A chloroplast transgenic approach to hyper-express and purify Human Serum Albumin, a protein highly susceptible to proteolytic degradation
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
"A chloroplast transgenic approach to hyper-express and purify Human Serum Albumin, a protein highly susceptible to proteolytic degradation" (2003). Faculty Bibliography 2000s. 3746.