Volume 36 Issue 2
Mar.  2014
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XIN Rui-Xiao, YI Fei, ZHANG Rui, XU Tao, XUAN Pu, LIU Rui, JIA Jun-Tao, CHEN Ji-Xiang. EXPRESSION AND IMMUNOPROTECTIVE ANALYSIS OF IRON-COFACTORED SUPEROXIDE DISMUTASE FROM VIBRIO HARVEYI[J]. ACTA HYDROBIOLOGICA SINICA, 2012, 36(2): 212-219. DOI: 10.3724/SP.J.1035.2012.00212
Citation: XIN Rui-Xiao, YI Fei, ZHANG Rui, XU Tao, XUAN Pu, LIU Rui, JIA Jun-Tao, CHEN Ji-Xiang. EXPRESSION AND IMMUNOPROTECTIVE ANALYSIS OF IRON-COFACTORED SUPEROXIDE DISMUTASE FROM VIBRIO HARVEYI[J]. ACTA HYDROBIOLOGICA SINICA, 2012, 36(2): 212-219. DOI: 10.3724/SP.J.1035.2012.00212
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EXPRESSION AND IMMUNOPROTECTIVE ANALYSIS OF IRON-COFACTORED SUPEROXIDE DISMUTASE FROM VIBRIO HARVEYI

  • Department of Marine Biology, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China;
    2. Shandong Entry-Exit Inspection and Quarantine Bureau, Qingdao 266002, China

  • Received Date: January 26, 2011
  • Rev Recd Date: November 28, 2011
  • Published Date: March 24, 2012
    Graphical Abstract
    • Abstract
  • Vibrio harveyi is one of the important bacterial pathogens of marine animals. Superoxide dismutases, which abate and clear the effect of O2- via catalyse dismutation of superoxide into hydrogen peroxide and oxygen, belong to the anti-oxidation defense system and play a central role in protection against oxidative stress. An iron-cofactored superoxide dismutase gene was cloned by PCR amplification from the chromosomal DNA of V. harveyi SF1. The ORF of the sod gene consists of 600 bp, Sequence analysis showed that homologies of the amino acid sequence with those of Fe-SOD ranged from 91% to 99%. The sod gene was subcloned into pET26b (+) for experssion. SDS-PAGE confirmed that the purified protein was 27 kD. The purified SOD belonged to Fe-SOD based on the analysis of sensitivity to hydrogen peroxid, chlorofrom-ethanol and ultraviolet-visible spectroscopy. Pyorgallol autoxidation method was used to determine SOD activity of the purified protein. The purified SOD had the maximal activity at pH 7 and was stable over a range of pH 6—8. The optimal temperature for enzyme activity was 20 . The protein ℃ n was stable under 40℃. Turbots (Scophthalmus maximus) were immunized with 50 ug of the purified SOD. The immunized fish were then challenged with 0.1 mL of V. harveyi (3.9×108 CFU/mL) four weeks later. The relative percentage survivals (RPS) of the immunized group were 80.00%. Specific antibody could be detected in the sera of the immunized fish with Western blot analysis.
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