Protein & Peptide Letters, 2012, 19, 725-731

725

Predicting Protein Crystallizability and Nucleation

Nuria Snchez-Puig1, Claude Sauter2, Bernard Lorber2, Richard Gieg2 and Abel Moreno1,*
1

Instituto de Qumica, Universidad Nacional Autnoma de Mxico, Circuito Exterior, C.U. Mxico, D.F. 04510
(Mxico); 2Architecture et Ractivit de lARN, Universit de Strasbourg, CNRS, IBMC, UPR9002, 15 rue Ren Descartes 67000 Strasbourg (France)
Abstract: The outcome of protein crystallization attempts is often uncertain due to inherent features of the protein or to
the crystallization process that are not fully under control of the experimentalist. The aim of this contribution is to propose
user-friendly tools that can increase the success rate of a protein crytallization project. Different bioinformatic approaches
to predict the crystallization feasibility (before any crystallization attempts are undertaken) are discussed and a novel approach to assess the nucleation process of a given protein is proposed. Practical examples illustrate these two points.

Keywords: Proteins, crystallization feasibility, crystal growth, X-ray diffraction, nucleation, surface energy.
1. INTRODUCTION
X-ray crystallography is a powerful technique for determining the 3D structure of proteins. However, as obvious as
it seems, it requires the production of protein crystals of suitable quality. Despite the existence of a huge variety of crystallization technologies and access to high-throughput
screening systems, statistics from the various structural programs indicate that only ~15% of expressed proteins yield
diffracting crystals. This represents a very low success rate
considering the cumulative difficulties of cloning, expressing
and purifying the proteins [1]. The reasons why some proteins do not crystallize are difficult to identify, but most
likely relate to the intrinsic physico-chemical properties of
the protein [2]. Therefore, it is useful to have user-friendly
tools that allow the experimenter to a priori select possible
successful protein targets for crystallization or to identify
problematic proteins. Proteins recalcitrant to crystallize
could be highly flexible [3] and even completely unstructured or despite being well folded would not nucleate properly for many reasons, such as a propensity to aggregate as
amorphous phase or difficulties to form stable crystal contacts [4]. Thus, getting good crystals can be very tricky and
often needs a combination of protein engineering, the use of
sophisticated crystallization techniques and a good understanding of the nucleation and crystal growth process.
This paper sketches different tools that may help to increase the success rate of a crystallization project. The first
part covers methods useful for crystallization prediction before any crystallization attempts are undertaken. The second
part presents a new way to predict the nucleation output together with some initial practical applications. Finally, considerations on the importance of impurities in protein crystal
growth are presented.

*Address correspondence to this author at the Instituto de Qumica, UNAM.

Circuito Exterior, C.U. Mxico, D.F. 04510 Mexico (Mexico);
Tel: +52-55-56224467; Fax: +52-55-56162217; E-mail: carcamo@unam.mx

1875-5305/12 $58.00+.00

2. PREDICTING PROTEIN CRYSTALLIZABILITY

Protein biochemistry has been dominated by the idea that
a well-defined structure is a pre-requisite for function. However, this structure-function paradigm has been re-evaluated
based on the growing evidence that many proteins contain
disordered domains and that polypeptide chains can exhibit
considerable flexibility or be completely unfolded in their
functional states [5]. For long, the structural characterization
of such proteins has been hindered due to their highly dynamic conformations. To overcome this bottleneck, crystallization data from the Structural Genomic consortia have been
used to create crystallizability predictors that require only the
protein sequence as input [6-14]. Depending on the program,
the score can be as simple as the phrase crystallizable protein or non-crystallizable protein like in the case of the
OB-Score program [10]. Programs such as XtalPred [13]
give the output as a number between 1 and 5, the highest
value standing for proteins "very difficult" to crystallize and
the lowest for proteins expected to be "optimal" to crystallize. These algorithms analyze protein characteristics such as
sequence length, isoelectric point, average hydropathy [15],
instability index [16] and existence of low complexity regions such as signal peptides [17], trans-membrane helices
[18] and coiled-coil regions [19]. Comparative analyses suggest that the programmes XtalPred [13], CRYSTALP2 [8],
ParCrys [11] and OB-score [10] have a similar prediction
accuracy of ~70%. A majority-vote based combination of
these methods only improved the prediction success rate to
74% [20]. The program MetaPPCP is a linear model treebased meta-predictor that uses the complementarity of the
state-of-the-art protein crystallization propensity predictors
and claims to improve the accuracy of predictions up to over
80% [9].
Besides analyzing the crystallization propensity, it is also
important to identify if the protein of interest is an intrinsically unstructured protein or not, since this may also hinder
its chances of crystallizing. There are several programs useful at predicting the propensity of regions longer than 40
2012 Bentham Science Publishers

