Rui Juan Du1, Bow Ho. 1. Department of Microbiology, National University of Singapore, 5 Science Drive 2, Singapore 117597.
Abstract
BACKGROUND: Heat Shock Protein (HSP) has been regarded as a pathogenic factor in Helicobacter pylori infection. Heat Shock Protein 20 (HSP20) of H. pylori is identified as Hs1V based on open reading frame predication of genome sequences. It is a homologue of HslV of E. coli, a peptidase involved in protein degradation. METHODS: The HSP20 gene was cloned and inserted into pET16b fused with His-tag. Recombinant HSP20 protein (rHSP20) was expressed and purified by nickel column. Rabbit anti-rHSP20 was purified by Protein A affinity chromatography and used as a probe to localize HSP20 in H. pylori by immuno-gold labeling and Western blotting. rHSP20 was also used as antigen to test for antibody against HSP20 in patients with H. pylori infection by enzyme-linked immunosorbant assay. RESULTS: Immuno-gold labeled transmission electron microscopy shows that HSP20 is located on the cell surface of H. pylori. Western blotting of 2-D gel shows that HSP20 has a pI of approximately 5.5 and a molecular weight of approximately 18 kDa. The ELISA result shows that there is no significant difference in antibody titre against rHSP20 in all sera tested. CONCLUSION: The presence of IgG to rHSP20 may imply an earlier exposure of the patients and normal subjects to H. pylori. However, the mechanism has not been established. HSP20 has been shown to localize on the surface of H. pylori. Surface localization of H. pylori HSP20 may provide the path to a better understanding of the role and function of HSP20 in bacteria-host interaction.
BACKGROUND: Heat Shock Protein (HSP) has been regarded as a pathogenic factor in Helicobacter pylori infection. Heat Shock Protein 20 (HSP20) of H. pylori is identified as Hs1V based on open reading frame predication of genome sequences. It is a homologue of HslV of E. coli, a peptidase involved in protein degradation. METHODS: The HSP20 gene was cloned and inserted into pET16b fused with His-tag. Recombinant HSP20 protein (rHSP20) was expressed and purified by nickel column. Rabbit anti-rHSP20 was purified by Protein A affinity chromatography and used as a probe to localize HSP20 in H. pylori by immuno-gold labeling and Western blotting. rHSP20 was also used as antigen to test for antibody against HSP20 in patients with H. pyloriinfection by enzyme-linked immunosorbant assay. RESULTS: Immuno-gold labeled transmission electron microscopy shows that HSP20 is located on the cell surface of H. pylori. Western blotting of 2-D gel shows that HSP20 has a pI of approximately 5.5 and a molecular weight of approximately 18 kDa. The ELISA result shows that there is no significant difference in antibody titre against rHSP20 in all sera tested. CONCLUSION: The presence of IgG to rHSP20 may imply an earlier exposure of the patients and normal subjects to H. pylori. However, the mechanism has not been established. HSP20 has been shown to localize on the surface of H. pylori. Surface localization of H. pyloriHSP20 may provide the path to a better understanding of the role and function of HSP20 in bacteria-host interaction.
Authors: Alexandra Grote; Denis Voronin; Tao Ding; Alan Twaddle; Thomas R Unnasch; Sara Lustigman; Elodie Ghedin Journal: PLoS Negl Trop Dis Date: 2017-03-30
Authors: Sumith Kumar; Bipul C Karmakar; Deepesh Nagarajan; Asish K Mukhopadhyay; Richard D Morgan; Desirazu N Rao Journal: Nucleic Acids Res Date: 2018-04-20 Impact factor: 16.971