Purification and Characterization of a Cu,Zn-SOD from Garlic (Allium sativum L.). Antioxidant Effect on Tumoral Cell Lines |
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Authors: | Imen Hadji M Nejib Marzouki Daniela Ferraro Elena Fasano Hafedh Majdoub Giovanni Pani Ferid Limam |
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Institution: | (1) National Institute of Applied Sciences and Technology, BP 676-1080, Tunis Cedex, Tunisia;(2) Institute of General Pathology, Catholic University of Rome, Rome, Italy;(3) Faculty of Sciences of Sfax, Sfax, Tunisia;(4) Center of Biotechnology, Borj-Cédria, BP 901-2050, Hammam-Lif, Tunisia |
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Abstract: | Crude garlic extract contains one Mn-superoxide dismutase designated as SOD1 and two Cu,Zn superoxide dismutases as SOD2 and
SOD3. The major isoform SOD2 was purified to homogeneity by Sephacryl S200-HR gel filtration, DEAE Sepharose ion exchange
chromatography, and chromatofocusing using PBE 94. SOD2 was purified 82-fold with a specific activity of 4,960 U/mg protein.
This enzyme was stable in a broad pH range from 5.0 to 10.0 and at various temperatures from 25 to 60°C. The native molecular
mass of SOD2 estimated by high performance liquid chromatography on TSK gel G2000SW column was 39 kDa. Sodium dodecyl sulfate–polyacrylamide
gel electrophoresis analysis showed a single band near 18 kDa, suggesting that native enzyme was homodimeric. The isoelectric
point as determined by chromatofocusing was 5. Analysis of its N terminal amino acid sequence revealed high sequence homology
with several other cytosolic Cu,Zn-SODs from plants. Exposure of cancer cell lines to garlic Cu,Zn-SOD2 led to a significant
decrease in superoxide content with a concomitant rise in intracellular peroxides, indicating that the enzyme is active in
mammalian cells and could, therefore, be used in pharmacological applications. |
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Keywords: | Cu Zn-SOD Antioxidant activity Garlic Purification Tumoral cell lines Reactive oxygen species |
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