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

International Tables for Crystallography (2012). Vol. F, ch. 5.2, p. 152   | 1 | 2 |

Section 5.2.2. Solvent in macromolecular crystals

E. M. Westbrooka*

aMolecular Biology Consortium, Argonne, Illinois 60439, USA
Correspondence e-mail:

5.2.2. Solvent in macromolecular crystals

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Crystals of biological macromolecules differ from crystals of smaller molecules in that a significant fraction of their volume is occupied by solvent (Adair & Adair, 1936[link]; Perutz, 1946[link]; Crick, 1957[link]). This solvent is not homogeneous: a part binds tightly to the macromolecule as a hydration shell, and the remainder remains free, indistinguishable from the solvent surrounding the crystal.

Hydration is essential for macromolecular stability: bound solvent is part of the complete macromolecule's structure (Tanford, 1961[link]). Diffraction-based studies of macromolecular crystals verify the presence of well defined bound solvent. Typically, 8–10% of the atomic coordinates in each Protein Data Bank file are those of bound water molecules. The consensus observation of protein hydration (Adair & Adair, 1936[link]; Perutz, 1946[link]; Edsall, 1953[link]; Coleman & Matthews, 1971[link]; Kuntz & Kaufmann, 1974[link]; Scanlon & Eisenberg, 1975[link]) is that every gram of dry protein is hydrated by 0.2–0.3 g of water: this is consistent both with the presence of a shell of hydration, the thickness of which is about one water molecule (2.5–3 Å), and with the rule-of-thumb that approximately one water molecule is found for every amino-acid residue in the protein's crystal structure. Matthews (1974[link]) suggests setting this hydration ratio, w, to 0.25 g water per gram of protein as a reasonable estimate for typical protein crystals.

Crystallographic structures also exhibit empty regions of `free' solvent. Such voids are to be expected: closely packed spheres occlude just 74% of the space they occupy, so to the extent that proteins are spherical, tight packing in their crystals would leave 26% of the crystal volume for free solvent. Although the distinction between free and bound solvent is not sharp (solvent-binding-site occupancies vary, as do their refined B factors), it is a useful convention and is consistent with many observed physical properties of these crystals.


Adair, G. S. & Adair, M. E. (1936). The densities of protein crystals and the hydration of proteins. Proc. R. Soc. London Ser. B, 120, 422–446.
Coleman, P. M. & Matthews, B. W. (1971). Symmetry, molecular weight, and crystallographic data for sweet potato β-amylase. J. Mol. Biol. 60, 163–168.
Crick, F. (1957). X-ray diffraction of protein crystals. Methods Enzymol. 4, 127–146.
Edsall, J. T. (1953). Solvation of proteins. In The Proteins, edited by H. Neurath & K. Bailey, Vol. 1, part B, pp. 549–726. New York: Academic Press.
Kuntz, I. D. & Kaufmann, W. (1974). Hydration of proteins and polypeptides. Adv. Protein Chem. 28, 239–345.
Matthews, B. W. (1974). Determination of molecular weight from protein crystals. J. Mol. Biol. 82, 513–526.
Perutz, M. F. (1946). The solvent content of protein crystals. Trans. Faraday Soc. 42B, 187–195.
Scanlon, W. J. & Eisenberg, D. (1975). Solvation of crystalline proteins: theory and its application to available data. J. Mol. Biol. 98, 485–502.
Tanford, C. (1961). Physical Chemistry of Macromolecules. New York: John Wiley & Sons.

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