Sequence-based identification of amyloidogenic ß-hairpins reveals a prostatic acid phosphatase fragment promoting semen amyloid formation.

ß-Structure-rich amyloid fibrils are hallmarks of several diseases, including Alzheimer's (AD), Parkinson's (PD), and type 2 diabetes (T2D). While amyloid fibrils typically consist of parallel ß-sheets, the anti-parallel ß-hairpin is a structural motif accessible to amyloidogenic proteins in their monomeric and oligomeric states. Here, to investigate implications of ß-hairpins in amyloid formation, potential ß-hairpin-forming amyloidogenic segments in the human proteome were predicted based on sequence similarity with ß-hairpins previously observed in Aß, α-synuclein, and islet amyloid polypeptide, amyloidogenic proteins associated with AD, PD, and T2D, respectively. These three ß-hairpins, established upon binding to the engineered binding protein ß-wrapin AS10, are characterized by proximity of two sequence segments rich in hydrophobic and aromatic amino acids, with high ß-aggregation scores according to the TANGO algorithm. Using these criteria, 2505 potential ß-hairpin-forming amyloidogenic segments in 2098 human proteins were identified. Characterization of a test set of eight protein segments showed that seven assembled into Thioflavin T-positive aggregates and four formed ß-hairpins in complex with AS10 according to NMR. One of those is a segment of prostatic acid phosphatase (PAP) comprising amino acids 185-208. PAP is naturally cleaved into fragments, including PAP(248-286) which forms functional amyloid in semen. We find that PAP(185-208) strongly decreases the protein concentrations required for fibril formation of PAP(248-286) and of another semen amyloid peptide, SEM1(86-107), indicating that it promotes nucleation of semen amyloids. In conclusion, ß-hairpin-forming amyloidogenic protein segments could be identified in the human proteome with potential roles in functional or disease-related amyloid formation.

Site Identification by Ligand Competitive Saturation-Biologics Approach for Structure-Based Protein Charge Prediction.

Protein-based therapeutics typically require high concentrations of the active protein, which can lead to protein aggregation and high solution viscosity. Such solution behaviors can limit the stability, bioavailability, and manufacturability of protein-based therapeutics and are directly influenced by the charge of a protein. Protein charge is a system property affected by its environment, including the buffer composition, pH, and temperature. Thus, the charge calculated by summing the charges of each residue in a protein, as is commonly done in computational methods, may significantly differ from the effective charge of the protein as these calculations do not account for contributions from bound ions. Here, we present an extension of the structure-based approach termed site identification by ligand competitive saturation-biologics (SILCS-Biologics) to predict the effective charge of proteins. The SILCS-Biologics approach was applied on a range of protein targets in different salt environments for which membrane-confined electrophoresis-determined charges were previously reported. SILCS-Biologics maps the 3D distribution and predicted occupancy of ions, buffer molecules, and excipient molecules bound to the protein surface in a given salt environment. Using this information, the effective charge of the protein is predicted such that the concentrations of the ions and the presence of excipients or buffers are accounted for. Additionally, SILCS-Biologics also produces 3D structures of the binding sites of ions on the proteins, which enable further analyses such as the characterization of protein surface charge distribution and dipole moments in different environments. Notable is the capability of the method to account for competition between salts, excipients, and buffers on the calculated electrostatic properties in different protein formulations. Our study demonstrates the ability of the SILCS-Biologics approach to predict the effective charge of proteins and its applicability in uncovering protein-ion interactions and their contributions to protein solubility and function.

Preserving the Integrity of Empirical Force Fields.

Generalized force fields (FFs) act as extensions to biomolecular FFs to provide a wide coverage of organic molecules. However, their precise application to an arbitrary molecule presents a separate challenge. We show that MATCH assigns different atom types and bonded and nonbonded parameters than CGenFF, and the AM1-BCC charge model, commonly used with GAFF/GAFF2, does not exactly reproduce the performance of the RESP charge model. The results indicate the need for caution when employing FFs to ensure their integrity with respect to their implementation and validation.

Computational design of a ß-wrapin's N-terminal domain with canonical and non-canonical amino acid modifications mimicking curcumin's proposed inhibitory function.

