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The nucleation of protein crystals as a race against time with on- and off-pathways

Published on Aug 1, 2017in Journal of Applied Crystallography2.867
· DOI :10.1107/S1600576717007312
Cecília Ferreira5
Estimated H-index: 5
,
Silvia Barbosa22
Estimated H-index: 22
+ 3 AuthorsPedro M. Martins11
Estimated H-index: 11
Abstract
  • References (46)
  • Citations (5)
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References46
Newest
The study of drug candidates for the treatment of amyloidosis and neurodegenerative diseases frequently involves in vitro measurements of amyloid fibril formation. Macromolecular crowding and off-pathway aggregation (OPA) are, by different reasons, two important phenomena affecting the scalability of amyloid inhibitors and their successful application in vivo. On the one hand, the cellular milieu is crowded with macromolecules that drastically increase the effective (thermodynamic) concentration...
4 CitationsSource
#1Cecília FerreiraH-Index: 5
#2Fernando RochaH-Index: 22
Last. Pedro M. MartinsH-Index: 11
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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...
2 CitationsSource
Abstract Protein crystal nucleation is a central problem in biological crystallography and other areas of science, technology, and medicine. Recent studies have demonstrated that protein crystal nuclei form within crucial precursors. Data for several proteins provided by these methods have demonstrated that the nucleation precursors are clusters consisting of protein dense liquid, which are metastable with respect to the host protein solution. The clusters are several hundred nanometers in size,...
14 CitationsSource
#2A. E. BlagovH-Index: 8
Last. Vladimir V. VolkovH-Index: 1
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From the base structure of tetragonal lysozyme crystals, the octamer cluster was selected as the possible element of crystal growth. The proposed model was verified by small-angle X-ray scattering (SAXS) measurements of lysozyme solutions. The results showed a noticeable presence of lysozyme dimers and octamers under crystallization conditions and the total absence of oligomers under conditions where crystal growth was impossible.
17 CitationsSource
#1Juan Manuel García-Ruiz (CSIC: Spanish National Research Council)H-Index: 31
#2Fermín Otálora (CSIC: Spanish National Research Council)H-Index: 23
Last. Alfonso García-Caballero (CSIC: Spanish National Research Council)H-Index: 3
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Mass transport takes place within the mesoscopic to macroscopic scale range and plays a key role in crystal growth that may affect the result of the crystallization experiment. The influence of mass transport is different depending on the crystallization technique employed, essentially because each technique reaches supersaturation in its own unique way. In the case of batch experiments, there are some complex phenomena that take place at the interface between solutions upon mixing. These transp...
7 CitationsSource
#1J. Ng (SGC: Structural Genomics Consortium)H-Index: 3
#2C. Dekker (Novartis)H-Index: 2
Last. F. von Delft (UJ: University of Johannesburg)H-Index: 14
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Although protein crystallization is generally considered more art than science and remains significantly trial-and-error, large-scale data sets hold the promise of providing general learning. Observations are presented here from retrospective analyses of the strategies actively deployed for the extensive crystallization experiments at the Oxford site of the Structural Genomics Consortium (SGC), where comprehensive annotations by SGC scientists were recorded on a customized database infrastructur...
17 CitationsSource
#1Rosa Crespo (University of Porto)H-Index: 6
#2Eva Villar-Alvarez (University of Santiago de Compostela)H-Index: 7
Last. Pedro M. Martins (University of Porto)H-Index: 11
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Some of the most prevalent neurodegenerative diseases are characterized by the accumulation of amyloid fibrils in organs and tissues. Although the pathogenic role of these fibrils has not been completely established, increasing evidence suggests off-pathway aggregation as a source of toxic/detoxicating deposits that still remains to be targeted. The present work is a step toward the development of off-pathway modulators using the same amyloid-specific dyes as those conventionally employed to scr...
15 CitationsSource
#1Robin Schubert (UHH: University of Hamburg)H-Index: 7
#2Arne Meyer (UHH: University of Hamburg)H-Index: 10
Last. Christian Betzel (UHH: University of Hamburg)H-Index: 39
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Crystallization of biological macromolecules such as proteins implies several prerequisites, for example, the presence of one or more initial nuclei, sufficient amounts of the crystallizing substance and the chemical potential to provide the free energy needed to force the process. The initiation of a crystallization process itself is a stochastic event, forming symmetrically assembled nuclei over kinetically preferred protein-dense liquid clusters. The presence of a spatial repetitive orientati...
