Abstract
BackgroundH. pylori is closely related to the occurrence and development of various digestive gastritis, peptic ulcer and mucosa-associated lymphoid tissue (MALT) lymphoma. H. pylori is also a class I carcinogen of gastric cancer. VacA is the only exocrine toxin of H. pylori, which plays a very important role in the pathogenesis of H. pylori. The production of VacA in natural circumstances is complex with heavy workload and low yield. Therefore, it is very important to obtain recombinant VacA protein which is stable and biologically active. This study therefore aims to explore the expression, purification and stable storage of VacA toxin of H. pylori in E.coli, and to provide experimental basis for further exploration of the role of VacA in H. pylori -induced inflammation of cancer.ResultsA 2502-bp fragment and VacA gene were identified. An 89.7-kDa VacA34–854 recombinant protein was expressed and purified from the recombinant engineering bacteria and was preserved stably in 50 mM acetic acid buffer (pH 2.9). The amount of the recombinant protein was larger in the inclusion bodies than in the supernatant. In addition, after a 24-h culture with VacA recombinant protein, GES-1 cells demonstrated evidence of apoptosis including early nuclear immobilization and clustering under inverted microscope and TEM. It was found that VacA recombinant protein induced apoptosis by TUNEL assay.ConclusionsA VacA recombinant protein that is stably and highly expressed and possesses pro-apoptotic activity is successfully constructed. The protein is stably preserved in 50 mM acetic acid buffer (pH 2.9).
Highlights
H. pylori is closely related to the occurrence and development of various digestive gastritis, peptic ulcer and mucosa-associated lymphoid tissue (MALT) lymphoma
One of the most crucial toxins produced by H. pylori is the vacuolating cytotoxin A (VacA), which has been demonstrated to play a vital role in the pathogenesis of peptic ulcer disease and gastric cancer [8, 9]
Considering the pivotal role of vacuolating cytotoxin A protein in H. pylori infection, VacA could be the best candidate for the construction of a recombinant vaccine [14]
Summary
H. pylori is closely related to the occurrence and development of various digestive gastritis, peptic ulcer and mucosa-associated lymphoid tissue (MALT) lymphoma. Helicobacter pylori infection can induce a chronic immune response including persistent oxidative stress in the stomach, further leading to DNA damage that eventually can lead to gastric cancer [6, 7]. One of the most crucial toxins produced by H. pylori is the vacuolating cytotoxin A (VacA), which has been demonstrated to play a vital role in the pathogenesis of peptic ulcer disease and gastric cancer [8, 9]. VacA promotes immune tolerance and persistent H. pylori infection through its activities on T Cells and antigen-presenting cells [13] These are all important virulence factors that H. pylori use to maintain a prolonged pro-inflammatory response while evading selfdestruction. Considering the pivotal role of vacuolating cytotoxin A protein in H. pylori infection, VacA could be the best candidate for the construction of a recombinant vaccine [14]
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have