ß-wrapins are engineered binding proteins of which different mutants can bind and sequester amyloidogenic proteins amyloid-ß (Aß), islet amyloid polypeptide (IAPP), and α-synuclein (α-syn), thereby inhibiting their aggregation into amyloid fibrils. ß-wrapin AS10 is capable of binding and sequestering all three amyloidogenic monomers with micro-molar affinity, with its N-terminal domains remaining flexible and non-functional. Here, we computationally investigated the hypothesis that the anti-amyloid properties of AS10 can be amplified by redesigning its currently non-functional N-terminal domain with particular combinations of canonical and non-canonical amino acids (ncAAs) that can mimic the binding and inhibitory anti-amyloid function of curcumin, using a combination of molecular docking and molecular dynamics simulations. Our simulations suggest that the inhibitory mechanism attributed to the binding of the computationally designed AS10 N-terminal domain to the Aß fibril can act simultaneously to its sequestering properties for Aß which are attributed to the core of AS10. Thus, our study proposes that the N-terminal domain of AS10 can be further modified to amplify its anti-amyloid properties, resulting in a ß-wrapin that may simultaneously prohibit elongation to existing amyloid fibrils and also sequester amyloid monomers.

Computational and Experimental Protocols to Study Cyclo-dihistidine Self- and Co-assembly: Minimalistic Bio-assemblies with Enhanced Fluorescence and Drug Encapsulation Properties.

Our published studies on the self- and co-assembly of cyclo-HH peptides demonstrated their capacity to coordinate with Zn(II), their enhanced photoluminescence and their ability to self-encapsulate epirubicin, a chemotherapy drug. Here, we provide a detailed description of computational and experimental methodology for the study of cyclo-HH self- and co-assembling mechanisms, photoluminescence, and drug encapsulation properties. We outline the experimental protocols, which involve fluorescence spectroscopy, transmission electron microscopy, and atomic force microscopy protocols, as well as the computational protocols, which involve structural and energetic analysis of the assembled nanostructures. We suggest that the computational and experimental methods presented here can be generalizable, and thus can be applied in the investigation of self- and co-assembly systems involving other short peptides, encapsulating compounds and binding to ions, beyond the particular ones presented here.

Montmorillonite clay-based sorbents decrease the bioavailability of per- and polyfluoroalkyl substances (PFAS) from soil and their translocation to plants.

Consumption of food and water contaminated with per- and polyfluoroalkyl substances (PFAS) presents a significant risk for human exposure. There is limited data on high affinity sorbents that can be used to reduce the bioavailability of PFAS from soil and translocation to plants and garden produce. To address this need, montmorillonite clay was amended with the nutrients carnitine and choline to increase the hydrophobicity of the sorbent and the interlayer spacing. In this study, the binding of PFOA (perfluorooctanoic acid) and PFOS (perfluorooctanesulfonic acid) to parent and amended clays was characterized. Isothermal analyses were conducted at pH 7 and ambient temperature to simulate environmentally-relevant conditions. The data for all tested sorbents fit the Langmuir model indicating saturable binding sites with high capacities and affinities under neutral conditions. Amended montmorillonite clays had increased capacities for PFOA and PFOS (0.51-0.71 mol kg-1) compared to the parent clay (0.37-0.49 mol kg-1). Molecular dynamics (MD) simulations suggested that hydrophobic and electrostatic interactions at the terminal fluorinated carbon chains of PFAS compounds were major modes of surface interaction. The safety and efficacy of the clays were confirmed in a living organism (Lemna minor), where clays (at 0.1% inclusion) allowed for increased growth compared to PFOA and PFOS controls (p ≤ 0.01). Importantly, soil studies showed that 2% sorbent inclusion could significantly reduce PFAS bioavailability from soil (up to 74%). Studies in plants demonstrated that inclusion of 2% sorbent significantly reduced PFAS residues in cucumber plants (p ≤ 0.05). These results suggest that nutrient-amended clays could be included in soil to decrease PFAS bioavailability and translocation of PFAS to plants.

Modification of a Single Atom Affects the Physical Properties of Double Fluorinated Fmoc-Phe Derivatives.

