Improved renaturation process of aggregated recombinant proteins through the design of hydrated ionic liquids

Abstract

Escherichia coli is the most commonly used host for recombinant protein expression. However, since inactive aggregation is obtained at a high rate and the renaturation rate is still low, the improvement of the treatment of aggregated recombinant proteins is a critical issue. This study aimed to evaluate the use of hydrated ionic liquids (ILs) to directly dissolve aggregated recombinant proteins and induce refolding. Selection of component ions and suitable water content resulted in a considerable improvement in the dissolution of the aggregated protein, followed by refolding behaviour. Ammonium or phosphonium cations with butyl groups coupled with dihydrogen phosphate anions were identified as appropriate ion pairs, showing high solubility and contributing to refolding. The column method showed great potential in the transition of dissolved and refolded proteins in hydrated ILs into the buffer solution, achieving both IL removal and protein purification. The successfully recovered activity levels of aggregated proteins indicate that this method is a promising approach for the development of novel treatments for aggregated recombinant proteins.