(Wiley / Blackwell, 2003) Fernández San Millán, Alicia; Mingo Castel, Ángel; Miller, Michael; Daniell, Henry; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua
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.
