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Helicobacter pylori cagT Protein:Expression,Purification and Immunogenicity

DING Hong-lei1,2,PAN Jing3,ZHANG Wei-jun2,ZHANG Yi2,LUO Ping2,XIE Qing-hua2,MAO Xu-hu2,ZOU Quan-ming21. Institute of Sericulture and System Biology,Southwest University,Chongqing 400715,China;2. Department of Clinical Microbiology and Immunology,Third Military Medical University & National Engineering Technological Research Center of Immune Biologicals,Chongqing 400038,China;3. Team 20 of Student Brigade,Third Military Medical University,Chongqing 400038,China  
[Objective] CagT is one of the key structural proteins in cytotoxin-associated gene pathogenicity island(cag PAI)of Helicobacter pylori.Cag PAI's structure will disappear and its function will be deleted when cagT gene is knocked out.The aim of this research is to analyse the structure cagT peotein and understand its pathogenicity.[Methods] The cagT gene was amplified by PCR with genomic DNA from H.pylori 26695 strain.The amplified product was cloned by T/A method,linked to pET-28a(+)vector,and then transferred into the host bacteria E.coli BL21(DE3).The cagT protein was expressed by IPTG inducing and purified by affinity chromatography.Antibody against cagT peotein was acquired by immunization of rabbits,and the antigenicity was evaluated through cagT antibody.[Results] CagT gene was cloned successfully,and the nucleotide sequence was found to be consistent with the sequence published in GenBank.The recombinant cagT protein was expressed in both soluble form and inclusion body.The highest antibody titer of the immunized rabbit sera was 1∶32.[Conclusion] A recombinant vector pET-28a(+)-cagT that can express cagT peotein efficiently was successfully constructed,and the recombinant protein showed perfect antigenicity.These results may lay a foundation for future study on structure and function of cagT.
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