726 Protein & Peptide Letters, 2012, Vol. 19, No. 7

consecutive residues to be disordered [21-31]. The most

widely used disorder predictors are those of the PONDR
family. The output of these predictors is a positiondependent score, which varies between 0 and 1 with a minimum threshold of 0.5 for a residue to be considered disordered. In addition, analysis of both, the amino acid composition and the sequence of the protein of interest, can also hint
to a protein being intrinsically unstructured. Statistical analysis has shown that intrinsically disordered proteins or segments thereof, consist of low complexity amino acid sequences. They commonly contain high levels of the amino
acids R, K, E, P and S, denoted as disorder promoting residues, and low levels of the amino acids C, W, Y, I and V,
considered to be order promoting residues [32]. The sequence of a protein can be used to calculate parameters such
as the net mean charge <R> and mean hydropathy <H>. A
combination of low mean hydropathy and relatively high net
mean charge are determinants for the absence of compact
structure in proteins [33, 34]. To calculate these values it is
necessary to consider that the mean net charge <R> is defined as the absolute value of the difference between the
numbers of positively and negatively charged residues divided by the total number of residues. The mean <H> is the
sum of normalized hydrophobicities of individual residues
divided by the total number of amino acids present in the
protein. Individual hydrophobicities are calculated as described in [15]. Disordered proteins display characteristic
<R> and <H> values that can be compared to those described in [34]. Analysis such as that mentioned above has
proven accurate at predicting disordered proteins [35, 36]. If
a protein fulfills all the criteria mentioned above it strongly
suggests that it belongs to the class of intrinsically disordered
proteins. Although very difficult, crystallizing an intrinsi-

Snchez-Puig et al.

cally unstructured protein may be achieved in complex with

an endogenous partner. However, expression and purification of such partners are not always possible, nor do all proteins have known binding partners. Furthermore, even when
the binding partners are known it is important to get the correct boundaries of the target protein such that any flexible
regions that do not participate in the interaction are removed.
A good example of such a strategy concerns the crystallization of the C-terminal transactivation domain of HIF-1

bound to the Taz1 domain of CBP and to FIH [37, 38].
In summary, different bioinformatics tools are available
that can help scientists to decide if a target protein is suitable
for crystallization. This knowledge can, in turn, led to redefine goals and to establish what type of project is worth pursuing. This is of practical and strategic importance since
working with a protein whose crystallization prediction is
low, may represent a long and hard process before getting
the structure and may not be worth enduring. A diagram with
a decision-tree summarizing the purpose of this section is
shown in (Figure 1). Finally, strategies to engineer the target
protein can be found in Ruggiero et al., on this special issue.
3. PREDICTING PROTEIN NUCLEATION
3.1. Background
Understanding protein crystal nucleation and crystal
growth is important to successfully crystallize a protein. Nucleation is the first step in any crystallization process and
deals with nucleus formation, nuclei size distribution and
nucleus growth rate until nuclei reach a detectable size.
Typically, nucleation involves different time scales occurring
simultaneously, namely 0.01 ns for molecular conformations,

Figure 1. Schematic view of protein primary structure analysis to predict the crystallization feasibility of a protein and possible decision
routes to take according to the results obtained.

