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Nucleation and Crystallization of Globular Proteins: What we Know and What is Missing

Published on Oct 1, 1996in Journal of Crystal Growth1.57
· DOI :10.1016/0022-0248(96)00358-2
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville),
Peter G. Vekilov47
Estimated H-index: 47
(UAH: University of Alabama in Huntsville)
+ 1 AuthorsB. R. Thomas17
Estimated H-index: 17
(UAH: University of Alabama in Huntsville)
Cite
Abstract
Recently, much progress has been made in understanding the nucleation and crystallization of globular proteins, including the formation of compositional and structural crystal defects. Insight into the interactions of (screened) protein macro-ions in solution, obtained from light scattering, small angle X-ray scattering and osmotic pressure studies, can guide the search for crystallization conditions. These studies show that the nucleation of globular proteins is governed by the same principles as that of small molecules. However, failure to account for direct and indirect (hydrodynamic) protein interactions in the solutions results in unrealistic aggregation scenarios. Microscopic studies of numerous proteins reveal that crystals grow by the attachment of growth units through the same layer-spreading mechanisms as inorganic crystals. Investigations of the growth kinetics of hen-egg-white lysozyme (HEWL) reveal non-steady behavior under steady external conditions. Long-term variations in growth rates are due to changes in step-originating dislocation groups. Fluctuations on a shorter timescale reflect the non-linear dynamics of layer growth that results from the interplay between interfacial kinetics and bulk transport. Systematic gel electrophoretic analyses suggest that most HEWL crystallization studies have been performed with material containing other proteins at percent levels. Yet, sub-percent levels of protein impurities impede growth step propagation and play a role in the formation of structural/compositional inhomogeneities. In crystal growth from highly purified HEWL solutions, however, such inhomogeneities are much weaker and form only in response to unusually large changes in growth conditions. Equally important for connecting growth conditions to crystal perfection and diffraction resolution are recent advances in structural characterization through high-resolution Bragg reflection profiling and X-ray topography.
  • References (104)
  • Citations (128)
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References104
Newest
Published on Oct 1, 1996in Journal of Crystal Growth1.57
Martin Muschol19
Estimated H-index: 19
(UAH: University of Alabama in Huntsville),
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville)
There have been numerous claims of large concentrations of prenucleation aggregates in supersaturated as well as undersaturated lysozyme solutions at high salt concentrations. The presence of these aggregates was derived from measurements of the light or neutron scattering intensity, ultracentrifugation and dialysis behavior, as well as over-simplified crystal growth kinetics considerations. In all these interpretations it has been assumed that lysozyme solutions are either ideal or that protein...
Published on Oct 1, 1996in Journal of Crystal Growth1.57
Yoshiki Higuchi32
Estimated H-index: 32
,
Takuya Okamoto1
Estimated H-index: 1
,
Noritake Yasuoka31
Estimated H-index: 31
The heterogeneity in a single crystal of hydrogenase has been recognized using synchrotron radiation. This heterogeneity has been estimated from five different positions of the crystal by analyzing the number of significant reflections above background levels and the mosaicity of the diffraction spots. The crystal showed the highest degree of mosaicity at one end of the crystal, the initial point of crystal growth. The degree of mosaicity decreases along the c-axis from the initial point of crys...
Peter G. Vekilov47
Estimated H-index: 47
,
Lisa A. Monaco5
Estimated H-index: 5
+ 2 AuthorsFranz Rosenberger30
Estimated H-index: 30
Nonuniform precipitant and impurity incorporation in protein crystals can cause lattice strain and, thus, possibly decrease the X-ray diffraction resolution. To address this issue, a series of crystallization experiments were carried out, in which initial supersaturation, NaCl concentration, protein purity level and crystallized fraction were varied. Lysozyme and protein impurities, as well as sodium and chloride were independently determined in the initial solution, supernatant and crystals. Th...
B. R. Thomas17
Estimated H-index: 17
,
Peter G. Vekilov47
Estimated H-index: 47
,
Franz Rosenberger30
Estimated H-index: 30
Hen egg-white lysozyme (HEWL) is widely used as a model protein, although its purity has not been adequately characterized by modern biochemical techniques. We have identified and quantified the protein heterogeneities in three commercial HEWL preparations by sodium dodecyl sulfate polyacrylamide gel electrophoresis with enhanced silver staining, reversedphase fast protein liquid chromatography (FPLC) and immunoblotting with comparison to authentic protein standards. Depending on the source, the...
