Preparation And Application Of Chiral Amino Acid-based Supramolecular Assemblies

As one of the core contents of supramolecular chemistry,self-assembly has attracted extensive attention from researchers.Molecular self-assembly refers to the process in which basic structural units spontaneously form stable and ordered structures through non-covalent bond interactions,and is considered to be an important means for creating new substances and generating new functions above the molecular level.Meanwhile,self-assembly is a very important bottom-up strategy for constructing supramolecular materials.Many biomolecules have been demonstrated to self-assemble into well-ordered supramolecular structures and play vital roles in biological systems.For example,phospholipids form biological membranes through hydrophilic and hydrophobic interactions,deoxyribonucleic acid(DNA)strands form double helix structures through hydrogen bonds and so on.Chirality is ubiquitous in nature and life,and the research related to supramolecular chirality has aroused immense interest.In the study of supramolecular chirality,the fundamental point is the choice of building blocks.Amino acids,as the basic units of peptides and proteins closely related to various physiological and pathological processes,are an important class of chiral small molecules.They not only have unique chiral structure,multiple reaction sites,and aboundant resources,but also have the good biocompatibility and biological activity.Importantly,the molecular chirality of amino acids can be transferred and amplified with the orderly stacking of molecules in the assembly process.Therefore,amino acids have become regarded as one of the ideal building blocks in fabricating supramolecular chiral nanostructures.With the rapid development of supramolecular chemistry,great progress has been made in the study of assembly mechanism.However,how to accurately regulate the structure of the assembly to achieve the ideal function,and then realize its potential application in the fields of chemistry,biology,medicine,etc.,is a huge challenge.In this study,we designed and developed chiral building blocks based on aspartic acid(Asp)and histidine(His),which spontaneously formed stable and ordered chiral phospholipid vesicles and chiral supramolecular gels through non-covalent bond interactions,such as hydrophobic interactions,π-π stacking,hydrogen bonds.We investigated the interactions and internal molecular mechanisims between them and biomolecules related to diseases(e.g.Alzheimer’s disease(AD),cancer),as well as the regulation of related pathological processes in living organisms,which provided more effective information for the realization of early diagnosis and precise treatment of diseases.The detailed experimental studies are as follows:In Chapter 2,a pair of chiral phospholipid molecules(abbreviated to L-/D-Asp-DPPE)based on chiral aspartic acid(L-/D-Asp)and1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine(DPPE)were designed and prepared to regulate conformational transition and fibrillation of β-amyloid(Aβ(1-40))associated with AD.First,L-or D-Asp-DPPE self-assembled into monolayer phospholipid vesicles with a uniform particle size of 100 nm(abbreviated to L-or D-Asp-DPPE vesicles)through thin-film rotary evaporation and extrusion method.Then,the self-assembled L-or D-Asp-DPPE vesicles were incubated with Aβ(1-40)monomers,and the influence of chiral phospholipid vesicles on the fibrillation process of Aβ(1-40)was investigated by aggregation kinetics experiment.Meanwhile,the aggregation morphology of Aβ(1-40)in the presence of L-/D-Asp-DPPE vesicles were observed by atomic force microscopy.These results indicated that both L-and D-Asp-DPPE vesicles inhibited the fibrillation of Aβ(1-40),especially D-Asp-DPPE vesicles.Further,mechanism studies and molecular dynamics simulations analyzed the interaction sites and binding modes of chiral phospholipid vesicles with Aβ(1-40).These results demonstrated that the intensive electrostatic binding between D-Asp on the head of D-Asp-DPPE and the residue lysine(K16)in the peptide remarkably promoted the adsorption of Aβ(1-40)monomers on the D-Asp-DPPE surface,prevented the random coil to β-sheet transition,and finally inhibited the amyloid fibrillation.By contrast,the binding affinity of L-Asp-DPPE surface with Aβ(1-40)monomers was substantially weaker,the monomers accumulated near the surface,and the conformational transition and fibrillation processes were not influenced strongly.In Chapter 3,L-and D-Asp-DPPE vesicles were applied to an AD cell model(mouse neuroblastoma(N2a)cells)to investigate the effect on the behavior of cells.First,cell viability assay was performed to analyze the cytotoxicity of L-/D-Asp-DPPE vesicles and to demonstrate their ability to rescue Aβ(1-40)aggregation-induced cytotoxicity.Then,apoptosis,mitochondrial membrane potential and oxidative stress experiments confirmed the protective effect of L-/D-Asp-DPPE vesicles on cells.Cell imaging experiments showed that the intracellular binding of L-/D-Asp-DPPE vesicles to Aβ(1-40).Further,three-month-old APPswe/PS1d9(APP/PS1)double-transgenic mice were treated with L-and D-Asp-DPPE vesicles through intranasal administration for three months.First,the Morris water maze test was conducted to investigate the improvement of L-/D-Asp-DPPE vesicles on cognitive deficits of APP/PS1 double-transgenic mice.Then,Aβ and glial fibrillary acidic protein(GFAP)immunohistochemical staining experiments confirmed the reduction of Aβ plaques and glial activation in the brain of mice treated with L-/D-Asp-DPPE vesicles.Hematoxylin-eosin(H&E)staining assay evaluated the biosafety of L-/D-Asp-DPPE vesicles on major organs in mice.These results showed that L-/D-Asp-DPPE vesicles could not only protect cells from cell damage caused by Aβ(1-40)aggregation,but also had a good therapeutic effect on AD model mice.In Chapter 4,a chiral gelator(abbreviated to L-Py His)based on chiral histidine(L-His)and pyrenecarboxylic acid was prepared,which self-assembled into supramolecular gels for specific dynamic response of sialic acid and sialylated glycans.First,at the single molecular level,the interactions between L-Py His small molecule and various saccharides were investigated by fluorescence titration experiments.Then,at the supramolecular level,the effects of adding various saccharides solutions on the assembly of chiral supramolecular gel were monitored by fluorescence spectrophotometer.Sialic acid and sialylated glycan could be specifically distinguished according to the changes of fluorescence signal.Meanwhile,the assembled morphologies of chiral supramolecular gels in the presence of various saccharides solutions were observed by helium ion microscope and atomic force microscope.Further,the detailed interaction sites of L-Py His and sialic acid were analyzed by nuclear magnetic resonance titration and quantum chemical calculation,indicating that the sialic acid or sialylated glycan could interact closely with two L-Py His molecules stacked together in the assemblies via hydrogen bonding interactions,thereby preventing the ordered accumulation of the gelators and realizing the specific recognition of sialic acid and sialylated glycan.