Predicting Protein Crystallizability and Nucleation

ns for surface structure and defect displacements [39], s for

surface step displacement [40], ms for growth of one atomic
layer [41], seconds for hydrodynamic transport, and minutes
for the homogenous nucleation phenomena [42]. There are
several methods to investigate the chemical and physical
interactions between the different molecules present during
crystal growth. For the nucleation step only few techniques
allow optimal resolution in terms of time scaling, quality of
the imaging measurement and visualization of the process. In
general, dynamic (DLS) and static light scattering (SLS)
methods are useful to analyze features such as homogeneity,
nucleation, protein-protein interactions and pre-crystallization conditions [43].
Here we present a method to predict the nucleation behavior of a protein based on the knowledge of hydrodynamic
parameters obtained by light scattering and of crystallogenesis parameters obtained under pre-crystallization conditions.
3.2. How to Relate Nucleation and Hydrodynamic Parameters
Following the classical theory of crystallization in solution and the energetics of crystal nucleation, in particular that
decribed by Kashiev et al. [44] for the calculation of W(n)
(i.e. the work necessary to assemble n protein molecules into
an n-sized cluster) and the revised formula by StefanSkapski-Turnbull [44] for the estimation of the specific surface energy of the cluster/solution interface, one can express
the Gibbs free energy (
G) of nucleation as a function of the
radius of the n-sized protein cluster as follows:

G = [(4/3r3)/] kT ln  + 4r2 
where r is the radius of the protein or protein-cluster,  is the
molar volume occupied by a protein unit in the crystal, k is
the Boltzmann constant, T is the absolute temperature,  is
the supersaturation (the ratio between the protein concentration C in mg mL1 and the solubility Ce which is the protein
concentration remaining in solution in equilibrium with the
crystals), and  is the specific surface energy of the cluster/solution interface. Details on the establisment of the
G =
f(r) formula will be published elsewhere.
The plot of
G versus r shows a maximum of
G representing the critical Gibbs free energy energy (
G*) of the
nucleation phenomenom and defines the critical radius (r*)
of the protein cluster, i.e. the nucleus from which a crystal
will grow (Figure 2). The
G = f(r) plot can be experimentally computed for any protein (or macromolecular assembly) provided experimental values of , , and  are available. They can be derived from light scattering (DLS), crystallographic and solubility data.
Experimental values of  are typically derived from
crystallographic data, but could also be roughly estimated
from the hydrodynamic radius of the protein obtained by
DLS measurements. Experimental estimates of  require
knowledge of the solubility or supersaturation of the protein
at the tested condition. This can be done via the revised
Stefan-Skapski-Turnbull formula [44]:
 = B [kT ln (Ce o)]/o2/3
where B is a numerical factor (ranging from  0.2 to 0.6, its
value being ~0.514 for spherical clusters) and o is the spe-

Protein & Peptide Letters, 2012, Vol. 19, No. 7

727

cific volume of the protein (estimated from o = Mr /N A,

with  the density of the crystal (g cm3), Na is the Avogadro
number and Mr is the molecular mass of the protein (g mol1)
or alternatively via the equation established previously in
[45]
2vo/r = kT ln  + A2 kT Mr Ce (1)
where both A2, the osmotic second virial coefficient (in
mol cm3 g2), and r the radius of the protein and can be derived from DLS measurements. To avoid confusion, it is
considered that the supersaturation  is an alternative expression defined as the ratio between the protein concentration C
(in mg mL1) and Ce (in mg mL1).
3.3. A Few Test Case Predictions of r*