Supramolecular hydrogels formed by the self-assembly of amino-acid based gelators are receiving increasing attention from the fields of biomedicine and material science. Self-assembled systems exhibit well-ordered functional architectures and unique physicochemical properties. However, the control over the kinetics and mechanical properties of the end-products remains puzzling. A minimal alteration of the chemical environment could cause a significant impact. In this context, we report the effects of modifying the position of a single atom on the properties and kinetics of the self-assembly process. A combination of experimental and computational methods, used to investigate double-fluorinated Fmoc-Phe derivatives, Fmoc-3,4F-Phe and Fmoc-3,5F-Phe, reveals the unique effects of modifying the position of a single fluorine on the self-assembly process, and the physical properties of the product. The presence of significant physical and morphological differences between the two derivatives was verified by molecular-dynamics simulations. Analysis of the spontaneous phase-transition of both building blocks, as well as crystal X-ray diffraction to determine the molecular structure of Fmoc-3,4F-Phe, are in good agreement with known changes in the Phe fluorination pattern and highlight the effect of a single atom position on the self-assembly process. These findings prove that fluorination is an effective strategy to influence supramolecular organization on the nanoscale. Moreover, we believe that a deep understanding of the self-assembly process may provide fundamental insights that will facilitate the development of optimal amino-acid-based low-molecular-weight hydrogelators for a wide range of applications.

Combining Experimental Isotherms, Minimalistic Simulations, and a Model to Understand and Predict Chemical Adsorption onto Montmorillonite Clays.

An attractive approach to minimize human and animal exposures to toxic environmental contaminants is the use of safe and effective sorbent materials to sequester them. Montmorillonite clays have been shown to tightly bind diverse toxic chemicals. Due to their promise as sorbents to mitigate chemical exposures, it is important to understand their function and rapidly screen and predict optimal clay-chemical combinations for further testing. We derived adsorption free-energy values for a structurally and physicochemically diverse set of toxic chemicals using experimental adsorption isotherms performed in the current and previous studies. We studied the diverse set of chemicals using minimalistic MD simulations and showed that their interaction energies with calcium montmorillonite clays calculated using simulation snapshots in combination with their net charge and their corresponding solvent's dielectric constant can be used as inputs to a minimalistic model to predict adsorption free energies in agreement with experiments. Additionally, experiments and computations were used to reveal structural and physicochemical properties associated with chemicals that can be adsorbed to calcium montmorillonite clay. These properties include positively charged groups, phosphine groups, halide-rich moieties, hydrogen bond donor/acceptors, and large, rigid structures. The combined experimental and computational approaches used in this study highlight the importance and potential applicability of analogous methods to study and design novel advanced sorbent systems in the future, broadening their applicability for environmental contaminants.

Self-Assembled Peptide Nano-Superstructure towards Enzyme Mimicking Hydrolysis.

The structural arrangement of amino acid residues in native enzymes underlies their remarkable catalytic properties, thus providing a notable point of reference for designing potent yet simple biomimetic catalysts. Herein, we describe a minimalistic approach to construct a dipeptide-based nano-superstructure with enzyme-like activity. The self-assembled biocatalyst comprises one peptide as a single building block, readily synthesized from histidine. Through coordination with zinc ion, the peptide self-assembly procedure allows the formation of supramolecular ß-sheet ordered nanocrystals, which can be used as basic units to further construct higher-order superstructure. As a result, remarkable hydrolysis activity and enduring stability are demonstrated. Our work exemplifies the use of a bioinspired supramolecular assembly approach to develop next-generation biocatalysts for biotechnological applications.

Protection of Oxygen-Sensitive Enzymes by Peptide Hydrogel.

