Protein phase behavior in aqueous solutions: Crystallization, liquid-liquid phase separation, gels, and aggregates

André C. Dumetz, Aaron M. Chockla, Eric W. Kaler, Abraham M. Lenhoff

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209 Scopus citations


The aggregates and gels commonly observed during protein crystallization have generally been considered disordered phases without further characterization. Here their physical nature is addressed by investigating protein salting-out in ammonium sulfate and sodium chloride for six proteins (ovalbumin, ribonuclease A, soybean trypsin inhibitor, lysozyme, and β-lactoglobulin A and B) at 4°C, 23°C, and 37°C. When interpreted within the framework of a theoretical phase diagram obtained for colloidal particles displaying short-range attractive interactions, the results show that the formation of aggregates can be interpreted theoretically in terms of a gas-liquid phase separation for aggregates that are amorphous or gel-like. A notable additional feature is the existence of a second aggregation line observed for both ovalbumin and ribonuclease A in ammonium sulfate, interpreted theoretically as the spinodal. Further investigation of ovalbumin and lysozyme reveals that the formation of aggregates can be interpreted, in light of theoretical results from mode-coupling theory, as a kinetically trapped state or a gel phase that occurs through the intermediate of a gas-liquid phase separation. Despite the limitations of simple theoretical models of short-range attractive interactions, such as their inability to reproduce the effect of temperature, they provide a framework useful to describe the main features of protein phase behavior.

Original languageEnglish (US)
Pages (from-to)570-583
Number of pages14
JournalBiophysical journal
Issue number2
StatePublished - Jan 15 2008

Bibliographical note

Funding Information:
We are grateful for support from the National Science Foundation (grant No. BES-0519191).


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