Published on Feb 1, 1996in Journal of Crystal Growth1.57
Hong Lin7
Estimated H-index: 7
(UAH: University of Alabama in Huntsville),
Peter G. Vekilov47
Estimated H-index: 47
(UAH: University of Alabama in Huntsville),
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville)
A model for the evolution of facet morphologies in growth from solutions is presented. The numerical model links, for the first time, bulk transport of solute and impurities in a solution growth cell with microscopic interfacial kinetics processes. The macroscopic transport is dealt with as in the 2D model [H. Lin, F. Rosenberger, J.I.D. Alexander and A. Nadarajah, J. Crystal Growth 151 (1995) 153] of a crystallization cell used for lysozyme in our laboratory. The microscopic kinetics is incorpo...
Published on Feb 1, 1996in Journal of Crystal Growth1.57
Peter G. Vekilov47
Estimated H-index: 47
(UAH: University of Alabama in Huntsville),
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville)
Interferometric microscopy was used to investigate the growth morphology and kinetics of {110} and {101} faces of tetragonal lysozyme crystals. Solutions were prepared from as-received Sigma and Seikagaku material, and Seikagaku lysozyme further purified by cation exchange liquid chromatography under salt-free conditions. The protein composition of the solutions was characterized by sodium dodecyl sulphate (SDS) electrophoresis with silver staining. We found that on crystals smaller than about 1...
Published on Dec 22, 1995in Journal of Chemical Physics3.00
Martin Muschol19
Estimated H-index: 19
(UAH: University of Alabama in Huntsville),
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville)
We have performed multiangle static and dynamic light scattering studies of lysozyme solutions at pH=4.7. The Rayleigh ratio Rθ and the collective diffusion coefficient Dc were determined as function of both protein concentration cp and salt concentration cs with two different salts. At low salt concentrations, the scattering ratio Kcp/Rθ and diffusivity increased with protein concentration above the values for a monomeric, ideal solution. With increasing salt concentration this trend was eventu...
Published on Nov 1, 1995in Journal of Crystal Growth1.57
Peter G. Vekilov47
Estimated H-index: 47
(UAH: University of Alabama in Huntsville),
Lisa A. Monaco5
Estimated H-index: 5
(UAH: University of Alabama in Huntsville),
Franz Rosenberger30
Estimated H-index: 30
(UAH: University of Alabama in Huntsville)
Abstract The growth morphology and kinetics of crystal facets were investigated by in-situ high-resolution optical interferometry, using tetragonal lysozyme as a model system. The protein composition of the growth solutions was characterized by high pressure liquid chromatography and gel electrophoresis. In relatively pure solutions, the facets exhibited depressions with lower vicinal slope at their edges. In solutions with ≤ 1% of protein impurities, that were found to be incorporated into the ...
Published on Nov 1, 1995in Journal of Crystal Growth1.57
Meirong Li2
Estimated H-index: 2
(UAH: University of Alabama in Huntsville),
Arunan Nadarajah7
Estimated H-index: 7
(UAH: University of Alabama in Huntsville),
Marc L. Pusey26
Estimated H-index: 26
(MSFC: Marshall Space Flight Center)
Abstract Although the faceted growth of tetragonal lysozyme crystals is known to occur by 2D nucleation and dislocation-led growth, the measured growth rates do not follow model predictions based on these mechanisms. One possible reason for this deviation is that these models ignore the highly aggregated state of lysozyme in supersaturated solutions. In this study a growth mechanism for tetragonal lysozyme crystals involving aggregation reactions leading to the formation of the growth unit, mass...
Published on Oct 2, 1995in Physical Review Letters9.23
T.A. Land15
Estimated H-index: 15
,
Alexander J. Malkin22
Estimated H-index: 22
(LLNL: Lawrence Livermore National Laboratory)
+ 2 AuthorsJ. J. De Yoreo22
Estimated H-index: 22
(LLNL: Lawrence Livermore National Laboratory)
The evolution of surface morphology and step dynamics during growth of single crystals of the protein Canavalin and of the cubic satellite tobacco mosaic virus crystals (STMV) have been investigated by in situ atomic force microscopy. These two crystals were observed to grow by very different mechanisms. Growth of Canavalin occurs on complex vicinal hillocks formed by multiple, independently acting screw dislocations. Small cluster were observed on the terraces. STMV on the other hand, was obser...
Cited By128
Newest
Published on Jun 20, 2019
Peter G. Vekilov47
Estimated H-index: 47
Published on Apr 22, 2019in Philosophical Transactions of the Royal Society A
D.V. Alexandrov18
Estimated H-index: 18
(UrFU: Ural Federal University),
I. G. Nizovtseva3
Estimated H-index: 3
(FSU: University of Jena)
A generalized theory of nucleation and growth of crystals in a metastable (supercooled or supersaturated) liquid is developed taking into account two principal effects: the diffusion mechanism of the particle-size distribution function in the space of particle radii and the unsteady-state growth rates of individual crystals induced by fluctuations in external temperature or concentration field. A system of the FokkerPlanck and balance integro-differential equations is formulated and analytically...
