Study On The Mechanism Of Amyloid Self-assembly

Amyloid self-assembly is a unique type of protein and peptide aggregate featuring with cross-? sheet molecular structure.Amyloid assembly has diverse morphological types including amyloid fibril,amyloid nano-sheet,and amyloid nano-ribbon.Among them,amyloid fibril is the most common type.Amyloid fibril is relevant to several well-known devastating human diseases such as Alzheimer's disease,Parkinson's disease,mad cow disease,and type II diabetes.In the human body,under pathological conditions,proteins can be misfolded to form insoluble fibrous aggregates called amyloid fibrils.Accumulation and depositions of these amyloid fibrils in tissues and organs ultimately lead to disease symptoms.The disease relevance of amyloid fibrillation has sparked great interests among scientists in the past several decades to better understand this intriguing protein aggregation phenomenon.Besides its clinical significance,amyloid self-assebly including amyloid fibril,amyloid nano-sheet,and amyloid nano-ribbon has also increasingly attracted interest among materials scientists in recent years.Amyloid self-assembly can be considered as a novel type of polypeptide-based biomaterials.Numerous applications of amyloid fibrils in biosensing,tissue engineering,drug delivery,and biomimetic catalysis have been discovered.It is of great significance to study the formation mechanism and its modulation factors of amyloid self-assembly process for the treatment of related diseases and for the development of new biomaterials based on amyloid self-assembly.In these aspects,this study is devoted to the following two parts of works:(1)the modulation mechanism of lysozyme amyloid fibrillation process;(2)the formation mechanism of amyloid nano-sheet by a short amyloid peptide.This work investigated the modulation mechanism on the amyloid fibrillation of hen egg white lysozyme(lysyozme)under the influences of two common surfactants,sodium dodecyl sulfate(SDS)and Triton X-100(TX-100),using atomic force microscopy(AFM)and Fourier transform infrared(FTIR)spectroscopy and discovered a new type of modulation effect.Detailed AFM investigations indicated that both SDS and TX-100 were able to induce some significant morphological changes of lysozyme amyloid fibrils.Both SDS and TX-100 could induce a morphological change which was characterized with alternating fibril regions with increased thicknesses along the fibril axis.In addition,TX-100 was also able to induce a morphological change which was characterized with fibril branching.In contrast,the positively charged cetyltrimethylammonium bromide displayed no such effect as comparedwith the negatively charged SDS and the nonionic TX-100.These intriguing modulation effects of SDS and TX-100 on amyloid fibrillation were discovered in this work for the first time and were hypothesized to be due to surfactant-induced surface-catalyzed secondary nucleation owing to the noncovalent interaction between the surfactants and lysozyme.With this mechanism,the surfactant molecule serve as some type of “bridging” molecule to anchor more lysozyme monomers onto the preformed lysozyme amyloid fibril surface to initiate the surface-catalyzed secondary nucleation process.New fibrils will grow using the anchoring points as new nucleation sites.If new fibrils grow along the pre-formed fibril,amyloid fibril with some thickening regions will form;if new fibrils sprout out of the pre-formed amyloid fibril,branched amyloid fibril morphology will form.In addition,to prove that there is intermolecular interaction betwwen the surfactants with amyloid fibril,FTIR study was performed and the results provided the spectroscopic evidencs for the formation of a SDS-amyloid fibril complex and TX-100-amyloid fibril complex through electrostatic or H-bonding interactions.Our work provides new insight into the modulation effffects of surfactants on amyloid fibrillation.In the study of the formation mechanism of amyloid nano-sheets formed by amyloid peptides,the mechanism of the formation of amyloid nanosheet by a 7-amino acid model amyloid short peptide was explored.By using AFM microscope technology to track the growth of the amyloid nano-sheet in real time,it was found that the growth of the amyloid nano-sheet follows the following rules.First,multiple monomeric polypeptide aggregates form a single ?-sheet structure,which is the initial nucleus.After that,the monomer peptide continued to use ?-sheet structures as templates for further longitudinal and lateral extension growth,and finally formed a complete nano-sheet structure.This work provides new insights into the formation mechanism and microstructure of amyloid nano-sheet.This research involves a variety of biophysical techniques,including atomic force microscopy(AFM),Fourier transform infrared(FTIR)spectroscopy,Thioflavin T(ThT)fluorescence spectroscopy,etc.The comprehensive application of these technologies makes us further deepen our understanding of the nucleation and growth mechanism and the micromolecular structure of amyloid self-assembly.We hope that our exploratory research work can provide useful reference for the prevention and treatment of amyloid diseases and the development of new nano-materials based on amyloid self-assembly.