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Intrinsically Disordered Protein Analysis pp 415–427Cite as

Identifying Solubility-Promoting Buffers for Intrinsically Disordered Proteins Prior to Purification

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 896))

Abstract

Intrinsically disordered proteins are anticipated to be more prone to aggregation than folded, stable proteins. Chemical additives included in the buffer can help maintain proteins in a soluble, monomeric state. However, the array of chemicals that impact protein solubility is staggering, precluding iterative testing of chemical conditions during purification. Herein, we describe a filter-based aggregation assay to rapidly identify chemical additives that maintain solubility for a protein of interest. A hierarchical approach to buffer selection is provided, in which the type of chemical which best improves solubility is first determined, followed by identifying the optimal chemical and its most effective concentration. Finally, combinations of chemical additives can be assessed if necessary. Although this assay can be applied to purified protein, partially purified protein, or aggregated protein, this protocol specifically details the use of this assay for crude cell lysate. This approach allows identification of solubility-promoting buffers prior to the initial protein purification.

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Acknowledgments

This work was supported by grants from the American Heart Association 422351 and the Texas A&M Health Science Center to SEB.

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Authors and Affiliations

  1. Department of Molecular and Cellular Medicine, Texas A&M Health Science Center, College Station, TX, USA

    Kelly A. Churion & Sarah E. Bondos

Authors
  1. Kelly A. Churion
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  2. Sarah E. Bondos
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Corresponding author

Correspondence to Sarah E. Bondos .

Editor information

Editors and Affiliations

  1. DEPT OF MOLECULAR MED, College of Medicine, University of South Florida, BRUCE B DOWNS BLVD 12901, TAMPA, 33612, Florida, USA

    Vladimir N. Uversky

  2. Dept. Biochemistry & Molecular Biology, Center for Computational Biology &, Indiana University, West 10th Street 410, Indianapolis, 46202, Indiana, USA

    A. Keith Dunker

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Churion, K.A., Bondos, S.E. (2012). Identifying Solubility-Promoting Buffers for Intrinsically Disordered Proteins Prior to Purification. In: Uversky, V., Dunker, A. (eds) Intrinsically Disordered Protein Analysis. Methods in Molecular Biology, vol 896. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-3704-8_28

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  • DOI: https://doi.org/10.1007/978-1-4614-3704-8_28

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  • Publisher Name: Springer, New York, NY

  • Print ISBN: 978-1-4614-3703-1

  • Online ISBN: 978-1-4614-3704-8

  • eBook Packages: Springer Protocols

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