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Phase knowledge enables rational screens for protein crystallization

Anderson, Megan J. and Hansen, Carl L. and Quake, Stephen R. (2006) Phase knowledge enables rational screens for protein crystallization. Proceedings of the National Academy of Sciences of the United States of America, 103 (45). pp. 16746-16751. ISSN 0027-8424. PMCID PMC1636526. doi:10.1073/pnas.0605293103.

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We show that knowledge of the phase behavior of a protein allows one to create a rational screen that increases the success rate of crystallizing challenging proteins. The strategy is based on using microfluidics to perform large numbers of protein solubility experiments across many different chemical conditions to identify reagents for crystallization experiments. Phase diagrams were generated for the identified reagents and used to design customized crystallization screens for every protein. This strategy was applied with a 75% success rate to the crystallization of 12 diverse proteins, most of which failed to crystallize when using traditional techniques. The overall diffraction success rate was 33%, about double what was achieved with conventional automation in large-scale protein structure consortia. The higher diffraction success rates are achieved by designing customized crystallization screens using the phase behavior information for each target. The identification of reagents based on an understanding of protein solubility and the use of phase diagrams in the design of individualized crystallization screens therefore promotes high crystallization rates and the production of diffraction-quality crystals.

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Additional Information:© 2006 by the National Academy of Sciences. Edited by Stephen L. Mayo, California Institute of Technology, Pasadena, CA, and approved September 21, 2006 (received for review June 23, 2006). Published online before print October 30, 2006, 10.1073/pnas.0605293103. We thank the laboratories of Frances Arnold, David Chan, Douglas Rees, Raymond Deshaies, Pamela Bjorkman, Axel Brunger, James Berger, Stefan Habelitz, Michael Merckel, Lee Hood, and Manal Swairjo for generously providing the materials used in this work. This work was supported by the National Institute of Health Director's Pioneer Award. Author contributions: M.J.A., C.L.H., and S.R.Q. designed research; M.J.A. performed research; C.L.H. contributed new reagents/analytic tools; M.J.A., C.L.H; and S.R.Q. analyzed data; and M.J.A., C.L.H., and S.R.Q. wrote the paper. The authors declare no conflict of interest. This article is a PNAS direct submission.
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Subject Keywords:microfluidics, protein phase behavior
Issue or Number:45
PubMed Central ID:PMC1636526
Record Number:CaltechAUTHORS:ANDpnas06
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:8514
Deposited By: Archive Administrator
Deposited On:17 Aug 2007
Last Modified:08 Nov 2021 20:50

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