To Explore The Effect Of Glutathionylation On FABP5 Binding And Transporting Fatty Acids Using Molecular Dynamics Simulations

Hyperoxia is a clinically important treatment that can effectively relieve cellular and tissue hypoxia.However,prolonged exposure to hyperoxia can cause oxidative stress in the lungs resulting in hyperoxic acute lung injury(HALI),which can seriously endanger the life and health of patients.Due to the complexity of the pathogenesis of hyperoxic acute lung injury,there is still a lack of effective drugs to treat this disease.Recently,it has been shown that fatty acid-binding protein 5(FABP5)reacts with glutathione under oxidative stress conditions to form glutathionylated FABP5 protein(FABP5-SSG)with the help of reactive oxygen species.This glutathione modification enhances the binding and transport of FABP5 to fatty acids such as linoleic acid(LA),resulting in an increased concentration of fatty acids in the nucleus,which enhances the downstream PPARβ/δ signalling pathway and reduces the levels of oxidative stressinduced inflammatory factors,ultimately leading to the alleviation of HALI.However,there is still a lack of mechanistic studies on how glutathionylation modifications enhance the binding and transport of fatty acids by FABP5.To investigate the effect of glutathione modification on the ability of FABP5 to bind and transport fatty acids,molecular dynamics simulations of 500 ns were performed for FABP5,FABP5+LA,FABP5-SSG and FABP5-SSG+LA,respectively.Adaptive steered molecular dynamics simulation were carried out for the FABP5+LA and FABP5-SSG+LA systems.The analysis of the simulation results revealed that.1.the results of stability analysis showed that glutathionylation of FABP5 resulted in a more stable conformation of the protein,and the results of RMSF analysis,secondary structure analysis and residues M35 and L60 distance analysis indicated that the glutathionylation modification had less effect on the α2 helix as well as the β3-β4loop interaction in FABP5,therefore,the glutathionylation modification may not affect the α1 helix and α2 helix by Therefore,glutathione modification may not alter the fatty acid transport capacity of FABP5 by affecting the nuclear localization signal in the α1and α2 helices.2.Combining the results of free energy analysis,hydrogen bonding analysis and salt bridge analysis showed that the glutathione-modified FABP5 protein had stronger interaction and binding ability with LA compared with the unmodified FABP5 protein.The glutathione modified residue C127 could form new hydrogen bonds as well as saltbridge interactions with LA,which directly enhanced the interaction between LA and protein.Meanwhile,the glutathione modified residue C127 could attract the carboxyl end of LA to form stronger hydrogen bonds and salt-bridge interactions at residues K61 and R129 as well as C127,which stabilized the LA conformation and formed stable interactions,which would facilitate the binding of LA to protein.3.The results of the adaptive stretching molecular dynamics simulations showed that more energy was required for LA dissociation from FABP5-SSG.The hydrogen bonding analysis in the dissociation process showed that LA formed less hydrogen bonds with FABP5 and more hydrogen bonds with FABP5-SSG during the dissociation process,and LA formed stronger interactions with surrounding residues during the dissociation process from FABP5-SSG,which made it more difficult to be separated from the protein.reason.Our data and analytical results deepen the understanding of the structure-function relationship of glutathione-modified FABP5 in HALI development and provide a solid theoretical basis for the development of HALI drugs targeting FABP5-related pathways.