Short Amphiphilic Peptides Research Articles

3D Cryo-Electron Microscopy Reveals the Structure of a 3-Fluorenylmethyloxycarbonyl Zipper Motif Ensuring the Self-Assembly of Tripeptide Nanofibers.

Short peptide-based supramolecular hydrogels appeared as highly interesting materials for applications in many fields. The optimization of their properties relies mainly on the design of a suitable hydrogelator through an empirical trial-and-error strategy based on the synthesis of various types of peptides. This approach is in part due to the lack of prior structural knowledge of the molecular architecture of the various families of nanofibers. The 3D structure of the nanofibers determines their ability to interact with entities present in their surrounding environment. Thus, it is important to resolve the internal structural organization of the material. Herein, using Fmoc-FFY tripeptide as a model amphiphilic hydrogelator and cryo-EM reconstruction approach, we succeeded to obtain a 3.8 Å resolution 3D structure of a self-assembled nanofiber with a diameter of approximately 4.1 nm and with apparently "infinite" length. The elucidation of the spatial organization of such nano-objects addresses fundamental questions about the way short amphiphilic N-Fmoc peptides lacking secondary structure can self-assemble and ensure the cohesion of such a lengthy nanostructure. This nanofiber is organized into a triple-stranded helix with an asymmetric unit composed of two Fmoc-FFY peptides per strand. The three identical amphiphilic strands are maintained together by strong lateral interactions coming from a 3-Fmoc zipper motif. This hydrophobic core of the nanofiber is surrounded by 12 phenyl groups from phenylalanine residues, nonplanar with the six Fmoc groups. Polar tyrosine residues at the C-term position constitute the hydrophilic shell and are exposed all around the external part of the assembly. This fiber has a highly hydrophobic central core with an internal diameter of only 2.4 Å. Molecular dynamics simulations highlight van der Waals and hydrogen bonds between peptides placed on top of each other. We demonstrate that the self-assembly of Fmoc-FFY, whether induced by annealing or by the action of a phosphatase on the phosphorylated precursor Fmoc-FFpY, results in two nanostructures with minor differences that we are unable to distinguish.

Self-assembling short peptide amphiphiles as versatile delivery agents: a new frontier in antibacterial research.

Self-assembling short peptide amphiphiles, crafted through a minimalistic approach, spontaneously generate well-ordered nanostructures, facilitating the creation of precise nanostructured biomaterials for diverse biomedical applications. The seamless integration of bioactive metal ions and nanoparticles endows them with the potential to serve as pioneering materials in combating bacterial infections. Nanomanipulation of these molecules' binary structures enables effective penetration of membranes, forming structured nanoarchitectures with antibacterial properties. Through a comprehensive exploration, we attempt to reveal the innovative potential of short peptide amphiphiles, particularly in conjugation with metal cations and nanoparticles, offering insights for future research trajectories.

Silver-Nanoparticle-Embedded Short Amphiphilic Peptide Nanostructures and Their Plausible Application to Reduce Bacterial Infections.

The microbiota-gut-brain axis (GBA) plays a critical role in the development of neurodegenerative diseases. Dysbiosis of the intestinal microbiome causes a significant alteration in the gut microbiota of Alzheimer's disease (AD) patients, followed by neuroinflammatory processes. Thus, AD beginning in the gut is closely related to an imbalance in gut microbiota, and hence a multidomain approach to reduce this imbalance by exerting positive effects on the gut microbiota is needed. In one example, a tyrosine-based short peptide amphiphile (sPA) was used to synthesize antibacterial AgNPs-sPA nanostructures. Such nanostructures showed high biocompatibility and low cytotoxicity, and therefore work as model drug delivery agents for addressing local bacterial infections. These may have therapeutic value for the treatment of microbiota-triggered progression of neurodegenerative diseases.

Design of short peptides and peptide amphiphiles as collagen mimics and an investigation of their interactions with collagen using molecular dynamics simulations and docking studies.

