title: Expression, subcellular localization, and enzyme activity of a recombinant human extra-cellular superoxide dismutase in tobacco (Nicotiana benthamiana L.).
abstract: Human extracellular superoxide dismutase (hEC-SOD) is an enzyme that scavenges reactive oxygen species (ROS). Because of its antioxidant activity, hEC-SOD has been used as a therapeutic protein to treat skin disease and arthritis in mammalian systems. In this study, codon-optimized hEC-SOD was expressed in tobacco (Nicotiana benthamiana L.) via a plant-based transient protein expression system. Plant expression binary vectors containing full-length hEC-SOD (f-hEC-SOD) and modified hEC-SOD (m-hEC-SOD), in which the signal peptide and heparin-binding domain were deleted, were constructed for the cytosolic-, endoplasmic reticulum (ER)-, and chloroplast-localizations in tobacco leaf mesophyll cells. The results demonstrated that f-hEC-SOD was more efficiently expressed in the cytosolic fractions than in the ER or chloroplasts of tobacco cells. Our data further indicated that differently localized f-hEC-SOD and m-hEC-SOD displayed SOD enzyme activities, suggesting that the hEC-SODs expressed by plants may be functionally active. The f-hEC-SOD was expressed up to 3.8% of the total leaf soluble protein and the expression yield was calculated to be 313.7 μg f-hEC-SOD per g fresh weight of leaf. Overall, our results reveal that it was possible to express catalytically active hEC-SODs by means of a transient plant expression system in tobacco leaf cells.
authors.affiliation: Department of Systems Biology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Republic of Korea.
authors.forename: Ki Youl
authors.initials: KY
authors.lastname: Park
authors.affiliation: Department of Systems Biology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Republic of Korea.
authors.forename: Eun Yu
authors.initials: EY
authors.lastname: Kim
authors.affiliation: Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Republic of Korea.
authors.forename: Weontae
authors.initials: W
authors.lastname: Lee
authors.affiliation: Laboratory of Dermatology-immunology, The Catholic University of Korea, Seoul 137-701, Republic of Korea. Electronic address: tykimder@catholic.ac.kr.
authors.forename: Tae-Yoon
authors.initials: TY
authors.lastname: Kim
authors.affiliation: Department of Systems Biology, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Republic of Korea. Electronic address: wtkim@yonsei.ac.kr.
authors.forename: Woo Taek
authors.initials: WT
authors.lastname: Kim
mesh_terms: D002478:Cells, Cultured; D003062:Codon; D004926:Escherichia coli; D015870:Gene Expression; D006801:Humans; D007700:Kinetics; D018515:Plant Leaves / Q000201:enzymology / Q000235:genetics; D030821:Plants, Genetically Modified; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics
chemical_list: D003062:Codon; D011994:Recombinant Proteins; C441642:SOD3 protein, human; D013482:Superoxide Dismutase
keywords: Human extracellular superoxide dismutase; Plant-based protein expression; Recombinant proteins; Subcellular localization; Tobacco (Nicotiana benthamiana L.)