on a method for X-ray single-crystal structure determination from a powder sample via a magnetically oriented microcrystal suspension (MOMS). The method was successfully applied to orthorhombic microcrystals (L-alanine, P212121). In this study, we apply this method to monoclinic microcrystals. Unlike most of the orthorhombic MOMSs, monoclinic MOMSs exhibit two or four orientations with the same magnetic energy (we refer to this as twin orientations), making data processing difficult. In this paper, we perform a MOMS experiment for a powder sample of monoclinic microcrystal (α-glycine, P21/n) to show that our method can also be applied to monoclinic crystals. The single-crystal structure determined in this work is in good agreement with the reported one performed on a real single crystal. Furthermore, the relationship between the crystallographic and magnetic susceptibility axes is determined. Full article
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8 pages, 1147 KiB  
Article
Analysis of Glulisine Crystallisation Utilising Phase Diagrams and Nucleants
by Yanmin Li, Lata Govada, Hodaya V. Solomon, Richard B. Gillis, Gary G. Adams and Naomi E. Chayen
Crystals 2019, 9(9), 462; https://doi.org/10.3390/cryst9090462 - 3 Sep 2019
Cited by 3 | Viewed by 3796
Abstract
Glulisine is a US Food and Drug Administration (FDA) approved insulin analogue, used for controlling hyperglycaemia in patients with diabetes mellitus (DM). It is fast acting which better approximates physiological insulin secretion, improving patient outcome. Crystallisation of Glulisine was analysed by its crystallisation [...] Read more.
Glulisine is a US Food and Drug Administration (FDA) approved insulin analogue, used for controlling hyperglycaemia in patients with diabetes mellitus (DM). It is fast acting which better approximates physiological insulin secretion, improving patient outcome. Crystallisation of Glulisine was analysed by its crystallisation phase diagram and nucleation-inducing materials. Both the hanging drop vapour diffusion and microbatch-under-oil methods were used and compared. We have shown that the same protein can have different solubility behaviours depending on the nature of the salt in the precipitating agent. In the case of Glulisine with magnesium formate, lowering the precipitant concentration drove the system further into supersaturation resulting in the formation of crystals and precipitation. This was the opposite effect to the usual scenario where raising the precipitant concentration leads to supersaturation. Glulisine with sodium potassium tartrate tetrahydrate (NaKT) followed the expected trend of forming crystals or precipitate at higher concentrations and clear drops at lower concentrations of the precipitant. The outcomes of crystallisation using the different crystallisation methods is also described. Glulisine was successfully crystallised and the crystals diffracted up to a resolution limit of 1.4 Å. Full article
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19 pages, 10309 KiB  
Article
The Role of Calcium and Strontium as the Most Dominant Elements during Combinations of Different Alkaline Earth Metals in the Synthesis of Crystalline Silica-Carbonate Biomorphs
by Mayra Cuéllar-Cruz and Abel Moreno
Crystals 2019, 9(8), 381; https://doi.org/10.3390/cryst9080381 - 24 Jul 2019
Cited by 13 | Viewed by 4367
Abstract
The origin of life from the chemical point of view is an intriguing and fascinating topic, and is of continuous interest. Currently, the chemical elements that are part of the different cellular types from microorganisms to higher organisms have been described. However, although [...] Read more.
The origin of life from the chemical point of view is an intriguing and fascinating topic, and is of continuous interest. Currently, the chemical elements that are part of the different cellular types from microorganisms to higher organisms have been described. However, although science has advanced in this context, it has not been elucidated yet which were the first chemical elements that gave origin to the first primitive cells, nor how evolution eliminated or incorporated other chemical elements to give origin to other types of cells through evolution. Calcium, barium, and strontium silica-carbonates have been obtained in vitro and named biomorphs, because they mimic living organism structures. Therefore, it is considered that these forms can resemble the first structures that were part of primitive organisms. Hence, the objective of this work was to synthesize biomorphs starting with different mixtures of alkaline earth metals—beryllium (Be2+), magnesium (Mg2+), calcium (Ca2+), barium (Ba2+), and strontium (Sr2+)—in the presence of nucleic acids, RNA and genomic DNA (gDNA). Our results allow us to infer that the stability of calcium followed by strontium had played an important role in the evolution of life since the Precambrian era until our current age. In this way, the presence of these two chemical elements as well as silica (in the primitive life) and some organic molecules give origin to a great variety of life forms, in which calcium is the most common dominating element in many living organisms as we know nowadays. Full article
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11 pages, 2954 KiB  
Article
Influence of Pyruvic Acid and UV Radiation on the Morphology of Silica-carbonate Crystalline Biomorphs
by Karina S. Pérez and Abel Moreno
Crystals 2019, 9(2), 67; https://doi.org/10.3390/cryst9020067 - 28 Jan 2019
Cited by 5 | Viewed by 3797
Abstract
In this work we report the effect of introducing pyruvic acid (PA) in the growing process of silica-carbonate biomorphs. Gas-diffusion and single-phase methods were performed, and different concentrations of pyruvic acid were tested. Moreover, influence of UV radiation on the morphogenesis of the [...] Read more.
