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Global Journal of Analytical Chemistry. Volume 2, Issue 4 (2011) pp. 152-167
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Review Article
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The mechanism of protein precipitation by salts, polymers and organic solvents
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Tsutomu Arakawaa, Yoshiko Kitab, Kentaro Shirakic, Satoshi Ohtaked
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a Alliance Protein Laboratories, Thousand Oaks, CA 91360, USA
b Department of Pharmacology, KEIO University School of Medicine, Tokyo 160-8582, Japan
c Institute of Applied Physics, University of Tsukuba, Tsukuba, Ibaraki 305-8573, Japan
d Aridis Pharmaceuticals, 5941 Optical Court, San Jose, CA 95138, USA
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Abstract |
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Precipitation of proteins by co-solvents is a key technology for protein purification and crystallization. Three classes of co-solvents, i.e., salting-out salts, polymers, and organic solvents, are primarily used to reduce protein solubility, leading to precipitation at sufficiently high concentrations. Salts, in particular ammonium sulfate, have been extensively used in differential precipitation and crystallization. Polymers, such as high molecular weight polyethylene glycol, have also been used for protein precipitation and crystallization ever since the discovery of salting-out effects by Hofmeister. Despite being a protein denaturant, organic solvents including various alcohols, acetone, and dioxane, can effectively precipitate proteins in the native state. The mechanism of salting-out effect of salts is mainly ascribed to their influence on water structure and surface tension. Polymers exert their effects mainly through excluded volume while organic solvents cause protein precipitation by charge repulsion. These mechanisms may be oversimplified, however, as their effects on protein solubility also depend to varying degrees on the chemical properties of the protein surface. In this review, we will summarize the available data that shed light on the interactions of co-solvents with the proteins and describe the mechanisms responsible for causing precipitation.
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Keywords |
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Protein precipitation; Attraction pressure; Cavity theory; Excluded volume; Charge repulsion
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