Abbreviated Journal Title
Plant Biotechnol. J.
chloroplast genetic engineering; biopharmaceuticals; genetically; modified crops; molecular farming; recombinant human blood proteins; RECOMBINANT HUMAN ALBUMIN; FOREIGN GENE-EXPRESSION; PSBA MESSENGER-RNA; TOBACCO CHLOROPLASTS; PLANTS; TRANSFORMATION; TRANSLATION; PLASTIDS; GENOME; OVEREXPRESSION; Biotechnology & Applied Microbiology; Plant Sciences
Human Serum Albumin (HSA) accounts for 60% of the total protein in blood serum and it is the most widely used intravenous protein in a number of human therapies. HSA, however, is currently extracted only from blood because of a lack of commercially feasible recombinant expression systems. HSA is highly susceptible to proteolytic degradation in recombinant systems and is expensive to purify Expression of HSA in transgenic chloroplasts using Shine-Dalgarno sequence (SD), which usually facilitates hyper-expression of transgenes, resulted only in 0.02% HSA in total protein (tp). Modification of HSA regulatory sequences using chloroplast untranslated regions (UTRs) resulted in hyper-expression of HSA (up to 11.1% tp), compensating for excessive proteolytic degradation. This is the highest expression of a pharmaceutical protein in transgenic plants and 500-fold greater than previous reports on HSA expression in transgenic leaves. Electron micrographs of immunogold labelled transgenic chloroplasts revealed HSA inclusion bodies, which provided a simple method for purification from other cellular proteins. HSA inclusion bodies could be readily solubilized to obtain a monomeric form using appropriate reagents. The regulatory elements used in this study should serve as a model system for enhancing expression of foreign proteins that are highly susceptible to proteolytic degradation and provide advantages in purification, when inclusion bodies are formed.
Plant Biotechnology Journal
Fernández-San Millán, Alicia; Mingo-Castel, Angel; Miller, Michael; and Daniell, Henry, "A chloroplast transgenic approach to hyper-express and purify Human Serum Albumin, a protein highly susceptible to proteolytic degradation" (2003). Faculty Bibliography 2000s. 3746.