Intact long-type DupA protein in Helicobacter pylori is an ATPase involved in multifunctional biological activities.
The function of intact long-type DupA protein in Helicobacter pylori was analyzed using immunoblotting and molecular biology techniques in the study. After cloning, expression and purification, ATPase activity of DupA protein was detected. Antibody was produced for localization and interaction proteins analysis. The dupA-deleted mutant was generated for adhesion and CagA protein translocation assay, susceptibility to different pH, IL-8 secretion assay, cytotoxicity to MKN-45 cells and proteins-involved apoptosis analysis. DupA protein exhibited an ATPase activity (129.5±17.8 U/mgprot) and located in bacterial membrane, while it did not involve the adhesion and CagA protein delivery of H. pylori. DupA protein involved the urease secretion as the interaction proteins. The wild type strain had a stronger growth in low pH than the dupA-deleted mutant (p < 0.001). IL-8 productions from GES-1 cells infected with the wild type strain were significantly higher than from those with the mutant (p < 0.001). The amounts of vital MKN-45 cells were decreased and the numbers of apoptotic cells were increased with the wild type strain, compared to those with the mutant after 12 h (p < 0.05). The increase of cleaved Caspase-3 and Bax was significantly higher and the decrease of Bcl-2 was more obvious in MKN-45 cells exposed to the wild type strain than that exposed to the mutant after 6 h. We demonstrate that intact long-type DupA protein located in membrane as ATPase is a true virulence factor associated with duodenal ulcer development involving the IL-8 induction and urease secretion, while it inhibits gastric cancer cell growth in vitro by activating the mitochondria-mediated apoptotic pathway.
School of Medical Science and Laboratory Medicine, Jiangsu University, Zhenjiang, Jiangsu 212013, PR China; Department of Clinical Lab, Weihai Municipal Hospital Affiliated to Dalian Medical University, Weihai, Shandong 264200, PR China.
Ming-yi
MY
Wang
School of Medical Science and Laboratory Medicine, Jiangsu University, Zhenjiang, Jiangsu 212013, PR China.
Cheng
C
Chen
School of Medical Science and Laboratory Medicine, Jiangsu University, Zhenjiang, Jiangsu 212013, PR China.
Chen
C
Shao
Department of Clinical Lab, Weihai Municipal Hospital Affiliated to Dalian Medical University, Weihai, Shandong 264200, PR China.
Shao-bo
SB
Wang
Department of Clinical Lab, Weihai Municipal Hospital Affiliated to Dalian Medical University, Weihai, Shandong 264200, PR China.
Ai-chu
AC
Wang
Department of Clinical Lab, Weihai Municipal Hospital Affiliated to Dalian Medical University, Weihai, Shandong 264200, PR China.
Ya-chao
YC
Yang
Department of Clinical Lab, Weihai Municipal Hospital Affiliated to Dalian Medical University, Weihai, Shandong 264200, PR China.
Xiao-yan
XY
Yuan
School of Medical Science and Laboratory Medicine, Jiangsu University, Zhenjiang, Jiangsu 212013, PR China. Electronic address: shaoshihe2006@aliyun.com.
Shi-he
SH
Shao
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D004847:Epithelial Cells / Q000276:immunology / Q000382:microbiology / Q000502:physiology; D017353:Gene Deletion; D016480:Helicobacter pylori / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D006863:Hydrogen-Ion Concentration; D016209:Interleukin-8 / Q000378:metabolism; D050296:Microbial Viability / Q000187:drug effects; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
C528428:DupA protein, Helicobacter pylori; D016209:Interleukin-8; D037521:Virulence Factors; D000251:Adenosine Triphosphatases
ATPase; Apoptosis; DupA protein; Helicobacter pylori; Urease