Short peptide sequences and bolaamphiphiles derived from natural proteins are gaining importance due to their ability to form unique nanoscale architectures for a variety of biological applications. In this work, we have designed six short peptides (triplet or monomeric forms) and two peptide bolaamphiphiles that either incorporate the bioactive collagen motif (Gly-X-Y) or sequences where Gly, Pro, or hydroxyproline (Hyp) are replaced by Ala or His. For the bolaamphiphiles, a malate moiety was used as the aliphatic linker for connecting His with Hyp to create collagen mimics. Stability of the assemblies was assessed through molecular dynamics simulations and results indicated that (Pro-Ala-His)3 and (Ala-His-Hyp)3 formed the most stable structures, while the amphiphiles and the monomers showed some disintegration over the course of the 200ns simulation, though most regained structural integrity and formed fibrillar structures, and micelles by the end of the simulation, likely due to the formation of more thermodynamically stable conformations. Multiple replica simulations (REMD) were also conducted where the sequences were simulated at different temperatures. Our results showed excellent convergence in most cases compared to constant temperature molecular dynamics simulation. Furthermore, molecular docking and MD simulations of the sequences bound to collagen triple helix structure revealed that several of the sequences had a high binding affinity and formed stable complexes, particularly (Pro-Ala-His)3 and (Ala-His-Hyp)3. Thus, we have designed new hybrid-peptide-based sequences which may be developed for potential applications as biomaterials for tissue engineering or drug delivery.

Self-assembly of wide peptide nanoribbons via the formation of nonpolar zippers between β-sheets

Because zipper motifs between complementary sidechains can allow molecular order to be maintained over larger length scales, they hold great promise for the preparation of hierarchical peptide self-assemblies with long-range ordering. Although polar zippers based on sidechain-sidechain hydrogen bonding have been demonstrated to promote significant lamination of β-sheets into wide and flat nanoribbons, it remains a challenge to achieve such well-defined nanostructures through nonpolar zippers. In this study, we designed a series of short amphiphilic peptides Ac-I3XGK-NH2, where X at the intramolecular hydrophobic/hydrophilic interface denotes different hydrophobic residues. Microscopic imaging and neutron scattering measurements indicated that only Ac-I3VGK-NH2 could self-assemble into wide and uniform nanoribbons. Spectroscopic and solid-state NMR analyses revealed that the peptide adopted an anti-parallel β-sheet secondary structure with a two-residue shifting. Under such a molecular conformation and alignment, homogenous and extensive nonpolar zippers between β-sheets, consisting of alternating Val4-Ile3 and Ile2-Ile1 sidechain contacts, would be formed, leading to significant β-sheet lamination that can cause wide and flat nanoribbon assembly. Finally, the failure of the peptide variants to form wide and flat nanoribbons indicated a stringent requirement of extensive nonpolar zipper formation between β-sheets for primary sequence, especially the interaction mode of hydrophobic sidechains.

Peptide Self-Assemblies from Unusual α-Sheet Conformations Based on Alternation of d/l Amino Acids.

Peptide self-assembly is a hierarchical process during which secondary structures formed in the initial stages play a critical role in determining the subsequent assembling processes and final structural ordering. Unusual secondary structures hold promise as a source to develop novel supramolecular architectures with unique properties. In this work, we report the design of a new peptide self-assembly strategy based on unusual α-sheet secondary structures. In light of the strong propensity of leucine toward forming helical conformations and its high hydrophobicity, we design two short amphiphilic peptides Ac-LDLLDLK-NH2 and Ac-DLLDLLDK-NH2 with alternating l- and d-form amino acids. Microscopic imaging, neutron scattering, and spectroscopic measurements indicate that the two heterochiral peptides form highly ordered wide nanotubes and helical ribbons with monolayer thickness, in sharp contrast to twisted nanofibrils formed by the homochiral peptide Ac-LLLLK-NH2. Molecular dynamics simulations from monomers to trimers reveal that the two heteropeptides fold into α-sheets instead of β-sheets, which readily pack into tubular architectures in oligomer simulations. Simulated circular dichroism spectra based on α-sheet oligomers validate the proposed α-sheet secondary structures. These results form an important basis for the rational design of higher-order peptide assemblies with novel properties based on unusual α-sheet secondary structures.

Antimicrobial Activity of Silver Nanoparticles Loaded Biomimetic Isomeric Short Lipopeptide Nanostructures

AbstractLipopeptide‐inspired lipidation of two short tripeptides, Phe‐Tyr‐β‐Ala and Tyr‐Phe‐β‐Ala leads to a pair of constitutional isomers of short peptide amphiphile (sPA). These two molecules were able to mimic lipopeptides properties and therefore could be used as potential drug delivery vehicles up to target sites. Further, these isomeric peptide amphiphiles were able to synthesize stable silver nanoparticles (AgNPs) in presence of sunlight without using any reducing agents. Interestingly, the presence of AgNPs significantly controls the secondary structures of these two isomeric sPAs and hence their self‐assembly. Therefore, such AgNPs‐sPA nanostructures are further used to develop antibacterial materials. Various microscopy and spectroscopy techniques were used to characterize these nanostructures. The antibacterial analysis demonstrated that owing to high biocompatibility and less cytotoxicity, designed sPA exhibited great potential in locally addressing bacterial infections. Thus, such a strategic design of lipidated tripeptide fragments can be tuned for potential therapeutic value for future applications.

