Studies On SjPGAM Of Schistosoma Japonicum

Schistosomiasis caused by schistosome, is a wide spread parasitic zoonosis that causes serious healthy problem to both human and animals. The research and development on Vaccine and drgus are important and hard problem to the prevention and control of the disease in the world. Phosphoglycerate mutas proteins together with their downstream effectors form a set of key glycolytic pathways. The PGAM glycolytic pathway is important in a wide variety of development processes including cell growth, cell differentiation, cell polarity and apoptosis. In this study ,SjPGAM was cloned and expressed in E coli,the enzymatic activities of rSjPGAM was determined. In addition, we construct and express the recombinant plasmid pET32a-BSjPGAM-BSjEnol and evaluate the immuno protective efficacy against the infection of Schistosoma japonicum induced with the recombinant protein in mice. Except this we observe the effects of Anti-schistosomiasis drugs Artemether on activity and gene expression of SjPGAM in order to reveal the potentiol of the epitope vaccine against Schistosoma japonicum and the mechanism of drug target of Artemether.1,Coloning,Expression and enzyme activities determination of PGAM of Schistosoma japonicum The recombinant plasmids pET28a-SjPGAM was constructed and soluble expressed in E. coli under optimized conditions,What‵s more,the recombinant protein with biological activity, The activity of the recombinant SjPGAM was 0.438 U/mg,The Michaelis-Menton constant(Km)of rSjPGAM was 3.78mmol,Kcat was 5.26s-1. The effects of different temperature,PH,divalent cation as well as EDTA on the enzymatic activities of rSjPGAM were evaluated.The optimal pH and temperature for the reaction were 7 and 37-40℃respectively.The enzyme was considerably sensitive to EDTA and could be inactivation by 6mM EDTA.All test divalent cation could enhanced the enzyme activities especially Ni²⁺.2,Evaluation of the immuno protective effecacy of the recombinant antigen BSjPGAM-BSjEnol of Schistosoma japonicum in mice We analysesed SjPGAM,SjEnol with bioinfomatic method which were stage differentially expressed of schistosomula,The peptides of SjPGAM and SjEnol containing the multivalent epitopes with higher binding capacity of human MHCâ…¡but low homology with the host were homology analyzed and screened through bioinfomatic analysis . The corresponding nucleotide sequence of selected epitopes were spliced, the recombinant plasmid pET32a-BSjPGAM-BSjEnol was constructed and expressed in Escherichia coli BL21 cells.Mice vaccination experiment showed that when compared with those of the blank control mice,the worm and egg reduction rate in group BSjPGAM-BSjEnol were 39.7% (P<0.01) and 64.9%(P<0.01) respectively.Induced higher immuno protection against schistosoma japonicum infection compared with those of SjPGAM and SjEonl .This study may have significance for the development of multi-epitope vaccine against schistosomiasis.3,Effection of Artemethe on SjPGAM gene expression and enzyme activities In the presence of Artemethe,enzyme activities were considerably inhibit,the difference is significantly comparing with control group;Rabbit were intramnscular injected with anti-schistosomiasis drug Artemether after infect with Schistosoma japonicum 6,9,13,17,41days later,Then collected the worm,detection the gene expression of SjPGAM using Real-Time quantitative PCR. The results demonstrate that the expression of SjPGAM was significantly inhibited with Artemethe of 7-day,10-day,14-day,18-day schistosomula as well as 42-day adult worm. SjPGAM were inhibited in protein and molecular levels with artemether .Indicated SjPGAM may be a potential target molecules for the anti-schistosomiasis drug Artemether.