A New Mechanism About The Confrontation Between Host And Pathogen:the S-glutathionylation Of Bacteria Virulence Effectors

S-Glutathionylation is the posttranslational modification of protein cysteine residues by the addition of glutathione (GSH, γ-Glu-Cys-Gly). Glutathionylation is regarded as a reversible posttranslational modification which have many physiological functions in the host cells, such us antioxidation, detoxification, regulate the cell signal transduction, cell proliferation and so on.Apart from these common function in the cells, researchers are surprised to find that GSH also has an important role in defense pathogen infection. There are some small moleculars which can replenish the depletion GSH during the infection and inhibit the pathogen infection. However, this mechanism is unknown. Therefore, we carried out its study, and guessed two possible mechanisms:one is the pathogen itself protein glutathione modification, making itself weaken to invade the host; Another possibility is that the host’s proteins Glutathione modification, thereby actively resist pathogen invasion.We found a phenomenon in the prior study:prokaryotic expression and purification of Shigella effector proteins OspF, GST-OspF activity is lower than many other fusion tags. We suspect that may GSH reagent caused some GST-OspF protein glutathione modification when elution in affinity purification process, thus reducing its activity, it is critical Shigella effcetor to invade the host. above the speculation, the pathogen itself glutathione protein modification occurred more convincing.Based on above questions, we start our research work as followed:Firstly, construction the reaction system in vitro to verify OspF which can occur such modifications in vitro, and to optimize the reaction conditions, further identified this modification by mass spectrometry. Then OspF glutathione in vitro after modification, research the influence of its enzyme activity and binding force with its substrate. We also discussed the effect of OspF protein in vivo redox stress environments. After that we research the OspF’S-Glutathionylation in vivo by cell linfection and transfection and pull down to. Finally, we also discussed the other pathogenic’s effector proteins such as SpvC, OspB, OspC, IpaH, YopJ.Taken together, these results indicated that:1) prokaryotic and eukaryotic vector expression OspF can glutathione modified in vitro, and this modification can be effected by redox regulation; 2) the modification of glutathione of OspF protein itself enzyme activity and substrate binding ability will be decreased; 3) Shigella infection in vivo expression of eukaryotic cells can indeed happen OspF glutathione modification, P-Erk will be decrease at first, then recovery in MAPK signaling pathway, and this modification by redox regulation; 4) many effectors of pathogens can also have the similar modification reactions in vitro...