Date of Award
Doctor of Philosophy
Chemical and Biological Engineering
Charles E. Glatz
Strategies to optimize the recovery of recombinant proteins from plant hosts have been investigated. The use of different crops, different product molecules, and different separation techniques was studied to suggest modes of operation that reduce the overall processing cost for obtaining purified recombinant proteins from plants.;Polyelectrolyte precipitation of beta-glucuronidase (GUS), an acidic model protein, with polyethyleneimine (PEI), a polycatonic polymer, from the extracts of canola, corn germ, soybean, and pea met with varying degrees of success. Precipitation from canola required the least amount of PEI and was capable of providing the highest enrichment. In contrast, soy and pea allowed for only slight enrichment. These results suggest that for ease of purification by PEI precipitation (and likely other charged based separations), the matrix components (proteins, carbohydrates, phenolics, lipids,) of canola are most suited for GUS (and likely other proteins with isoelectric point <7), while soy and pea may be more advantageous as a host for basic recombinant proteins.;Expanded bed adsorption (EBA) processed a crude plant extract if the proper combination of column inlet design, resin, operating conditions and crude feed properties was chosen. The chemistry of the EBA resin ultimately dictated what size of corn endosperm solids could be eluted during the expanded bed adsorption process. Application with DEAE and SP resins with higher density, capable of operating at higher flow velocities (>450 cm/h) were not affected by solids of <44 mum. However, processes employing Streamline resins with the same functionality, requiring a lower superficial velocity (180 cm/h), could not completely elute solids of <44 mum.;The large size (390 kDa) of the target antibody to be captured in this study resulted in extreme internal diffusion limitations during adsorption. Smaller resin particles allowed enhanced uptake rates of the protein, but breakthrough performance was poor for all resins evaluated. A process simulation analysis for recovery of the antibody showed that complete clarification of the corn endosperm extract, followed by packed bed operation, would require approximately the same capital expenditure as the EBA process. However, the packed bed mode would require roughly one-fourth the operating expenditure.
Digital Repository @ Iowa State University, http://lib.dr.iastate.edu
Todd J. Menkhaus
Menkhaus, Todd J., "Recovery of recombinant proteins from transgenic plants: strategies in implementation and optimization " (2002). Retrospective Theses and Dissertations. 1417.