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Plant Biotechnol J. 2019 Jun;17(6):1094-1105. doi: 10.1111/pbi.13040. Epub 2018 Dec 8.

Cost-effective production of tag-less recombinant protein in Nicotiana benthamiana.

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Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology, Pohang, Korea.
Department of Life Science, Pohang University of Science and Technology, Pohang, Korea.


Plants have recently received a great deal of attention as a means of producing recombinant proteins. Despite this, a limited number of recombinant proteins are currently on the market and, if plants are to be more widely used, a cost-effective and efficient purification method is urgently needed. Although affinity tags are convenient tools for protein purification, the presence of a tag on the recombinant protein is undesirable for many applications. A cost-effective method of purification using an affinity tag and the removal of the tag after purification has been developed. The family 3 cellulose-binding domain (CBM3), which binds to microcrystalline cellulose, served as the affinity tag and the small ubiquitin-related modifier (SUMO) and SUMO-specific protease were used to remove it. This method, together with size-exclusion chromatography, enabled purification of human interleukin-6 (hIL6) with a yield of 18.49 mg/kg fresh weight from leaf extracts of Nicotiana benthamiana following Agrobacterium-mediated transient expression. Plant-produced hIL6 (P-hIL6) contained less than 0.2 EU/μg (0.02 ng/mL) endotoxin. P-hIL6 activated the Janus kinase-signal transducer and activator of transcriptional pathways in human LNCaP cells, and induced expression of IL-21 in activated mouse CD4+ T cells. This approach is thus a powerful method for producing recombinant proteins in plants.


Nicotiana benthamiana ; bdSENP1; bdSUMO; cellulose-binding domain; human interleukin-6; plant-based expression system; proteolytic tag removal

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