Expression of Human Extracellular Superoxide Dismutase in Chinese Hamster Ovary Cells and Characterization of the Product

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 84; no. 19; pp. 6634 - 6638
• Main Authors: Tibell, Lena, Hjalmarsson, Karin, Edlund, Thomas, Skogman, Gunnar, Engström, Åke, Marklund, Stefan L.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01-10-1987; National Acad Sciences
• Subjects: amino acid composition; Amino acids; Amino Acids - analysis; Animals; Biological and medical sciences; Biotechnology; cDNA; Cell Line; Chemical composition; CHO cells; Cloning, Molecular; Complementary DNA; Cricetinae; DNA; DNA - metabolism; Enzymes; Fundamental and applied biological sciences. Psychology; Gels; Genetic engineering; Genetic technics; Genetic Vectors; Humans; Lectins; man; Methods. Procedures. Technologies; Molecular cloning; Plasmids; Protein Biosynthesis; Simian virus 40 - genetics; superoxide dismutase; Superoxide Dismutase - genetics; Superoxides; Transcription, Genetic; Characterization; Human; Cell culture; Established cell line; Product; Superoxide dismutase; Genetic engineering; Molecular cloning; Gene expression; CHO cell line
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.84.19.6634

A complementary DNA clone from human placenta, encoding human extracellular superoxide dismutase (EC-SOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1), has recently been isolated and characterized. An expression plasmid, based on the EC-SOD complementary DNA, was transfected into Chinese hamster ovary cells (CHO-K1). The transfected cells secreted human EC-SOD to the culture medium. The secreted recombinant (r) EC-SOD was isolated in high yield with a three-step procedure beginning with immobilized monoclonal anti-EC-SOD antibodies. The properties of the rEC-SOD were compared with native (n) EC-SOD isolated from human umbilical cords. The specific activities and aminoterminal amino acid sequences were identical. The amino acid compositions were virtually identical and very similar to the composition deduced from the complementary DNA sequence. Both rEC-SOD and nEC-SOD contained 4 Cu and 4 Zn atoms per molecule, and the presence of Zn in EC-SOD is thus now established. The rEC-SOD produced is type C, since its affinity for heparin-Sepharose was identical to that of nEC-SOD type C. Both enzymes bound to concanavalin A, lentil lectin, and wheat germ lectin and are thus glycoproteins. rEC-SOD and nEC-SOD seem to have the same subunit structure and composition as analyzed by polyacrylamide gel electrophoresis and gel chromatography.