Amyloid‐β Inspired Short Peptide Amphiphile Facilitates Synthesis of Silver Nanoparticles as Potential Antibacterial Agents

An amyloid-β inspired biocompatible short peptide amphiphile (sPA) molecule was used for controlled and targeted delivery of bioactive silver nanoparticles via transforming sPA nanostructures. Such sPA-AgNPs hybrid structures can be further used to develop antibacterial materials to combat emerging bacterial resistance. Due to the excellent antibacterial activity of silver, the growth of clinically relevant bacteria was inhibited in the presence of AgNPs-sPA hybrids. Bacterial tests demonstrated that the high biocompatibility and low cytotoxicity of the designed sPA allow it to work as a model drug delivery agent. It therefore shows great potential in locally addressing bacterial infections. The results of our study suggest that these nanodevices have the potential to trap and then engage in the facile delivery of their chemical payload at the target site, thereby working as potential delivery materials. This system has potential therapeutic value for the treatment of microbiota triggered progression of neurodegenerative diseases.

Short peptide amphiphile nanostructures facilitate sunlight-induced nanowelding of gold nanosheets.

An effortless thermoplasmonic welding of multi-shaped gold nanosheets is achieved by ordinary and simple sunlight irradiation. A light-matter interaction occurred via the nanogaps of smaller nanosheets, leading to the enhancement of the electromagnetic field and thus effectively concentrating the heat at the welding point. The sPA peptide nanostructure facilitates the nanowelding of small caged gold nanostructures.

Construction and immobilization of recombinant Bacillus subtilis with D-allulose 3-epimerase

D-allulose-3-epimerase (DPEase) is the key enzyme for isomerization of D-fructose to D-allulose. In order to improve its thermal stability, short amphiphilic peptides (SAP) were fused to the N-terminal of DPEase. SDS-PAGE analysis showed that the heterologously expressed DPEase folded correctly in Bacillus subtilis, and the protein size was 33 kDa. After incubation at 40 °C for 48 h, the residual enzyme activity of SAP1-DSDPEase was 58%. To make the recombinant B. subtilis strain reusable, cells were immobilized with a composite carrier of sodium alginate (SA) and titanium dioxide (TiO2). The results showed that 2% SA, 2% CaCl2, 0.03% glutaraldehyde solution and a ratio of TiO2 to SA of 1:4 were optimal for immobilization. Under these conditions, up to 82% of the activity of immobilized cells could be retained. Compared with free cells, the optimal reaction temperature of immobilized cells remained unchanged at 80 °C but the thermal stability improved. After 10 consecutive cycles, the mechanical strength remained unchanged, while 58% of the enzyme activity could be retained, with a conversion rate of 28.8% achieved. This study demonstrated a simple approach for using SAPs to improve the thermal stability of recombinant enzymes. Moreover, addition of TiO2 into SA during immobilization was demonstrated to increase the mechanical strength and reduce cell leakage.

Hydrophobized Short Peptide Amphiphile Functionalized Gold Nanoparticles as Antibacterial Biomaterials

AbstractIn search of new antibacterial agents and therapeutic strategies, during this antibiotic resistance crises, we developed a facile and effective antibacterial nanomaterial by using di‐phenylalanine based hydrophobized short peptide amphiphile (HsPA) and AuNPs. Such peptide amphiphile functionalised nanoparticles can provide a universal platform for medicinal applications owing to their unique physicochemical properties. The nanostructures of sPA and HsPA‐AuNPs hybrids were confirmed by spectroscopic and microscopy techniques. The antibacterial activity of these nanostructures were tested against clinically relevant bacteria such as Escherichia coli and found satisfactory. The HsPA acted as a model drug (Au(III) and AuNPs) delivery agent. Therefore such stimuli responsive, HsPA‐AuNPs system may open new therapeutic paradigms for emerging the theranostics.

Unexpected Role of Achiral Glycine in Determining the Suprastructural Handedness of Peptide Nanofibrils.

