Analytical Ultracentrifugation: A Valuable Tool to Recognize Crystallization Conditions of Proteins

  • Joachim Behlke
  • Otto Ristau


The use of analytical ultracentrifugation to elucidate crystallization conditions for proteins is discussed. The methods are based on careful enhancement of the protein concentration by sedimentation, leading to a partial formation of associates. The results can be monitored either from the broadening of the moving boundary in a Schlieren pattern or from the considerable concentration increase near the cell base. The conditions under which highly charged proteins are able to undergo a self-association process (nucleation) as prerequisite for crystallization can be determined from the second virial coefficient, to which the excluded volume and the net charge mainly contribute. Crystallization experiments are successful when the repulsive forces of charged proteins are screened by addition of neutral salts and the net charge contribution becomes smaller than that of the excluded volume. Protein crystallization can be considered as an open association event with binding constants in the millimolar range. An exact description of single steps by using solutions of the Lamm differential equation is not possible because of the unknown association kinetics and the large number of parameters that must be estimated. However, based on fitting of simulated concentration profiles we can exclude a statistical binding mode for nucleation.


Sedimentation Velocity Virial Coefficient Crystallization Condition Analytical Ultracentrifugation Sedimentation Equilibrium 
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Copyright information

© Springer Science+Business Media New York 2001

Authors and Affiliations

  • Joachim Behlke
    • 1
  • Otto Ristau
    • 1
  1. 1.Max Delbrück Center for Molecular MedicineBerlinGermany

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