cloning, expression, purification and antigenicity of recombinant ureb332-hpaa fusion protein from helicobacter pylori
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abstract
objective: helicobacter pylori is a widely distributed gram negative bacterium that infects the human stomach and duodenum. some antibiotic regimens are subjected to cure the infection but the cost of drugs, poor patient compliance and emerging of antibiotic-resistant strains are limiting the usefulness of these antibiotic therapies. therefore, interest in developing a h. pylori vaccine is growing up rapidly. the aim of this study was to construct a recombinant vector containing fusion genes encoding a fragment of b subunit from helicobacter pylori (h. pylori) urease (ureb332) and helicobacter pylori adhesion a (hpaa) and expressed it in e. coli bl21, as well as determining its antigenicity as a vaccine candidate of h. pylori. materials and methods: the target genes encoding ureb332 and hpaa amplified from standard h. pylori chromosome by pcr, digested by restricted endonuclease enzyme and inserted into the prokaryotic expression vector pet28a(+) which was digested by corresponding restricted endonuclease enzyme. the target fusion protein was expressed in the bl21 (de3) e.coli. furthermore, ureb332-hpaa antigenicity was studied by western blotting after ni-nta agarose resin purification. results: enzyme digestion analysis, pcr and sequencing showed that the target genes were inserted correctly into the recombinant vector. the fusion protein ureb332-hpaa was recognized by the rabbit anti h. pylori polyclonal antibody and the human sera infected with h. pylori. conclusion: our results in addition to favorable properties of hpaa and ureb antigens, support the application of rureb332-hpaa fusion protein, as a good candidate for the development of h. pylori vaccine.
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Journal title:
modares journal of medical sciences: pathobiologyPublisher: tarbiat modares university
ISSN 1562-9554
volume 13
issue 2 2010
Keywords
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