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Circulating Fluidized Beds for Continuous Protein Refolding


Many recombinant expression systems deposit their valuable heterologous protein products in the form of insoluble inclusion bodies. These must be separated from cell debris, resolubilized by denaturing the proteins and then refolded into the native configuration in suitable buffers. This refolding is often facilitated by binding the protein as it is solubilized to ion exchange resin or other adsorbents (matrix-assisted refolding). The protein must be eluted from the adsorbent for further purification and the adsorbent often requires regeneration before reuse in a subsequent cycle of operation.

The cleanup, binding and elution steps are usually done in batch mode, with the aid of expanded bed chromatography columns. Continuous production is only possible by repeating the batches.

Description of the Invention:

It has been proven feasible to conduct the various steps of heterologous protein refolding in a unique arrangement of connected vessels, each handling one of the steps. Employing principles of circulating fluidized beds, the technology allows continuous and simple adsorption, desorption and adsorbent regeneration, without mixing of buffers from the various stages. The technology handles the adsorbent at gentle velocities, minimizing the attrition often seen from repeated batch handling.

Potential Advantages:

• Continuous handling of heterologous inclusion proteins
• Multi-stage treatment of proteins with minimum exposure times in closed vessels
• Adaptable to other multiple-step protein purification or treatment systems
• Unlimited scale-up capability

Potential Applications:

• Biopharmaceuticals
• Recombinant protein recovery and refolding
• Multi-step enzyme purification
• Cellular debris removal in downstream processing.

Patent Information:

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