Chromatography is the heart of protein purification and the development of the chromatography process is facilitated by automated chromatography systems. Purification protocols often follow the general three step scheme of capture, intermediate and final polish. In this case study, we present the process development for a recombinant E. coli protein using new generation chromatography resins. The target protein has an acidic pI and is expressed at high levels as a soluble, properly folded intracellular protein. Thus, the lysate is full of nucleic acids and cytosolic contaminants, which are also generally acidic. Using an automated chromatography system, we screened four anion exchange resins and compared their capture capacity verses a dextran/agarose resin. A hydrophilic epoxide AEX resin with large pores provided the highest binding capacity and recovery. The capture step was then scaled up from 1 ml resin to 100 ml resin. The intermediate purification step was developed by screening HIC resins and the final polish step was developed using a second anion exchange chromatography resin.

Development of an efficient purification process for a recombinant <i>E. coli</i> protein

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

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Development of an efficient purification process for a recombinant E. coli protein