G = f(r) plots have been computed for three test case
proteins, namely hen egg white lysozyme (Mr 14300), thaumatin (Mr 22200) and apoferritin (Mr 443000) based on inhouse experimental data. Table 1 shows the main results,
including the estimated critical nucleus radius (r*) and the
critical Gibbs free energy of nucleation (
G*), for the
aforementioned proteins. Large variations appear when comparing the numerical values of these two crystallogenesis
parameters for the three test-case proteins, with the largest
values of both r* and
G* for thaumatin. Interestingly, the
value of 50 protein molecules forming the nucleus of
lysozyme crystals compares well with data from the literature obtained by other means (2044 protein units) [46, 47]
but deviates significantly from other estimates where size of
the nuclei were either smaller [48] or much larger [49, 50].
Of course, predicted values are subject to errors on the experimental data since the parameters such as , , and , are
determined using biophysical methods. For instance, errors
of 10% of the values (due to experimental conditions) lead to
maximal deviation of 20% on r* in some cases (see article of
Peter Vekilov published in this special issue on nucleation).
Application to other proteins together with a detailed description experimental procedures, critical analysis of the
results and comparison with data from literature obtained by
other experimental approaches will be published elsewhere.
4. CONCLUSIONS AND PERSPECTIVES
Purity is the first variable that is essential to obtain good
crystals. It has been demonstrated for a long time that macromolecular contaminants and micro-heterogeneities that are
present within a protein batch poison the faces of growing
crystals and alter the crystal packing. In order to understand
the effects of these impurities on crystal growth and the correlation with the surface energy it is necessary to estimate
the value of the surface energy  using any of the revised
equations shown in this contribution. These calculations
were applied to three model proteins (lysozyme, thaumatin
and apo-ferritin) to test our approach. From these results we
conclude that the lower the value of the surface energy the
higher the propensity of a protein to be poisoned in its crystal
growth mechanism.
Concerning the crystal growth there is no unique recipe
to crystallize soluble or non-soluble proteins, most of them
need special and particular care based on multifactorial elements (precipitants, temperatures, pH values, transport prop-

728 Protein & Peptide Letters, 2012, Vol. 19, No. 7

Figure 2. Gibbs energy as a function of number of protein units for (A) Lysozyme, (B) Thaumatin and (C) Ferritin.

Snchez-Puig et al.

Predicting Protein Crystallizability and Nucleation

Table 1.

729

Physicochemical Crystallogenesis Parameters for Different Proteins

Parameters

 = C / Ce
0, cm

Protein & Peptide Letters, 2012, Vol. 19, No. 7

Hen egg white Lysozyme

Thaumatin

Apo-Ferritin

2.5

2.7

43

0.2 x 10

-19

0.3 x 10

-19

5.0 x 10-19

, J cm-2

10 x 10-8

20 x 10-8

0.24 x 10-8

, cm3

2.4 x 10-19

5.3 x 10-19

62 x 10-19

50 5

220 22

30 3

r* (in protein units)

-1

G*, J mol

36 x 10

1500 x 10

24 x 103

* means critical; r* values are given assuming an error of 10% on , , and
.

erties, etc.). The propensity of any protein to be crystallized

depends on the nature of the protein, but it can be hinted
using some of the tools presented here. However, these aspects are not usually taken into account and academic labs
seldom predict the crystallization propensity or analyze their
target protein before starting a difficult structural biology
project. The strategy shown in Figure 1 should be followed
as a normal procedure before using kits to search for the
crystallization conditions.

worth mentioning that temperature also plays an important

role (it should be kept constant) while searching for the crystallization conditions as well as the presence of impurities.
All these ideas and procedures should be taken into account
when trying to crystallize a new protein. Choosing the correct strategies and a good understanding of the protein of
interest may provide the experimenters with higher chances
of crystallization success in a shorter time.

Finally, after having understood the physicochemical

crystallization behavior of the target protein, or having analyzed the propensity to crystallize, it is possible to move into
crystallization trials. Figure 3 shows the schematic view of
de novo protein crystallization strategy to be followed. It is

ACKNOWLEDGEMENTS

Figure 3. Schematic strategies for de novo protein crystallization.

The authors thank Universit de Strasbourg and French

CNRS and Agence National pour la Recherche (ANR-09BLAN-0091-03) for support. A.M. acknowledges DGAPAUNAM project PAPIIT No. IN201811 for financial support

730 Protein & Peptide Letters, 2012, Vol. 19, No. 7

for this research and CNRS for sponsorship during his sabbatical visit in Strasbourg. N. S-P acknowledges the financial
support from UNAM-DGAPA project PAPIIT No.
IN204010.
REFERENCES
[1]

[2]
[3]

[4]

[5]
[6]
[7]

[8]

[9]
[10]

[11]
[12]

[13]

[14]

