Preparation of Protein Crystals for X-Ray Structural Study

  • Soichi Takeda
Part of the Methods in Molecular Medicine book series (MIMM, volume 129)


The knowledge of accurate molecular structures obtained by X-ray protein crystallography is now inevitable for rational drug design and for understanding the molecular basis underlying genetic disorders found in patients. However, preparing protein crystals suitable for structural analysis is currently the bottleneck in structure determination by this method. The intent of this chapter is to present current methods of preparing protein crystals for structural studies for a wide range of biologists who have access to macromolecules but do not know how to handle them for crystallization. The chapter includes the pretreatment of a protein prior to the crystallization experiment, initial screens, and optimization of the crystallization conditions for further X-ray study. Finally, handling considerations that are important for a protein intended for crystallization experiments are discussed.

Key Words

X-ray crystallography structural biology crystallization protein structure rational drug design synchrotron 


  1. 1.
    Drenth, J. (1994) Principles of Protein X-Ray Crystallography. (Cantor, C.R., ed.), Springer-Verlag, New York.Google Scholar
  2. 2.
    Blundell, T. L. and Johnson, L. N. (eds.) (1976) Protein Crystallography. Academic, Inc., San Diego.Google Scholar
  3. 3.
    Blow, D. (ed.) (2002) Outline of Crystallography for Biologists. Oxford University Press, New York.Google Scholar
  4. 4.
    Ducruix, A. and Giege, R. (eds.) (1999) Crystallization of Nucleic Acids and Proteins: A Practical Approach. Oxford University Press, New York.Google Scholar
  5. 5.
    McPherson, A. (ed.) (1999) Crystallization of Biological Macromolecules. Cold Spring Harbor Laboratory, New York.Google Scholar
  6. 6.
    Terese, M. B. (ed.) (1999) Protein Crystallization: Techniques, Strategies, and Tips. International University Line, La Jolla.Google Scholar
  7. 7.
    McRee, D. E. (ed.) (1999) Practical Protein Crystallography. Academic, San Diego.Google Scholar
  8. 8.
    Jancarik, J. and Kim, S. H. (1991) Sparse matrix sampling: a screening method for crystallization of protein. J. Appl. Cryst. 24, 409–411.CrossRefGoogle Scholar
  9. 9.
    Vassylyev, D. G., Takeda, S., Wakatsuki, S., Maeda, K., and Maeda, Y. (1998) Crystal structure of troponin C in complex with troponin I fragment at 2.3-A resolution. Proc. Natl. Acad. Sci. USA 95, 4847–4852.CrossRefPubMedGoogle Scholar
  10. 10.
    Takeda, S., Yamashita, A., Maeda, K., and Maeda, Y. (2003) Structure of the core domain of human cardiac troponin in the Ca(2+)-saturated form. Nature 424, 35–41.CrossRefPubMedGoogle Scholar
  11. 11.
    Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.CrossRefPubMedGoogle Scholar
  12. 12.
    Takeda, S., Kobayashi, T., Taniguchi, H., Hayashi, H., and Maeda, Y. (1997) Structural and functional domains of the troponin complex revealed by limited digestion. Eur. J. Biochem. 246, 611–617.CrossRefPubMedGoogle Scholar
  13. 13.
    Pace, C. N., Vajdos, F., Fee, L., Grimsley, G., and Gray, T. (1995) How to measure and predict the molar absorption coefficient of a protein. Protein Sci. 4, 2411–2423.CrossRefPubMedGoogle Scholar

Copyright information

© Humana Press Inc. 2006

Authors and Affiliations

  • Soichi Takeda
    • 1
  1. 1.Department of Cardiac PhysiologyNational Cardiovascular Research InstituteJapan

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