Purification of chimeric amine dehydrogenase using a tailor-made aqueous two-phase system - A case study

Abstract Enzymes used as biocatalysts in evolving green processes have already reached industrial competitiveness to classical chemical catalysts. After production of these enzymes, aqueous two-phase extraction using an aqueous two-phase system (ATPS) has been shown a promising and sustainable alternative to common cost-intense chromatographic purification. However, state-of-the-art ATPS design is based on “trial-and-error” screening approaches leading to un-optimized ATPS. Within this work, we demonstrate the applicability of our method for ATPS design to an industrially relevant chimeric amine dehydrogenase (cFL1). The method includes thermodynamic modeling of ATPS phase composition and investigations on the conformational and colloidal cFL1 stability to rapidly select a tailor-made ATPS, offering optimal process and processing conditions, outclassing the previously selected ATPS (identified by random screening) by a factor of 4. All investigations and the selection of the sodium glutamate-polyethylene glycol 2000 ATPS were performed in less than 4 weeks, demonstrating proof of concept for a fast and sustainable ATPS selection in an early stage of downstream processing development.