Purification of an acidic recombinant protein from transgenic tobacco.
Tobacco has proven to be a promising alternative for the production of recombinant therapeutic proteins and offers numerous advantages over other plants as a host system. However, the recovery and purification steps needed to obtain a protein at high recovery and purity have not been well investigated. In this study, a process was developed to purify a model acidic protein, recombinant beta-glucuronidase (rGUS) from transgenic tobacco leaf tissue, in three main steps after extraction: polyelectrolyte precipitation, hydrophobic interaction chromatography (HIC), and hydroxyapatite chromatography (HAC). Using this three-step process, up to 40% of the initial rGUS activity could be recovered to near homogeneity as judged by SDS-PAGE. This work demonstrates that acidic recombinant proteins expressed in tobacco may be purified to high yield with high purity in a minimal amount of steps that are suitable for scale-up. Furthermore, the general steps used in this process may suggest that a wide variety of acidic recombinant proteins may be purified in a similar manner from transgenic tobacco or other leafy crops.
Holler
Chris
C
Department of Biological Systems Engineering, Virginia Polytechnic Institute and State University, 210 Seitz Hall, Blacksburg, Virginia 24061, USA.
Zhang
Chenming
C
D002845:Chromatography / Q000379:methods; D005966:Glucuronidase / Q000302:isolation & purification* / Q000502:physiology*; D018515:Plant Leaves / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D014026:Nicotiana / Q000502:physiology*