Macrocyclic Peptides Self-Assemble into Robust Vesicles with Molecular Recognition Capabilities

In this study, we developed macrocyclic peptide building blocks that formed self-assembled peptide vesicles with molecular recognition capabilities. Macrocyclic peptides were significantly different from conventional amphiphiles, in that they could self-assemble into vesicles at very high hydrophilic-to-total mass ratios. The flexibility of the hydrophobic self-assembly segment was critical for vesicle formation. The unique features of this peptide vesicle system include a homogeneous size distribution, unusually small size, and robust structural and thermal stability. The peptide vesicles successfully entrapped a hydrophilic model drug, released the payload very slowly, and were internalized by cells in a highly efficient manner. Moreover, the peptide vesicles exhibited molecular recognition capabilities, in that they selectively bound to target RNA through surface-displayed peptides. This study demonstrates that self-assembled peptide vesicles can be used as strong intracellular delivery vehicles that recognize specific biomacromolecular targets

Helix Stabilized, Thermostable, and Protease-Resistant Self-Assembled Peptide Nanostructures as Potential Inhibitors of Protein–Protein Interactions

Self-assembled peptide nanostructures with actively folded secondary structures have potential to mimic the function of proteins. We here show that α-helix-stabilized self-assembled peptide nanostructures (αSSPNs), whose sizes are comparable to those of proteins, have potential to be developed as protein–protein interaction (PPI) inhibitors along with several unprecedented properties. Using p53-MDM2 PPI as a model system, the molecular recognition and modulation of PPIs by αSSPN grafted with a p53 α-helix (p53 αSSPN) were investigated. The competition assay showed that the p53 αSSPN can inhibit the p53-MDM2 interaction. Interestingly, the p53 αSSPN was far more resistant to degradation by the protease chymotrypsin than the monomeric p53 peptide and had high thermal stability. These results suggest that the αSSPN scaffold holds great potential to be developed as a novel class of PPI inhibitors

Tuning Oligovalent Biomacromolecular Interfaces Using Double-Layered α‑Helical Coiled-Coil Nanoassemblies from Lariat-Type Building Blocks

The target affinity and selectivity of many biomacromolecules depend on the three-dimensional (3D) distribution of multiple ligands on their surfaces. Here, we devised a self-assembly strategy to control the target-tailored 3D distribution of multiple α-helical ligands on a coiled-coil core scaffold using novel lariat-type supramolecular building blocks. Depending on the coiled-coil composition and ligand grafting sites in the lariat building blocks, the structural and functional features of the self-assembled peptide nanostructures (SPNs) could be variably fine-tuned. Using oligovalent protein–RNA (Rev-RRE) interactions as a model system, we demonstrate that longer grafting reinforces the helicity of the peptide ligands, whereas shorter grafting strengthens the target binding affinity of the SPNs in both monovalent and oligovalent interactions. This supramolecular approach should be useful in developing precisely controllable multivalent ligands for biomacromolecular interactions

Chameleon-like Self-Assembling Peptides for Adaptable Biorecognition Nanohybrids

We present here the development of adaptable hybrid materials in which self-assembling peptides can sense the diameter/curvature of carbon nanotubes and then adjust their overall structures from disordered states to α-helices, and <i>vice versa</i>. The peptides within the hybrid materials show exceptionally high thermal-induced conformational stability and molecular recognition capability for target RNA. This study shows that the context-dependent protein-folding effects can be realized in artificial nanosystems and provides a proof of principle that nanohybrid materials decorated with structured and adjustable peptide units can be fabricated using our strategy, from which smart and responsive organic/inorganic hybrid materials capable of sensing and controlling diverse biological molecular recognition events can be developed

Multiplexing Natural Orientation: Oppositely Directed Self-Assembling Peptides

We explore here the possibility that polypeptide chains with directional multiplicity might provide for the control of peptide self-assembly processes. We tested this new possibility using an oppositely directed peptide (ODP) supramolecular system. The ODP could make it possible to form a βαβ motif with antiparallel β-sheets, which does not exist in nature. Furthermore, the designed ODPs were able to self-assemble into discrete, homogeneous, and structured protein-like controlled nano-objects. ODPs represent a simple but powerful unnatural self-assembling peptide system that can become a basic scaffold for fabricating more complex and elaborate artificial nanostructures

Macromolecular Sensing of RNAs by Exploiting Conformational Changes in Supramolecular Nanostructures

