Identifying solubility-promoting buffers for intrinsically disordered proteins prior to purification. | Academic Article individual record
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.

authors
author list (cited authors)
Churion, K. A., & Bondos, S. E.
publication date
2012
publisher
keywords
  • Proteins
  • Buffers
  • Protein Solubility
  • Chaperone
  • Kosmotrope
  • Blotting, Western
  • Chaotrope
  • Solubility
  • Electrophoresis, Polyacrylamide Gel
  • Filter-based Aggregation Assay
  • Purification
  • Precipitation
  • Osmolyte
  • Chemical Fractionation
  • Filtration
altmetric score

1.0

citation count

8