Mechanism Of Lysozyme Amyloid Fibrillation And The Inhibition And Modulation Effects Of Dye Molecules On Related Processes

Amyloid fibrillation is a unique aggregation behavior of proteins and peptides.Under proper conditions,proteins and peptides form fibrillar aggregates with cross-?supramolecular configuration through self-assembly process.Amyloid fibrillation is closely related to many major diseases affecting human health,including Alzheimer's disease and Parkinson's disease.In addition,as a new type of nano materials,amyloid fibril also shows many potential applications in the field of materials sciences.Therefore,the research on amyloid fibrillation has been a hot topic at home and abroad in recent years.This thesis work focuses on two aspects: the formation mechanism investigation of amyloid fibrillation,and the inhibition and modulation of amyloid fibrillation.In the study of mechanism of amyloid fibrillation,we will study the amyloidogenic system based on lysozyme.Lysozyme(hen egg white lysozyme)is a widely used model system in the study of amyloid fibrillation.People have carried out various studies using this model system,but there is still a lack of research on the nucleation and growth mechanism of lysozyme amyloid fibrillation.We used atomic force microscopy(AFM)to track the process of lysozyme amyloid fibrillation under different p H conditions and different concentrations.Through in-depth analysis of AFM images at various time-points,we proposed the mechanism of lysozyme amyloid fibrillation,and discussed the difference between this mechanism and the existing mechanism reported in the literature.We propose that in the early stage of lysozyme amyloid fibrillation,a multi-sheet structure is formed as the protofibril structure,while in the existing mechanism,it is believed that a single-sheet structure will appear as the protofibril structure in the early stage of amyloid fibrillation;then,the protofibril will further thicken and branch under the mechanism of secondary nucleation,and the protofibril will grow through primary nucleation.The existence of the secondary nucleation mechanism leads to the rich morphology of the lysozyme amyloid system;while in the existing mechanism,the thickening and and morphology of amyloid fibril is believed to be realized through the intertwining of multiple protofibrils.In the study of the inhibition and modulation of amyloid fibrillation,three dye molecules,brilliant blue G,acid chrome blue K and eriochrome black T,were found to be able to inhibit and modulate lysozyme amyloid fibrillation.Using AFM and and Fourier transform infrared spectroscopy,we found that the three dyes have dual effects on lysozyme amyloid fibrillation,that is,the dyes have both inhibitory and modulation effects,and such effects show a concentration dependent behavior.In other words,at high dye concentration,the three dyes can completely inhibit the occurrence of amyloid fibrillation;while at low dye concentration,the three dyes can affect the morphology of amyloid fibrils.Compared with the control group,under the modulation effect of the three dyes,the amyloid systems show fibril thickening,fibril branching,and fibril intertwining to a greater extent.We speculate that these morphological effects of dyes are mainly due to the secondary nucleation induced by the interaction between dyes and amyloid fibrils.These observed inhibitory and modulation effects were verified at different temperatures.In addition,we also studied the structure of amyloid fibrils under the influence of dyes by infrared spectroscopy peak fitting technology and chemometrics analysis method,and found that the secondary structures of amyloid fibrils are different among these amyloid systems under the different effects of different dyes.This thesis include four chapters.The first chapter is the introduction;the second chapter is about the formation mechanism of lysozyme amyloid fibrillation;the third chapter is about the effects of dye molecules on lysozyme amyloid fibrillation;the forth chapter is the conclusion and perspectives.