[15]
[16]

[17]

[18]

[19]
[20]
[21]

Fox, B.G.; Goulding, C.; Malkowski, M.G.; Stewart, L.; Deacon,

A. Structural genomics: from genes to structures with valuable materials and many questions in between. Nat. Methods, 2008, 5(2),
129-132.
Dale, G.E.; Oefner, C.; D'Arcy, A. The protein as a variable in
protein crystallization. J. Struct., Biol., 2003, 142(1), 88-97.
Malawski, G.A.; Hillig, R.C.; Monteclaro, F.; Eberspaecher, U.;
Schmitz, A.A.; Crusius, K.; Huber, M.; Egner, U.; Donner, P.;
Muller-Tiemann, B. Identifying protein construct variants with increased crystallization propensity--a case study. Protein Sci., 2006,
15(12), 2718-2728.
Moon, A.F.; Mueller, G.A.; Zhong, X.; Pedersen, L.C. A synergistic approach to protein crystallization: combination of a fixed-arm
carrier with surface entropy reduction. Protein Sci, 2010, 19(5),
901-913.
Dyson, H.J.; Wright, P.E. Intrinsically unstructured proteins and
their functions. Nat. Rev. Mol. Cell. Biol., 2005, 6(3), 197-208.
Chen, K.; Kurgan, L.; Rahbari, M. Prediction of protein crystallization using collocation of amino acid pairs. Biochem. Biophys. Res.
Commun., 2007, 355(3), 764-769.
Kandaswamy, K.K.; Pugalenthi, G.; Suganthan, P.N.; Gangal, R.
SVMCRYS: an SVM approach for the prediction of protein crystallization propensity from protein sequence. Protein Pept. Lett.,
2010, 17(4), 423-430.
Kurgan, L.; Razib, A.A.; Aghakhani, S.; Dick, S.; Mizianty, M.;
Jahandideh, S. CRYSTALP2: sequence-based protein crystallization propensity prediction. BMC Struct. Biol., 2009, 9, 50.
Mizianty, M.J.; Kurgan, L. Meta prediction of protein crystallization propensity. Biochem. Biophys. Res. Commun., 2009, 390(1),
10-15.
Overton, I.M.; Barton, G.J. A normalised scale for structural genomics target ranking: the OB-Score. FEBS Lett., 2006, 580(16),
4005-4009.
Overton, I.M.; Padovani, G.; Girolami, M.A.; Barton, G.J. ParCrys:
a Parzen window density estimation approach to protein crystallization propensity prediction. Bioinformatics, 2008, 24(7), 901-907.
Overton, I.M.; van Niekerk, C.A.; Barton, G.J. XANNpred: neural
nets that predict the propensity of a protein to yield diffractionquality crystals. Proteins, 2006, 79(4), 1027-1033.
Slabinski, L.; Jaroszewski, L.; Rychlewski, L.; Wilson, I.A.; Lesley, S.A.; Godzik, A. XtalPred: a web server for prediction of protein crystallizability. Bioinformatics, 2007, 23(24), 3403-3405.
Smialowski, P.; Schmidt, T.; Cox, J.; Kirschner, A.; Frishman, D.
Will my protein crystallize? A sequence-based predictor. Proteins,
2006, 62(2), 343-355.
Kyte, J.; Doolittle, R.F. A simple method for displaying the hydropathic character of a protein. J. Mol. Biol., 1982, 157(1), 105132.
Guruprasad, K.; Reddy, B.V.; Pandit, M.W. Correlation between
stability of a protein and its dipeptide composition: a novel approach for predicting in vivo stability of a protein from its primary
sequence. Protein Eng., 1990, 4(2), 155-161.
Plewczynski, D.; Slabinski, L.; Tkacz, A.; Kajan, L.; Holm, L.;
Ginalski, K.; Rychlewski, L. The RPSP: Web server for prediction
of signal peptides. Polymer, 2007, 48(19), 5493-5496.
Krogh, A.; Larsson, B.; von Heijne, G.; Sonnhammer, E.L. Predicting transmembrane protein topology with a hidden Markov model:
application to complete genomes. J. Mol. Biol., 2001, 305(3), 567580.
Lupas, A.; Van Dyke, M.; Stock, J. Predicting coiled coils from
protein sequences. Science, 1991, 252(5010), 1162-1164.
Kurgan, L.; Mizianty, M. Sequence-based protein crystallization
propensity prediction for structural genomics: review and comparative analysis. Nat. Sci., 2009, 1, 93-106.
Dosztanyi, Z.; Csizmok, V.; Tompa, P.; Simon, I. IUPred: web
server for the prediction of intrinsically unstructured regions of

Snchez-Puig et al.

