Study On The Interaction Between Lipopolysaccharide Binding Protein (LBP) And Peroxiredoxin4(Prx4)

Toll-like receptors (TLRs) are highly conserved molecules, which were first discovered in Drosophila, and related to the proteins of regulating dorsal and ventral patterning during embryogenesis. TLRs play central roles in recognizing microbial molecules and activing the innate immune response. The homologous proteins of TLRs were also found in immune cells of mammalian, which were called the TLR family. Toll-like receptor4(TLR4) is the major receptor that recognizes and responds the cndotoxin/lipopolysaccharide (LPS).The TLR4/CD14signal pathway is the most important way in inflammation reaction induced by LPS. Lipopolysaccharide binding protein (LBP) catalyzes the transfer of LPS to a binding site of membrane-bound (m) CD14, which is one kind of cellular LPS-rcceptors.This LPS-LBP-mCD14complex induces the activation of NF-κB through the TLR4-MD2-MyD-88pathway, inducing the activation of the expression of genes coding proinflammatory factors subsequently, which in turn, leads to the release of the inflammation factors, such as TNF-a, and triggers systemic inflammatory response. In addition, LBP transfers LPS to soluble CD14(sCD14), resulting in activation of mCD14-negative cells, such as endothelial and epithelial, inducing a series of biological effects. Peroxiredoxin4(Prx4) plays roles in oxidation resistance through scavenging ROS in the extracellular space. Prx4can act as a regulator to prevent H2O2-induced activation of NF-κB by reducing H2O2in a TRx-dependent mannar in the cytoplasm. Furthermore, Prx4is found to function as a cytokine in the activation of NF-κB.The TLRs signal pathways related proteins as baits, and adult liver cDNA library genetic coding proteins as preys, we found nearly500protein-protein interactions related with human liver by Yeast Two-Hybrid in the preliminary work, and LBP/Prx4is one pair of them.To date, there is no report about this interaction yet. Because the couple of proteins play an important role in many signal pathways and are found in many complexes, we deduced that this interaction is reliable and probably has important phygiological significance. To confirm this interaction, we applied GST pull-down.. co-locolization and co-immunoprecipitation to investigate the interaction, the results of GST pull-down and co-immunoprecipitation indicated the interaction between LBP and Prx4was positive, and the result of co-locolization indicated that LBP and Prx4localize in cytoplasm. In order to gather more information, large amounts of Prx4have been produced in baculovirus-insect cell expression system. In the future, we will study the LBP bond to the reduced Prx4, or the oxidized Prx4in vitro, which will lay the foundation for studing its physiological function.