Published on Jan 1, 2018
James M. Van Alstine3
Estimated H-index: 3
(KTH: Royal Institute of Technology),
Günter Jagschies1
Estimated H-index: 1
(GE Healthcare),
Karol M. Łącki
Abstract As with many other alternative separations methods, crystallization and aqueous polymer two-phase extraction (ATPE) have been well-studied academically and are amenable to predictive and process modeling. Their solution nature allows them to be optimized using high-throughput microtiter plate-based methodologies. Their liquid nature allows them to be readily scaled and, in comparison with some other separation methods, they do not require overt energy use or capital investment. Crystall...
Published on Jan 1, 2016in CrystEngComm3.38
Ivaylo L. Dimitrov4
Estimated H-index: 4
(BAS: Bulgarian Academy of Sciences),
Dobryana P. Koleva1
Estimated H-index: 1
(BAS: Bulgarian Academy of Sciences),
Feyzim V. Hodzhaoglu5
Estimated H-index: 5
(BAS: Bulgarian Academy of Sciences)
The aggregation of lysozyme in crystallizing solutions prior to crystal nucleation and during crystal growth has been the subject of numerous investigations over the past two decades. Nevertheless, it remains a controversial rather than a well-recognized phenomenon. In this study, we investigate the growth of tetragonal lysozyme crystals in quiescent solutions at the early crystal growth stage using seven different protein concentrations ranging from 25 mg ml−1 to 55 mg ml−1, 5% NaCl (w/v), 0.1 ...
Published on Nov 1, 2015
Alexander McPherson53
Estimated H-index: 53
,
Lawrence J. DeLucas24
Estimated H-index: 24
Published on Aug 31, 2015in Russian Chemical Reviews4.61
Konstantin M. Boyko8
Estimated H-index: 8
(RAS: Russian Academy of Sciences),
Vladimir O. Popov19
Estimated H-index: 19
(RAS: Russian Academy of Sciences),
M. V. Kovalchuk11
Estimated H-index: 11
(RAS: Russian Academy of Sciences)
Published on Jul 1, 2015in Bioprocess and Biosystems Engineering2.37
Dariusch Hekmat14
Estimated H-index: 14
(TUM: Technische Universität München)
Since about 170 years, salts were used to create supersaturated solutions and crystallize proteins. The dehydrating effect of salts as well as their kosmotropic or chaotropic character was revealed. Even the suitability of organic solvents for crystallization was already recognized. Interestingly, what was performed during the early times is still practiced today. A lot of effort was put into understanding the underlying physico-chemical interaction mechanisms leading to protein crystallization....
Published on May 1, 2015in Scientific Reports4.01
Marlus Alves dos Santos4
Estimated H-index: 4
,
Francesco Brugnera Teixeira1
Estimated H-index: 1
+ 12 AuthorsRafael Gonçalves Barbosa Gomes2
Estimated H-index: 2
Structural studies of proteins normally require large quantities of pure material that can only be obtained through heterologous expression systems and recombinant technique. In these procedures, large amounts of expressed protein are often found in the insoluble fraction, making protein purification from the soluble fraction inefficient, laborious, and costly. Usually, protein refolding is avoided due to a lack of experimental assays that can validate correct folding and that can compare the co...
Published on Jan 1, 2015in Faraday Discussions3.71
Zoubida Hammadi10
Estimated H-index: 10
(AMU: Aix-Marseille University),
Romain Grossier10
Estimated H-index: 10
(AMU: Aix-Marseille University)
+ 4 AuthorsStéphane Veesler28
Estimated H-index: 28
(AMU: Aix-Marseille University)
Do the differing properties of materials influence their nucleation mechanisms? We present different experimental approaches to study and control nucleation, and shed light on some of the factors affecting the nucleation process.
Published on Sep 1, 2014in European Physical Journal B1.44
Giuseppe Pellicane16
Estimated H-index: 16
(UKZN: University of KwaZulu-Natal),
Lev Sarkisov21
Estimated H-index: 21
(Edin.: University of Edinburgh)
In this article we explore a problem of effective interactions between two rotationally restrained lysozyme molecules forming a crystal contact in aqueous solution. We perform non-equilibrium molecular dynamics simulations in order to estimate the interaction energy as a function of the distance between the two proteins obtained from direct application of the Jarzynski equality (JE), and compare it with that calculated by means of another non-equilibrium approach (Forward-Reverse method) and con...
View next paperInteractions in undersaturated and supersaturated lysozyme solutions: Static and dynamic light scattering results