[22]
[23]

[24]
[25]

[26]

[27]
[28]

[29]

[30]

[31]

[32]

[33]
[34]

[35]

[36]
[37]

[38]

[39]
[40]
[41]

[42]
[43]
[44]

proteins based on estimated energy content. Bioinformatics, 2005,

21(16), 3433-3434.
Garbuzynskiy, S.O.; Lobanov, M.Y.; Galzitskaya, O.V. To be
folded or to be unfolded? Protein Sci., 2004, 13(11), 2871-2877.
Linding, R.; Jensen, L.J.; Diella, F.; Bork, P.; Gibson, T.J.; Russell,
R.B. Protein disorder prediction: implications for structural proteomics. Structure, 2003, 11(11), 1453-1459.
Linding, R.; Russell, R.B.; Neduva, V.; Gibson, T.J. GlobPlot:
Exploring protein sequences for globularity and disorder. Nucleic
Acids Res., 2003, 31(13), 3701-3708.
Obradovic, Z.; Peng, K.; Vucetic, S.; Radivojac, P.; Brown, C.J.;
Dunker, A.K. Predicting intrinsic disorder from amino acid sequence. Proteins, 2003, 53 Suppl 6, 566-572.
Prilusky, J.; Felder, C.E.; Zeev-Ben-Mordehai, T.; Rydberg, E.H.;
Man, O.; Beckmann, J.S.; Silman, I.; Sussman, J.L. FoldIndex: a
simple tool to predict whether a given protein sequence is intrinsically unfolded. Bioinformatics, 2005, 21(16), 3435-3438.
Romero, P.; Obradovic, Z.; Li, X.; Garner, E.C.; Brown, C.J.;
Dunker, A.K. Sequence complexity of disordered protein. Proteins,
2001, 42(1), 38-48.
Vullo, A.; Bortolami, O.; Pollastri, G.; Tosatto, S.C. Spritz: a
server for the prediction of intrinsically disordered regions in protein sequences using kernel machines. Nucleic Acids Res., 2006,
34, W164-168.
Ward, J.J.; Sodhi, J.S.; McGuffin, L.J.; Buxton, B.F.; Jones, D.T.
Prediction and functional analysis of native disorder in proteins
from the three kingdoms of life. J. Mol. Biol., 2004, 337(3), 635645.
Xue, B.; Dunbrack, R.L.; Williams, R.W.; Dunker, A.K.; Uversky,
V.N. PONDR-FIT: a meta-predictor of intrinsically disordered
amino acids. Biochim. Biophys. Acta, 2010, 1804(4), 996-1010.
Yang, Z.R.; Thomson, R.; McNeil, P.; Esnouf, R.M. RONN: the
bio-basis function neural network technique applied to the detection of natively disordered regions in proteins. Bioinformatics,
2005, 21(16), 3369-3376.
Williams, R.M.; Obradovic, Z.; Mathura, V.; Braun, W.; Garner,
E.C.; Young, J.; Takayama, S.; Brown, C.J.; Dunker, A.K. The protein non-folding problem: amino acid determinants of intrinsic order and disorder. Pac. Symp. Biocomput., 2001, 89-100.
Uversky, V.N. Cracking the folding code. Why do some proteins
adopt partially folded conformations, whereas other don't? FEBS
Lett., 2002, 514(2-3), 181-183.
Uversky, V.N.; Gillespie, J.R.; Fink, A.L. Why are "natively unfolded" proteins unstructured under physiologic conditions? Proteins, 2000, 41(3), 415-427.
Brocca, S.; Samalikova, M.; Uversky, V.N.; Lotti, M.; Vanoni, M.;
Alberghina, L.; Grandori, R. Order propensity of an intrinsically
disordered protein, the cyclin-dependent-kinase inhibitor Sic1. Proteins, 2009, 76(3), 731-746.
Sanchez-Puig, N.; Veprintsev, D.B.; Fersht, A.R. Binding of
natively unfolded HIF-1alpha ODD domain to p53. Mol. Cell,
2005, 17(1), 11-21.
Dames, S.A.; Martinez-Yamout, M.; De Guzman, R.N.; Dyson,
H.J.; Wright, P.E. Structural basis for Hif-1 alpha /CBP recognition
in the cellular hypoxic response. Proc. Natl. Acad. Sci. USA, 2002,
99(8), 5271-5276.
Hewitson, K.S.; McNeill, L.A.; Riordan, M.V.; Tian, Y.M.; Bullock, A.N.; Welford, R.W.; Elkins, J.M.; Oldham, N.J.; Bhattacharya, S.; Gleadle, J.M.; Ratcliffe, P.J.; Pugh, C.W.; Schofield, C.J.
Hypoxia-inducible factor (HIF) asparagine hydroxylase is identical
to factor inhibiting HIF (FIH) and is related to the cupin structural
family. J. Biol. Chem., 2002, 277(29), 26351-26355.
De Sancho, D.; Best, R.B. What is the time scale for alpha-helix
nucleation? J. Am. Chem. Soc., 2011, 133(17), 6809-6816.
Vekilov, P.G. What determines the rate of growth of crystals from
solution? Cryst. Growth Des., 2007, 7(12), 2796-2810.
Durbin, S.D.; Feher, G. Studies of crystal growth mechanisms of
proteins by electron microscopy. J. Mol. Biol., 1990, 212(4), 763774.
DeMattei, R.C.; Feigelson, R.S. Controlling Nucleation In Protein
Solutions. J. Cryst. Growth, 1992, 122(1-4), 21-30.
George, A.; Wilson, W.W. Predicting protein crystallization from a
dilute solution property. Acta Crystallogr. Sect D Biol. Crystallogr., 1994, 50(Pt 4), 361-365.
Kashchiev, D.; van Rosmalen, G.M. Review: Nucleation in solutions revisited. Cryst. Res. Technol., 2003, 38(7-8), 555-574.

