Purification of Recombinant Human Serum Albumin from Transgenic Rice Extract

Abstract. Transgenic plant systems have successfully been used to express a variety of recombinant proteins, including rice seed-expressed recombinant human serum albumin (rHSA). The development of an efficient and integrated rHSA extraction and purification process would allow rHSA to be used for various medical applications such as a stabilizer in vaccinations without the risk of contamination of pathogens and viruses. Extraction is a critical step that determines recombinant protein concentration and purity and the type and quantity of impurities that must be removed during purification. Previous studies indicated that pH 3.5 extraction resulted in high purity (78%) but rHSA was unstable and degraded over time. Extraction at pH 4.5 and 6.0 provided stable rHSA yields but higher native rice protein concentrations, which would increase purification requirements. To develop an integrated process, the effects of extraction conditions on subsequent purification using ion exchange chromatography must be evaluated. The impact of extraction conditions on purification efficiency were evaluated through adsorption kinetics with pure rHSA. Once the optimal conditions for rHSA were identified, adsorption of rHSA in transgenic rice extracts were evaluated. Anion exchange chromatography with Q Sepharose FastFlow^® resin was selected because of the acidic pI of rHSA at 4.7. A low buffering capacity of 25 mM NaAcetate at pH 8 and conductivity of 5 mS/cm was chosen to improve binding of rHSA and resin using the ÄKTA Pure Chromatography System. Purification results of rHSA were analyzed for the development of an integrated, cost-efficient downstream process.