Cloning and Expression of Helicobacter pylori HpaA Gene

• Main Authors: Moein Farshchian, Saman Hoseinkhani, Javad Atoofi, Shahin Najar Peerayeh
• Format: Article
• Language: English
• Published: Royan Institute (ACECR), Tehran 2009-01-01
• Series: Cell Journal
• Subjects: Helicobacter pylori; hpaA; Recombinant Protein; pET28a
• Online Access: http://celljournal.org/library/upload/article/Pirayeh.pdf

Objective: Helicobacter pylori is associated with chronic gastritis, peptic ulcers, gastric adenocarcinomaand gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Antibiotictherapies do not protect from potential re-infection and have a risk for development of drugresistance. Therefore, prophylactic vaccine mediated protection against H. pylori is an attractiveclinical interest. H. pylori adhesin A (HpaA) is a conserved surface lipoprotein and playsimportant roles in the pathogenesis of infection. In this study the recombinant protein (rHpaA)was over-expressed in E.coli.Materials and Methods: The hpaA gene was amplified by PCR. Prokaryote expression vectorpET28a-hpaA was constructed, and used to transform E.coli BL21DE3. The expressionof recombinant protein induced by IPTG was examined by SDS-PAGE. Western blot wereused to determine immunoreactivity of rHpaA by a rabbit polyclonal antibodies against wholecell of H. pylori.Results: The hpaA gene nucleotide sequence in the recombinant plasmid vector of pET-28-a-hpaA was consistent with that of H.pylori hpaA as published in the GenBank. SDS-PAGEdemonstrated that the constructed prokaryotic expression efficiently produced rHpaA at the1.5 mmol/L of IPTG. HpaA fusion protein was able to react with the rabbit polyclonal antibodyagainst whole cells of H. pylori.Conclusion: A prokaryotic expression system pET-28a-hpaA-BL21 with high efficiency of H.pylori hpaA gene was successfully established and the HpaA fusion protein showed satisfactoryimmunoreactivity. These results indicate that production of a specific recombinant proteinis an alternative and potentially more expeditious strategy for development of H. pylori vaccine.