Abstract
HP0232 is an acid-inducible gene that may code secreted protein involving in flagella motility in Helicobacter pylori. However, there has no further information in their structure and biological function. In order to study the function of HP0232 protein. In the beginning, I had amplified HP0232 DNA fragments in the presence of genomic DNA of H. pylori and its primers by means of PCR. The HP0232 fragments were subcloned into commercially available pQE30 vectors (Qiagen). The HP0232-pQE30 vectors were transformed into E. coli SG13009. Clones transformed with HP0232-pQE30 vector expressed His-tagged HP0232 protein were collected after IPTG induction protocols. Large amount of soluble HP0232 protein from E. coli containing HP0232-pQE30 vectors were purified from affinity column packed with Ni-NTA resin. Native proteins were prepared for structure genomics study and functional assays. HP0232 proteins were used to immunize rabbits to produce anti-HP0232 antibodies. And then, the detection limit of 1:5000 diluted HP0232 antibody was approximately 3ng HP0232 protein. After purifying HP0232 polyclonal antibody, the detection limit of 1:5000 diluted HP0232 antibody was 0.05ng. In order to see the phenomenon of acid-induced of HP0232 in protein level. The HP0232 protein expression from H. pylori in medium at pH 5.5 at different period of time were detected through Western blot analysis. And these results indicate that HP0232 from H. pylori could be induced in medium at pH 5.5 for 4 hr. And at pH 2.5 (contains 10 mM urea as physiologically condition), the HP0232 protein could be detected in extracellular medium by Western blotting analysis. In addition, basing on the motility test, I have found high concentration of purified HP0232 protein (above 2 μg protein per spot) would inhibit the motility of H. pylori. Otherwise, low concentration of HP0232 (below 2 μg protein per spot) would increase the motility of H. pylori. And 0.9 μg HP0232 protein per spot had most significant influence on motility. And heat inactivated HP0232 wouldn’t have any effect on the motility of H. pylori. Basing on the results above, I concluded that HP0232 is an acid-inducible gene that code protein involving in flagella motility in Helicobacter pylori.