Helical supramolecular architectures play important structural and functional roles in biological systems. Although their occurrence is widely perceived to correlate to fundamental chiral units including l-amino acids and d-sugars, the detailed relationship between molecular and supramolecular handedness is still unclear. At the same time, although achiral units are practically always in close proximity to chiral ones by covalent linkage along a polymeric chain, their effect on supramolecular handedness has received relatively less attention. Here, we designed a set of short amphiphilic peptides, in which an achiral glycine residue was incorporated at the interface between the hydrophobic and hydrophilic segments. We observed that glycine incorporation caused dramatic variations in suprastructural handedness in self-assembled peptide nanofibrils, and the effect of the hydrophilic charged residue at the C-terminus on supramolecular handedness was demolished, leading to chiral truncation. Furthermore, molecular dynamics simulations and quantum chemistry calculations revealed that the unanticipated role of the glycine residue in regulating supramolecular handedness originated from its effect on the conformational preference of single β-strands. Importantly, reduced density gradient analyses on single β-strands indicated that, due to the lack of a side chain in glycine, intricate noncovalent interactions were produced among the neighboring amino acid side chains of the incorporated glycine and its local backbone, resulting in diverse β-strand conformations.

PH-Responsive Self-Assembly of Designer Aromatic Peptide Amphiphiles and Enzymatic Post-Modification of Assembled Structures.

Supramolecular fibrous materials in biological systems play important structural and functional roles, and therefore, there is a growing interest in synthetic materials that mimic such fibrils, especially those bearing enzymatic reactivity. In this study, we investigated the self-assembly and enzymatic post-modification of short aromatic peptide amphiphiles (PAs), Fmoc-LnQG (n = 2 or 3), which contain an LQG recognition unit for microbial transglutaminase (MTG). These aromatic PAs self-assemble into fibrous structures via π-π stacking interactions between the Fmoc groups and hydrogen bonds between the peptides. The intermolecular interactions and morphologies of the assemblies were influenced by the solution pH because of the change in the ionization states of the C-terminal carboxy group of the peptides. Moreover, MTG-catalyzed post-modification of a small fluorescent molecule bearing an amine group also showed pH dependency, where the enzymatic reaction rate was increased at higher pH, which may be because of the higher nucleophilicity of the amine group and the electrostatic interaction between MTG and the self-assembled Fmoc-LnQG. Finally, the accumulation of the fluorescent molecule on these assembled materials was directly observed by confocal fluorescence images. Our study provides a method to accumulate functional molecules on supramolecular structures enzymatically with the morphology control.

An overview of latest advances in exploring bioactive peptide hydrogels for neural tissue engineering.

Neural tissue engineering holds great potential in addressing current challenges faced by medical therapies employed for the functional recovery of the brain. In this context, self-assembling peptides have gained considerable interest owing to their diverse physicochemical properties, which enable them to closely mimic the biophysical characteristics of the native ECM. Additionally, in contrast to synthetic polymers, which lack inherent biological signaling, peptide-based nanomaterials could be easily designed to present essential biological cues to the cells to promote cellular adhesion. Moreover, injectability of these biomaterials further widens their scope in biomedicine. In this context, hydrogels obtained from short bioactive peptide sequences are of particular interest owing to their facile synthesis and highly tunable properties. In spite of their well-known advantages, the exploration of short peptides for neural tissue engineering is still in its infancy and thus detailed discussion is required to evoke interest in this direction. This review provides a general overview of various bioactive hydrogels derived from short peptide sequences explored for neural tissue engineering. The review also discusses the current challenges in translating the benefits of these hydrogels to clinical practices and presents future perspectives regarding the utilization of these hydrogels for advanced biomedical applications.

Insights into the co-assemblies formed by different aromatic short-peptide amphiphiles

The mechanism of co-assembly of different aromatic dipeptides has been studied using a combination of microscopy and spectroscopy techniques. At an equimolar ratio, the kinetics of the process is favored giving rise to alternate copolymers.