Predicting Protein Crystallizability and Nucleation

[45]

[46]
[47]

Protein & Peptide Letters, 2012, Vol. 19, No. 7

Jurez-Martnez, G.; Garza, C.; Castillo, R.; Moreno, A. A dynamic light scattering investigation of the nucleation and growth of
thaumatin crystals. J. Cryst. Growth, 2001, 232, 119-131.
Baird, J.K.; Scott, S.C.; Kim, Y.W. Theory of the effect of pH and
ionic strength on the nucleation of protein crystals. J. Cryst.
Growth, 2001, 232(1-4), 50-62.
Michinomae, M.; Mochizuki, M.; Ataka, M. Electron microscopic
studies on the initial process of lysozyme crystal growth. J. Cryst.
Growth, 1999, 197(1-2), 257-262.

Received: August 8, 2011

Revised: August 29, 2011

Accepted: February 11, 2012

[48]

[49]
[50]

731

Tanaka, S.; Ito, K.; Hayakawa, R.; Ataka, M. Size and number
density of precrystalline aggregates in lysozyme crystallization
process. J. Chem. Phys., 1999, 111(22), 10330-10337.
Georgalis, Y.; Schuler, J.; Frank, J.; Soumpasis, M.D.; Saenger, W.
Protein crysatllization screening through scattering techniques.
Adv. Colloid Interface Sci., 1995, 58(1), 57-86.
Niimura, N.; Minezaki, Y.; Ataka, M.; Katsura, T. Aggregation in
supersaturated lysozyme solutions studied by time-resolved smallangle neutron scattering. J. Cryst. Growth, 1995, 154(1-2), 136144.