Here, we report on a ratiometric fluorescence biosensor based on self-assembled peptide nanostructures (SPN), which can respond to conformational changes induced by RNA ligand binding. The design of the SPN biosensor was inspired by the conformational stabilization and multimerization behaviors of the HIV-1 Rev protein induced by cooperative protein–protein and protein–RNA interactions. Because conformation-sensitive SPN biosensors can orchestrate binding and signal transduction events, they can be developed as highly sophisticated and smart nanomaterials for biosensing

Nanomorphological Diversity of Self-Assembled Cyclopeptisomes Investigated via Thermodynamic and Kinetic Controls

The physicochemical and biological characteristics of vesicles are dependent on the type of self-assembly building blocks and methods of preparation. In this report, we designed a vesicle-forming linear and cyclic peptide building blocks and investigated the effect of molecular topology and thermodynamic and kinetic controls on the stability and morphological features of the self-assembled vesicles. Comparison of topological effect on self-assembly revealed that the strong association of the aromatic hydrophobic segments is observed only in the cyclic peptide, which is most likely the results of constrained structure along with the restriction in the molecular degree of freedom. Consequently, the formation of stable vesicles could be observed only with the cyclic peptide. Further investigation with cyclic peptide building blocks revealed that depending on the control methods, vesicles with a variety of structural features, such as polygonal, wrinkled, round, round-patched, and round-fused vesicles, could be fabricated. Our results demonstrate that existing vesicle structures constitute only a fraction of the possible structural diversity and that macrocyclic peptides can provide a wealth of opportunities in vesicle engineering

Inhibition of Multimolecular RNA–Protein Interactions Using Multitarget-Directed Nanohybrid System

Multitarget-directed ligands (MTDLs) are hybrid ligands obtained by covalently linking active pharmacophores that can act on different targets. We envision that the concept of MTDLs can also be applied to supramolecular bioinorganic nanohybrid systems. Here, we report the inhibition of multimolecular RNA–protein complexes using multitarget-directed peptide–carbon nanotube hybrids (SPCHs). One of the most well-characterized and important RNA–protein interactions, a Rev-response element (RRE) RNA:Rev protein:Crm1 protein interaction system in human immunodeficiency virus type-1, was used as a model of multimolecular RNA–protein interactions. Although all previous studies have targeted only one of the interaction interfaces, that is, either the RRE:Rev interface or the RRE–Rev complex:Crm1 interface, we here have developed multitarget-directed SPCHs that could target both interfaces because the supramolecular nanosystem could be best suited for inhibiting multimolecular RNA–protein complexes that are characterized by large and complex molecular interfaces. The results showed that the single target-directed SPCHs were inhibitory to the single interface comprised only of RNA and protein in vitro, whereas multitarget-directed SPCHs were inhibitory to the multimolecular RNA–protein interfaces both in vitro and in cellulo. The MTDL nanohybrids represent a novel nanotherapeutic system that could be used to treat complex disease targets

pH-Dependent In-Cell Self-Assembly of Peptide Inhibitors Increases the Anti-Prion Activity While Decreasing the Cytotoxicity

The first step in the conventional approach to self-assembled biomaterials is to develop well-defined nanostructures in vitro, which is followed by disruption of the preformed nanostructures at the inside of the cell to achieve bioactivity. Here, we propose an inverse strategy to develop in-cell gain-of-function self-assembled nanostructures. In this approach, the supramolecular building blocks exist in a unimolecular/unordered state in vitro or at the outside of the cell and assemble into well-defined nanostructures after cell internalization. We used block copolypeptides of an oligoarginine and a self-assembling peptide as building blocks and investigated correlations among the nanostructural state, antiprion bioactivity, and cytotoxicity. The optimal bioactivity (i.e., the highest antiprion activity and lowest cytotoxicity) was obtained when the building blocks existed in a unimolecular/unordered state in vitro and during the cell internalization process, exerting minimal cytotoxic damage to cell membranes, and were subsequently converted into high-charge-density vesicles in the low pH endosome/lysosomes in vivo, thus, resulting in the significantly enhanced antiprion activity. In particular, the in-cell self-assembly concept presents a feasible approach to developing therapeutics against protein misfolding diseases. In general, the in-cell self-assembly provides a novel inverse methodology to supramolecular bionanomaterials

Differential Self-Assembly Behaviors of Cyclic and Linear Peptides

Here we ask the fundamental questions about the effect of peptide topology on self-assembly. The study revealed that the self-assembling behaviors of cyclic and linear peptides are significantly different in several respects, in addition to sharing several similarities. Their clear differences included the morphological dissimilarities of the self-assembled nanostructures and their thermal stability. The similarities include their analogous critical aggregation concentration values and cytotoxicity profiles, which are in fact closely related. We believe that understanding topology-dependent self-assembly behavior of peptides is important for developing tailor-made self-assembled peptide nanostructures