Exploring The Effect Of Carbon Nanoparticle And Small Molecules On The Aggregation Of Prion Protein

Prion diseases were serious neurodegenerative diseases caused by the misfolding and aggregation of prion protein.Currently,there was still no effective treatment.When the infection occured,the mortality rate of what was 100%.Studies have found that carbon nanomaterials and some polyphenolic compounds could affect the aggregation and folding of amyloid proteins?such as A?protein,human amylin?,but it was still unclear how to affect the misfolding and aggregation of prion proteins.In this paper,we investigated the effects of carbon nanomaterials and anthraquinone compounds on the misfolding and aggregation of prion proteins using molecular dynamics simulation and biological experiment methods at the atomic and molecular level,respectively.The first part of the study tested the effects of graphene and carbon nanotubes on prion protein aggregation.Firstly,we used thioflavin T fluorescence experiments to prove that graphene and carbon nanotubes could strongly inhibit the aggregation of prion protein and the inhibition of graphene and carbon nanotubes on prion protein aggregation exhibited a concentration-dependent manner.When the concentration of graphene and carbon nanotubes was 50?g/ml or higher,the inhibition rate was higher than 50%.When the concentration of graphene and carbon nanotubes were 100?g/ml 200?g/ml,the fluorescence intensity was very low,and the aggregation of prion protein was almost completely inhibited.Comparison of the inhibitory effect between graphene and carbon nanotubes on the aggregation of prion protein showed that the inhibitory effect of graphene was more obvious.To obtain the inhibition mechanism of carbon nanoparticles on prion protein aggregationa at the atomic level,molecular dynamics simulation was performed to study the effect of carbon nanoparticles on the aggregation of Pr P_127-147 tetramer.By calculating the Root-Mean-Square Deviation?RMSD?of the skeletal atoms?C,C?,N,O?and the number of hydrogen bonds along the trajectories indicated that Pr P_127-147 tetramer without carbon nanoparticles had the greater conformattional changes and carbon nanoparticles could reduce the interaction between peptides and peptides.The representative conformations of the trajectories were obtained using cluster analysis.The results showed that the Pr P_127-147 tetramer could form distinct?-sheet structure without carbon nanoparticles.When in the presence of carbon nanotubes and graphene,the Pr P_127-147tetramer mainly presented as irregularly coil structure.It showed that carbon nanoparticles could inhibit the aggregation of prion protein.By analyzing the number of contacts between the Pr P_127-147tetramer and the carbon nanoparticles and the free energy contribution of each residue,it was found that the number of contacts between Pr P_127-147 and graphene was much greater than the number of contacts with the carbon nanotubes.And the binding energies of the residues in the graphene system were relatively low,explaining that the graphene had stronger inhibition ability.In addition,the?-?interactions between aromatic amino acid residues?Y128,F141,and Y145?and carbon nanoparticles were important for the inhibition of aggregation,which were more outstanding in graphene systems.Overall,our research results clarified that carbon nanoparticles could inhibit the aggregation of prion proteins at atomic level and graphene had stronger inhibition effect than carbon nanotubes.The second part of the thesis was to screen small molecule inhibitors of prion protein aggregation from anthraquinones and their derivatives.Firstly,11 compounds were screened to inhibit prion protein fibrosis using a single concentration of 100?M.Then concentration-dependent inhibition of prion protein fibrosis experiments were carried out to screen positive compounds.Concentration gradient of the small molecule compound was set to 5?M,10?M,20?M,40?M and 80?M.Among them,the five compounds?chrysophanol,physcion,catechin,AE-562/12222209,AE-848/01284025?were obtained to strongly inhibit prion protein aggregation in a concentration dependent manner.Then,the five small molecules were further tested to obtain their binding affinity to prion protein using microscale thermophoresis.Finally,compounds,chrysophanol,AE-848/01284025 and physcion were obtained.Among the three compounds,Chrysophanol had the highest binding affinity with a binding constant of 13.3±3.5?M.For the other two compounds,the binding affinity were much weaker.The three compounds were then subjected to a depolymerization experiment of prion protein,and finally,chrysophanol was demonstrated to have a certain depolymerization effect on prion protein fibers.The results obtained in this paper could provide a theoretical basis for understanding the mechanisms of carbon nanomaterials affecting the aggregation of prion protein.The small molecule inhibitors of anthraquinones could provide a material basis for the discovery of anti-prion compounds.