In this work we report the effect of introducing pyruvic acid (PA) in the growing process of silica-carbonate biomorphs. Gas-diffusion and single-phase methods were performed, and different concentrations of pyruvic acid were tested. Moreover, influence of UV radiation on the morphogenesis of the samples was analyzed. Since PA decomposes in CO2 and other compounds under UV radiation, here we demonstrate that PA decomposition enables a source of carbonate ions to induce the precipitation of silica-carbonate biomorphs in absence of environmental CO2. We also found that high concentrations [0.5 M] of PA inhibit the formation of biomorphs, while lower concentrations [0.01 M] results in common life-like structures. However [0.1 M] of PA provokes the precipitation of carbonates of alkaline earth metals in non-usual crystalline habits, i.e., semi-spherical smoothed shapes sized between 10 and 70 µm and homogeneously growth on a glass substrate. Full article
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9 pages, 6026 KiB  
Article
Synthesis of Bimetallic Nanoparticles of Cd4HgS5 by Candida Species
by Araceli Romero-Núñez, Gonzalo González, Josué E. Romero-Ibarra, Arturo Vega-González, Gustavo Cruz-Jiménez, Orlando Hernández-Cristóbal, Ramón A. Zárraga-Núñez, Armando Obregón-Herrera, Everardo López-Romero, Mario Pedraza-Reyes and Mayra Cuéllar-Cruz
Crystals 2019, 9(2), 61; https://doi.org/10.3390/cryst9020061 - 24 Jan 2019
Cited by 3 | Viewed by 2896
Abstract
In recent decades, it has been demonstrated that bimetallic nanoparticles (NPs) possess a number of advantages over monometallic NPs, as the combination of metals results in important changes to their physicochemical properties. Synthesis of bimetallic NPs can be achieved through a number of [...] Read more.
In recent decades, it has been demonstrated that bimetallic nanoparticles (NPs) possess a number of advantages over monometallic NPs, as the combination of metals results in important changes to their physicochemical properties. Synthesis of bimetallic NPs can be achieved through a number of methods, yet there are serious difficulties in controlling these protocols. Biological methods based on the use of microorganisms exhibit important advantages over traditional methods, which makes the search for organisms such as bacteria, yeast and fungi endowed with these abilities an important task. In this context, it has been found that Candida species are able to biosynthesize monometallic NPs, but their ability to form bimetallic NPs has not been investigated. CdHgS is a bimetallic NP of special interest, as it has been found useful in a number of applications; however, its preparation by traditional methods poses certain limitations, and the ability to obtain it through biological procedures has never been demonstrated. With this in mind, the major purpose of this study is to evaluate whether several Candida species were able to synthesize bimetallic NPs of CdHgS in a Cd4HgS5 phase. To our knowledge, this is the first report on the biological synthesis of bimetallic NPs in Candida species. Full article
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11 pages, 31692 KiB  
Article
Synthesis and Characterization of a Monoclinic Crystalline Phase of Hydroxyapatite by Synchrotron X-ray Powder Diffraction and Piezoresponse Force Microscopy
by Ricardo Pérez-Solis, José Juan Gervacio-Arciniega, Boby Joseph, María Eugenia Mendoza and Abel Moreno
Crystals 2018, 8(12), 458; https://doi.org/10.3390/cryst8120458 - 8 Dec 2018
Cited by 19 | Viewed by 6410
Abstract
In this work, we report the synthesis of a monoclinic hydroxyapatite [Ca10(PO4)6(OH)2] (hereafter called HA) prepared by the sol-gel method assisted by ultrasound radiation at room temperature. The characterization of both the monoclinic and the [...] Read more.
In this work, we report the synthesis of a monoclinic hydroxyapatite [Ca10(PO4)6(OH)2] (hereafter called HA) prepared by the sol-gel method assisted by ultrasound radiation at room temperature. The characterization of both the monoclinic and the hexagonal phases were performed by powder X-ray diffraction (PXRD) and using synchrotron radiation (SR). The measurement of the piezoelectricity was performed by piezoresponse force microscopy (PFM). The synthesis produced a mixture of monoclinic and hexagonal hydroxyapatite (HA). We also discuss the importance of stabilizing the monoclinic phase at room temperature with ultrasound irradiation. The existence of the monoclinic phase has important advantages in terms of showing piezoelectric properties for applications in the new medical rehabilitation therapies. Rietveld refinement of the PXRD data from SR indicated the monoclinic phase to be of about 81%. Finally, piezoelectric force microscopy was used to distinguish the phases of hydroxyapatite by measuring the average piezoelectric coefficient deff = 10.8 pm/V. Full article
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Review

Jump to: Research

35 pages, 12684 KiB  
Review
Applications of X-ray Powder Diffraction in Protein Crystallography and Drug Screening
by Maria Spiliopoulou, Alexandros Valmas, Dimitris-Panagiotis Triandafillidis, Christos Kosinas, Andrew Fitch, Fotini Karavassili and Irene Margiolaki
Crystals 2020, 10(2), 54; https://doi.org/10.3390/cryst10020054 - 21 Jan 2020
Cited by 19 | Viewed by 10311
Abstract
Providing fundamental information on intra/intermolecular interactions and physicochemical properties, the three-dimensional structural characterization of biological macromolecules is of extreme importance towards understanding their mechanism of action. Among other methods, X-ray powder diffraction (XRPD) has proved its applicability and efficiency in numerous studies of [...] Read more.
Providing fundamental information on intra/intermolecular interactions and physicochemical properties, the three-dimensional structural characterization of biological macromolecules is of extreme importance towards understanding their mechanism of action. Among other methods, X-ray powder diffraction (XRPD) has proved its applicability and efficiency in numerous studies of different materials. Owing to recent methodological advances, this method is now considered a respectable tool for identifying macromolecular phase transitions, quantitative analysis, and determining structural modifications of samples ranging from small organics to full-length proteins. An overview of the XRPD applications and recent improvements related to the study of challenging macromolecules and peptides toward structure-based drug design is discussed. This review congregates recent studies in the field of drug formulation and delivery processes, as well as in polymorph identification and the effect of ligands and environmental conditions upon crystal characteristics. These studies further manifest the efficiency of protein XRPD for quick and accurate preliminary structural characterization. Full article
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