11 CitationsSource
#1Marko Ristic (CSIRO: Commonwealth Scientific and Industrial Research Organisation)H-Index: 2
#2Nicholas Rosa (CSIRO: Commonwealth Scientific and Industrial Research Organisation)H-Index: 2
Last. Janet Newman (CSIRO: Commonwealth Scientific and Industrial Research Organisation)H-Index: 27
view all 4 authors...
There is strong evidence to suggest that a protein sample needs to be well folded and uniform in order to form protein crystals, and it is accepted knowledge that the formulation can have profound effects on the behaviour of the protein sample. The technique of differential scanning fluorimetry (DSF) is a very accessible method to determine protein stability as a function of the formulation chemistry and the temperature. A diverse set of 252 soluble protein samples was subjected to a standard fo...
9 CitationsSource
#1Maria A. Vorontsova (UH: University of Houston)H-Index: 8
#2Dominique Maes (Vrije Universiteit Brussel)H-Index: 26
Last. Peter G. Vekilov (UH: University of Houston)H-Index: 47
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The two-step mechanism of nucleation of crystals in solutions posits that the formation of crystal nuclei occurs within structures of extended lifetimes, in which the nucleating solute is at high concentration. The validity of this mechanism has been demonstrated for proteins, small-molecule organic and inorganic materials, colloids, and polymers. Due to large molecule sizes, proteins are an ideal system to study the details of this nucleation pathway, in particular the formation mechanisms of t...
37 CitationsSource
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#2A. S. Boikova (RAS: Russian Academy of Sciences)H-Index: 2
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#1Jorge A. Ferreira (Katholieke Universiteit Leuven)H-Index: 6
#2Filipa Castro (University of Minho)H-Index: 6
Last. Simon Kuhn (Katholieke Universiteit Leuven)H-Index: 18
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S.K. acknowledges funding from the European Research Council under the ERC Starting Grant Agreement no. 677169–MicroParticleControl. F.C acknowledges FCT (postdoctoral fellowship [SFRH/BPD/96132/2013]) under the project POCI-01-0145-FEDER-006939 (Laboratory for Process Engineering, Environment, Biotechnology and Energy – UID/EQU/00511/2013) funded by the European Regional Development Fund (ERDF), through COMPETE2020 – Programa Operacional Competitividade e Internacionalizacao (POCI) and by natio...
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#1Yong YuH-Index: 1
#2Kai LiH-Index: 6
Last. Ji-Cheng LiH-Index: 1
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Space is expected to be a convection-free, quiescent environment for the production of large-size and high-quality protein crystals. However, the mechanisms by which the diffusion environment in space improves the quality of the protein crystals are not fully understood. The interior of a microfluidic device can be used to simulate a microgravity environment to investigate the protein crystallization mechanism that occurs in space. In the present study, lysozyme crystals were grown in a prototyp...
1 CitationsSource
#1Alexandra SilvaH-Index: 7
#2Zsuzsa SárkányH-Index: 4
Last. Pedro M. MartinsH-Index: 11
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Drug discovery frequently relies on the kinetic analysis of physicochemical reactions that are at the origin of the disease state. Amyloid fibril formation has been extensively investigated in relation to prevalent and rare neurodegenerative diseases, but thus far no therapeutic solution has directly arisen from this knowledge. Other aggregation pathways producing smaller, hard-to-detect soluble oligomers are increasingly appointed as the main reason for cell toxicity and cell-to-cell transmissi...
1 CitationsSource
#1Christo N. NanevH-Index: 1
Three-dimensional protein molecule structures are essential for acquiring a deeper insight of the human genome, and for developing novel protein-based pharmaceuticals. X-ray diffraction studies of such structures require well-diffracting protein crystals. A set of external physical factors may promote and direct protein crystallization so that crystals obtained are useful for X-ray studies. Application of electric fields aids control over protein crystal size and diffraction quality. Protein cry...
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