›› 2011, Vol. 29 ›› Issue (2): 1-81-86.

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Cloning and Expression of Metalloprotease Gene from Schistosoma japoncum and its Immunoprotective Efficiency

 XU  Bin1, JU  Chuan1,3, LEI  Yan2, MO  Xiao-Jin1, FENG  Zheng1, HU  Hua-Nian1, HU  Wei1*   

  1. 1 National Institute of Parasitic Diseases，Chinese Center for Disease Control and Prevention；Key Laboratory of Parasite and Vector Biology，MOH；WHO Collaborating Centre for Malaria，Schistosomiasis and Filariasis，Shanghai 200025，China；2 Biotechnology School，East China University of Science and Technology，Shanghai 200237，China；3 Institute of Entomology，Central China Normal University，Wuhan 430079，China
  • Online:2011-04-30 Published:2012-09-27

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Abstract: Objective   To clone and express a metalloprotease gene of Schistosoma japonicum, purify the expressed protein，and investigate the induced immune response in mice and its localization in the parasite.  Methods  Specific primers were designed according to the EST sequence and used for amplification of the encoding sequence from the S. japonicum cDNA clone containing S. japonicum metalloprotease. The gene was subcloned into pET-28a plasmid and expressed，and the recombinant protein was purified with HisoTag affinity chromatography. Western blotting was used to analyze the immunogenicity. Eighteen C57BC/6 mice were divided into two groups.  Mice in group A were immunized each with 25 μg purified recombinant SjB04 at every 2 weeks for 3 times. Mice in group B received only adjuvant as control. Each mouse was challenged by （40±2） cercariae at the third week after the last immunization. Fecal samples were collected for 6 days from 37th days after challenge. Eggs per gram feces and rate of egg reduction were calculated. S. japonicum adult worms were collected from infected mice，and used for preparing frozen sections and indirect immunofluorescence staining with specific polyclone antibody to S. japonicum metalloprotease.  Results  The metalloprotease gene SjB04 was cloned，sequenced and expressed. The immunofluorescence localization showed that SjB04 protein distributed mainly in the intestinal epithelium of the adult worm. The recombinant protein was specifically recognized by the S. japonicum-infected rabbit sera，showing that the expressed product possessed antigenicity. Mice immunized with the recombinant protein revealed a reduction in number of adult worms，eggs in feces by 27.1% and 57.8%，respectively.   Conclusion   The recombinant protein of S. japonicum metalloprotease has been obtained with Mr 36 500. The protein locates in the intestinal epithelium of adult worm. Immunization with the SjB04 protein induces significant reduction of fecal eggs.

Key words: Schistosoma japoncum, Metalloprotease, Immune protection