International
Tables for
Crystallography
Volume F
Crystallography of biological macromolecules
Edited by M. G. Rossmann and E. Arnold

© International Union of Crystallography 2006
International Tables for Crystallography (2006). Vol. F. ch. 4.1, p. 81   | 1 | 2 |

Section 4.1.2.1. General considerations

R. Giegéa* and A. McPhersonb

a Unité Propre de Recherche du CNRS, Institut de Biologie Moléculaire et Cellulaire, 15 rue René Descartes, F-67084 Strasbourg CEDEX, France, and bDepartment of Molecular Biology & Biochemistry, University of California at Irvine, Irvine, CA 92717, USA
Correspondence e-mail:  R.Giege@ibmc.u-strasbg.fr

4.1.2.1. General considerations

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Many methods can be used to crystallize macromolecules (McPherson, 1982[link], 1998[link]; Ducruix & Giegé, 1999[link]), the objectives of which bring the macromolecules to an appropriate state of supersaturation. Although vapour-phase equilibrium and dialysis techniques are favoured, batch and free interface diffusion methods are often used (Fig. 4.1.2.1[link]). Besides the current physical and chemical parameters that affect crystallization (Table 4.1.2.1[link]), macromolecular crystal growth is affected by the crystallization method itself and the geometry of the arrangements used. Generally, in current methods, growth is promoted by the non-equilibrium nature of the crystallization process, which seldom occurs at constant protein concentration. This introduces changes in supersaturation and hence may lead to changes in growth mechanism. Crystallization at constant protein concentration can, however, be achieved in special arrangements based on liquid circulation cells (Vekilov & Rosenberger, 1998[link]).

Table 4.1.2.1 | top | pdf |
Factors affecting crystallization

Physical Chemical Biochemical
Temperature variation pH Purity of the macromolecule or impurities
Surface Precipitant type Ligands, inhibitors, effectors
Methodology or approach to equilibrium Precipitant concentration Aggregation state of the macromolecule
Gravity Ionic strength Post-translational modifications
Pressure Specific ions Source of macromolecule
Time Degree of supersaturation Proteolysis or hydrolysis
Vibrations, sound or mechanical perturbations Reductive or oxidative environment Chemical modifications
Electrostatic or magnetic fields Concentration of the macromolecules Genetic modifications
Dielectric properties of the medium Metal ions Inherent symmetry of the macromolecule
Viscosity of the medium Crosslinkers or polyions Stability of the macromolecule
Rate of equilibration Detergents, surfactants or amphophiles Isoelectric point
Homogeneous or heterogeneous nucleants Non-macromolecular impurities History of the sample
[Figure 4.1.2.1]

Figure 4.1.2.1 | top | pdf |

Principles of the major methods currently used to crystallize biological macromolecules. (a) Batch crystallization in three versions. (b) Dialysis method with Cambridge button. (c) Vapour diffusion crystallization with hanging and sitting drops. (d) Interface crystallization in a capillary and in an arrangement for assays in microgravity. The evolution of the concentration of macromolecule, [M], and crystallizing agent (precipitant), [CA], in the different methods is indicated (initial and final concentrations in the crystallization solutions are [M]i, [M]f, [CA]i and [CA]f, respectively; [CA]res is the concentration of the crystallizing agent in the reservoir solution, and k is a dilution factor specified by the ratio of the initial concentrations of crystallizing agent in the drop and the reservoir). In practice, glass vessels in contact with macromolecules should be silicone treated to obtain hydrophobic surfaces.

References

First citation Ducruix, A. & Giegé, R. (1999). Editors. Crystallization of proteins and nucleic acids: a practical approach, 2nd ed. Oxford: IRL Press.Google Scholar
 First citation McPherson, A. (1982). The preparation and analysis of protein crystals. New York: John Wiley and Sons.Google Scholar
 First citation McPherson, A. (1998). Crystallization of biological macromolecules. Cold Spring Harbor and New York: Cold Spring Harbor Laboratory Press.Google Scholar
 First citation Vekilov, P. G. & Rosenberger, F. (1998). Protein crystal growth under forced solution flow: experimental setup and general response of lysozyme. J. Cryst. Growth, 186, 251–261.Google Scholar








































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