A chloroplast transgenic approach to hyper-express and purify human serum albumin, a protein highly susceptible to proteolytic degradation
Fecha
2003Versión
Acceso abierto / Sarbide irekia
Tipo
Artículo / Artikulua
Versión
Versión publicada / Argitaratu den bertsioa
Impacto
|
10.1046/j.1467-7652.2003.00008.x
Resumen
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. E ...
[++]
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. [--]
Materias
Chloroplast genetic engineering,
Biopharmaceuticals,
Genetically modified crops,
Molecular farming,
Recombinant human blood proteins
Editor
Wiley / Blackwell
Publicado en
Plant Biotechnology Journal, (2003) 1, pp. 71–79
Departamento
Universidad Pública de Navarra/Nafarroako Unibertsitate Publikoa. IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Versión del editor
Entidades Financiadoras
This study was supported in part by grants from NIH RO1 GM 63879 and Chlorgen Inc. to H.D., and ‘Dirección General de Industria, Gobierno de Navarra’ (Spain) to A.M.C.