Controllable Self-assembly Of Rationally Designed α-helical Peptides

Self-assembly is a highly active area of research across different scientific disciplines including chemistry,physics,biology,materials science and engineering.Over the past two decades,numerous studies on molecular self-assembly have brought us closer to thoroughly understanding the rationale of self-assembly as well as establishing the relationship between the delicate structures and intriguing functions of self-assembled systems found in nature.It is now known that self-assembly is primarily driven by weak,non-covalent interactions such as hydrogen bonding（H-bonding）,hydrophobic interactions,electrostatic interactions and metal ion coordination,and the resulting supramolecular architectures rely heavily on the combination and cooperation of these multiple non-covalent interactions.In the meantime,self-assembly has been developed into a promising strategy to construct supramolecular materials from simple molecules,with well-defined nanostructures and tailored properties for specific applications.Among a number of self-assembling molecules,peptides are of particular interest,not only because of their biological origin and biodegradability but also because of their specific bioactivity derived from rationally designed sequences.Many ordered assemblies from natural and synthetic peptides have shown potential applications in tissue engineering and regenerative medicine.On the other hand,the structural hallmarks of peptides,such as the string of amide bonds along the main polypeptide chain,pendant side chains,and the asymmetry ofα-carbons（S enantiomers）confer an extreme abundance,great diversity,and even directionality on their non-covalent interactions,compared to conventional organic molecules and synthetic polymers in which the driving force for their self-assembly is straightforward and more easily controlled.In this thesis,a group of 28-residueα-helical peptides including NN（K IAALKAK NAALKAE IAALEAE NAALEA）,NK（K IAALKKK NAALKAE IAALEKE NAALEA）and HH（K IAALKAK HAALKAE IAALEAE HAALEA）were designed and synthesized on a CEM Liberty microwave synthesizer according to Fmoc solid-phase chemistry.The key of peptide design lies in the incorporation of two asparagine（Asn or N）or histidine（His or H）residues at the a positons of the second and fourth heptads,which allow one sequence to pack into homodimers with sticky ends through specific interhelical Asn-Asn or metal complexation interactions,followed by their longitudinal association into ordered nanofibers.This is in contrast to classical self-assembling helical peptide systems consisting of two complementary peptides.Helical peptides NN,NK,and HH that display distinct hierarchical events.The aggregation of NN is a self-assembly process with a remarkable hierarchy:intrachain H-bonding initially promotes the peptide molecules to fold into amphipathicα-helices;their subsequent dimerization and elongation into nanofibers then arise from the close collaboration of interchain hydrophobic interactions,ionic bonds,and buried H-bonds;then,weak hydrophobic interactions from positions b,c,and f promote fiber thickening.At a higher level of hierarchy,the hydrophobic interaction between NN protofibers can cause significant thickening at elevated temperatures.The introduction of two positively charged Lysine（Lys or K）residues at positions 7f and 21f resulting a markable variations in the peptide NK assembly process.Although two charged Lys residues at f positions are unable to cause any significant inhibition to peptide folding and dimerization,they can effectively impede the association of the dimeric coiled coils into long nanofibers by forming the electrostatic repulsion between them,leading to the absence of ordered nanostructures or the formation of much shorter nanofibers.For His-containing helical peptide HH,Cu²⁺coordination can drive peptide HH folding and assembly to form well-ordered nanostructures in a distinctive mode at p H 5.0.Such an interaction also needs to act in concert with other non-covalent interactions,such as intrachain H-bonding in promoting peptide folding and with hydrophobic interactions and ionic pairing in directing coiled-coil dimerization and association into long nanofibers.In the case of HH,local metal ion coordination and electrostatic interactions were likely to play roles in the packing of dimeric coiled-coil nanofibers,in addition to the hydrophobic adhesion.This packing tends to occur along a preferred direction,leading to fibers with a ribbon-like morphology.Due to the binding of Cu²⁺,the HH nanofibers were net positively charged and the electrostatic repulsions between fibers could prevent further thickening with concentration,one reason for the high degree of cross-sectional monodispersity.The oblate ellipsoidal HH aggregates and their high polydispersity are most likely resulting from the complex Cu²⁺coordination at the mildly basic p H condition.In addition to the imidazole nitrogens,terminal amines and main chain amide nitrogens can also bind Cu²⁺at neutral and basic p H due to partial or complete deprotonation,which virtually disfavors the assembly of dimeric coiled coils along the longitudinal direction and even destabilizes their helical structures.In a summary,this thesis demonstrated the exact roles and collaborative interactions of the four main non-covalent interactions,including H-bonding,hydrophobic interactions,electrostatic interactions and metal ion coordination during the hierarchical self-assembling processes of rational designed helical peptides.Different nanostructures,for example,long and short nanofibers,thin and thick fibers,uniform metal ion-entrapped nanofibers and polydisperse globular stacks,can be prepared by harnessing these interactions at different levels of hierarchy.