Influence Of Natural Small Molecules On The Abnormal Aggregation And Neurotoxicity Of Neurodegenerative Diseases-related Proteins

Alzheimer’s disease and Parkinson’s disease seriously threaten human health. These diseases involve the disorder folding and aggregation of amyloid-β (Aβ) peptide and a-synuclein (Aβ) protein, respectively. Normally soluble proteins are aggregated into soluble toxic aggregates or deposited as abnormally insoluble amyloid fibrils during the disorder folding and aggregation process that disrupt tissue structure and cause disease. Accordingly, new attempts have been carried out to search for the therapeutic agents which can inhibit the formation of toxic aggregtes and remold the preformed fibrils into not-toxic components instead of toxic fibril precursors. Nonetheless, only limited experimental results of those agents have been reported and the action mechanism of the agents is still ambiguous.Metal ions are known to trigger the structural transformation of Ap peptide and AS protein, and induce the formation of the toxic aggregate species. Therefore, inhibiting of the protein folding modulated by metal ions would be significant.To date, some small molecules have been explored to treat neurodegenerative diseases. Among those compounds, curcumin, a natural product, is considered capable of binding both metal ions and neurodegenerative disease-related proteins. Nevertheless, little is know about the combined action of curcumin and metal ions on Aβ and AS fibrillation and neurotoxicity. In addition, trehalose has attracted special attention. It was found to be effective in inhibiting Ap peptide aggregation. The striking similarity between the fibrillation of AS and Aβ enables us to hypothesize that trehalose might be able to disrupt AS aggregation and potentially contribute to PD therapy.In the present thesis, we attempt to develop approaches treating neurodegenerative diseases by investigations of the potency of curcumin on metal-induced protein fibrillation and neurotoxicity, and the impact of trehalose on the aggregation and neurotoxicity of AS protein. The founding of this thesis is as follows:(1) Influence of curcumin on the Fe(Ⅲ) and AI(Ⅲ)-induced conformation transition of silk fibroinSilk fibroin (SF) which have essential and sufficient amino acid sequences for the protein fibrillation on nucleation-dependent conformation transition mechanism similar to that for the fibrillation of neurodegenerative proteins, was used as a model protein to reveal the influence of curcumin on Fe(Ⅲ) and Al(Ⅳ)-induced conformation transition of proteins. It was demonstrated that Fe(Ⅲ) and Al(Ⅲ) at certain concentration could induce the conformation of SF into β-sheet, while curcumin could inhibit the event. In particular, Al(Ⅲ)-curcumin complexes formed at [Al(Ⅲ)]/[curcumin] ratios from1:1to1:2could reverse the conformation of SF from β-sheet into random coil.(2) Inhibitory effect of curcumin on the AI(Ⅲ)-induced AP42aggregation and neurotoxicity in vitroIt was demonstrated that although Al(Ⅲ) can promote the AP42fibrillation dose-dependently, leading to the high neurotoxicity to PC12cells, curcumin can inhibit the events. Besides, we found that curcumin is able not only to inhibit the formation of Al(Ⅲ)-induced Aβ42fibrillation, but also to form the Al(Ⅲ)-curcumin complexes which in turn can remold the preformed, mature, ordered Aβ42fibrils into the low toxic amorphous aggregates.(3) Trehalose inhibits fibrillation of wild-type and A53T mutant a-synuclein protein and disaggregates existing aggregatesIt was found that when co-incubated with trehalose, wild-type a-synuclein in vitro preferred assembling into the large amorphous aggregates instead of the neurotoxic β-sheet-rich fibrillar aggregates during the prophase incubation. On the other hand, the large amorphous aggregates could be disassembled into the small amorphous particles, even into the random coil structure after long time of incubation. In addition, trehalose at low concentration disaggregates preformed A53T AS protofibrils and fibrils into small aggregates or even random coil structure, while trehalose at high concentration slows down the structural transition into β-sheet structure and completely prevents the formation of mature A53T AS fibrils.(4) Trehalose inhibits wild-type and A53T mutant a-synuclein protein overexpression and neurotoxicity in transduced PC12cellsIt was demonstrated that trehalose at concentration lower than1.0mM and Al(Ⅲ)-curcumin complexes formed at [Al(Ⅲ)]/[curcumin] ratios of1:1inhibited the overexpression of wild-type and A53T mutant a-synuclein in the transduced PC12cells and protected the cells against the neurotoxicity induced by a-synuclein. Besides, although H2O2and aluminum ions increased the expression and neurotoxicity of a-synuclein protein in the cells, Al(Ⅲ)-curcumin complexes formed at [Al(Ⅲ)]/[curcumin] ratios of1:1could inhibit the events.In conclusion, Al(Ⅲ)-curcumin complexes and trehalose are worth potentially developing as a therapy agent against the neurodegenerative disorders in the future.