Antimicrobial and anti-inflammatory activities of short dodecapeptides derived from duck cathelicidin: Plausible mechanism of bactericidal action and endotoxin neutralization

Antimicrobial peptides (AMPs) have gained increasing attention to combat antibiotic-resistant pathogens. dCATH (duck cathelicidin) is a 20-residue avian cathelicidin with potent bactericidal activity. However, its therapeutic application is limited due to high mammalian cell cytotoxicity. To develop therapeutically useful AMPs with enhanced antimicrobial and cell-selective property, we designed a series of 12-meric (dodeca) short amphiphilic peptides based on dCATH. Among these, Trp and Lys-rich dCATH 12–4 and dCATH 12–5 exhibited higher selectivity towards bacterial cells than erythrocytes and macrophages. Additionally, these AMPs significantly reduced NO and TNF-α secretion in LPS-stimulated macrophage cells, suggesting their anti-inflammatory properties. Various fluorophore-based studies and confocal microscopic observations demonstrated that dCATH 12–4 and dCATH 12–5 could penetrate the bacterial cell membrane and accumulate in the cytoplasm, without disrupting membrane integrity. Results from the microscopic examination and gel-retardation DNA binding assay suggested that both the designed AMPs could bind with bacterial DNA, subsequently leading to cell death via arrest of DNA synthesis. Fluorescence spectroscopy and flow cytometry analysis revealed that the designed AMPs induced strong binding to LPS oligomers which resulted in dissociation of LPS aggregates, thereby preventing LPS from binding to the carrier protein lipopolysaccharide-binding protein (LBP) or alternatively to CD14 receptors of macrophage cells. Additionally, both dCATH 12–4 and dCATH 12-5 demonstrated synergistic actions with various conventional antibiotics against antibiotic resistant pathogens, thus indicating their ability as promising adjuncts to combination therapy. In summary, these findings contribute to the design of short AMPs with bactericidal and immunomodulatory properties for combating bacterial infection and sepsis.

Self-assembly of a Sequence-shuffled Short Peptide Amphiphile Triggered by Metal Ions into Terraced Nanodome-like Structures.

We highlight the structural diversity of strategically designed two short peptide amphiphiles (sPAs) and describe their structure-function relationship studies. The shuffling of two key amino acids, that is, tyrosine and phenylalanine, in a designed sPA lead to a pair of constitutional isomers. Such small and strategic alteration can bring a substantial change in the self-assembling pattern. Inspired from the naturally occurring metallopeptides, bioactive transition-metal ions were used for constructing the unusual nanostructures. Use of appropriate metal ions created bigger differences between the properties of these isomers and hence the self-assembly. Coordination of appropriate transition metal ions modifies the internal nanoscale structures of sPA, thus leading to the formation of vertically stacked terraced layers with decreasing size, which possess a high degree of dimensional regularity. We propose that such metal-induced terraced nanodome-like hierarchical self-assembly may have relevance for specific biotechnology applications.

Aldolase Cascade Facilitated by Self‐Assembled Nanotubes from Short Peptide Amphiphiles

Early evolution benefited from a complex network of reactions involving multiple C-C bond forming and breaking events that were critical for primitive metabolism. Nature gradually chose highly evolved and complex enzymes such as lyases to efficiently facilitate C-C bond formation and cleavage with remarkable substrate selectivity. Reported here is a lipidated short peptide which accesses a homogenous nanotubular morphology to efficiently catalyze C-C bond cleavage and formation. This system shows morphology-dependent catalytic rates, suggesting the formation of a binding pocket and registered enhancements in the presence of the hydrogen-bond donor tyrosine, which is exploited by extant aldolases. These assemblies showed excellent substrate selectivity and templated the formation of a specific adduct from a pool of possible adducts. The ability to catalyze metabolically relevant cascade transformations suggests the importance of such systems in early evolution.

Aldolase Cascade Facilitated by Self‐Assembled Nanotubes from Short Peptide Amphiphiles

AbstractEarly evolution benefited from a complex network of reactions involving multiple C−C bond forming and breaking events that were critical for primitive metabolism. Nature gradually chose highly evolved and complex enzymes such as lyases to efficiently facilitate C−C bond formation and cleavage with remarkable substrate selectivity. Reported here is a lipidated short peptide which accesses a homogenous nanotubular morphology to efficiently catalyze C−C bond cleavage and formation. This system shows morphology‐dependent catalytic rates, suggesting the formation of a binding pocket and registered enhancements in the presence of the hydrogen‐bond donor tyrosine, which is exploited by extant aldolases. These assemblies showed excellent substrate selectivity and templated the formation of a specific adduct from a pool of possible adducts. The ability to catalyze metabolically relevant cascade transformations suggests the importance of such systems in early evolution.

Transition metal ions induced secondary structural transformation in a hydrophobized short peptide amphiphile

Transition metal ions mediate the secondary structural transformation of hydrophobized sPA and can be applied to the design and development of stimuli-responsive nanomaterials.