Molecular oxygen (O2) is a highly reactive oxidizing agent and is harmful to many biological and industrial systems. Although O2 often interacts via metals or reducing agents, a binding mechanism involving an organic supramolecular structure has not been described to date. In this work, the prominent dipeptide hydrogelator fluorenylmethyloxycarbonyl-diphenylalanine is shown to encage O2 and significantly limit its diffusion and penetration through the hydrogel. Molecular dynamics simulations suggested that the O2 binding mechanism is governed by pockets formed between the aromatic rings in the supramolecular structure of the gel, which bind O2 through hydrophobic interactions. This phenomenon is harnessed to maintain the activity of the O2-hypersensitive enzyme [FeFe]-hydrogenase, which holds promising potential for utilizing hydrogen gas for sustainable energy applications. Hydrogenase encapsulation within the gel allows hydrogen production following exposure to ambient O2. This phenomenon may lead to utilization of this low molecular weight gelator in a wide range of O2-sensitive applications.

Enhanced adsorption of per- and polyfluoroalkyl substances (PFAS) by edible, nutrient-amended montmorillonite clays.

Humans and animals are frequently exposed to PFAS (per- and polyfluoroalkyl substances) through drinking water and food; however, no therapeutic sorbent strategies have been developed to mitigate this problem. Montmorillonites amended with the common nutrients, carnitine and choline, were characterized for their ability to bind 4 representative PFAS (PFOA, PFOS, GenX, and PFBS). Adsorption/desorption isothermal analysis showed that PFOA, PFOS (and a mixture of the two) fit the Langmuir model with high binding capacity, affinity and enthalpy at conditions simulating the stomach. A low percentage of desorption occurred at conditions simulating the intestine. The results suggested that hydrophobic and electrostatic interactions, and hydrogen bonding were responsible for sequestering PFAS into clay interlayers. Molecular dynamics (MD) simulations suggested the key mode of interaction of PFAS was through fluorinated carbon chains, and confirmed that PFOA and PFOS had enhanced binding to amended clays compared to GenX and PFBS. The safety and efficacy of amended montmorillonite clays were confirmed in Hydra vulgaris, where a mixture of amended sorbents delivered the highest protection against a PFAS mixture. These important results suggest that the inclusion of edible, nutrient-amended clays with optimal affinity, capacity, and enthalpy can be used to decrease the bioavailability of PFAS from contaminated drinking water and diets.

Hydroxylated Chalcones as Aryl Hydrocarbon Receptor Agonists: Structure-Activity Effects.

Hydroxylated chalcones are phytochemicals which are biosynthetic precursors of flavonoids and their 1,3-diaryl-prop-2-en-1-one structure is used as a scaffold for drug development. In this study, the structure-dependent activation of aryl hydrocarbon receptor (AhR)-responsive CYP1A1, CYP1B1, and UGT1A1 genes was investigated in Caco2 colon cancer cells and in non-transformed young adult mouse colonocytes (YAMC) cells. The effects of a series of di- and trihydroxychalcones as AhR agonists was structure dependent with maximal induction of CYP1A1, CYP1B1, and UGT1A1 in Caco2 cells observed for compounds containing 2,2'-dihydroxy substituents and this included 2,2'-dihydroxy-, 2,2',4'-trihydroxy-, and 2,2',5'-trihydroxychalcones. In contrast, 2',4,5'-, 2'3',4'-, 2',4,4'-trihydroxy, and 2',3-, 2',4-, 2',4'-, and 2',5-dihydroxychalcones exhibited low to non-detectable AhR activity in Caco2 cells. In addition, all of the hydroxychalcones exhibited minimal to non-detectable activity in YAMC cells, whereas 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced CYP1A1, CYP1B1, and UGT1A1 in Caco2 and YAMC cells. The activity of AhR-active chalcones was confirmed by determining their effects in AhR-deficient Caco2 cells. In addition, 2,2'-dihydroxychalcone induced CYP1A1 protein and formation of an AhR-DNA complex in an in vitro assay. Simulation and modeling studies of hydroxylated chalcones confirmed their interactions with the AhR ligand-binding domain and were consistent with their structure-dependent activity as AhR ligands. Thus, this study identifies hydroxylated chalcones as AhR agonists with potential for these phytochemicals to impact AhR-mediated colonic pathways.

Insights into the interactions of bisphenol and phthalate compounds with unamended and carnitine-amended montmorillonite clays.

Montmorillonite clays could be promising sorbents to mitigate toxic compound exposures. Bisphenols A (BPA) and S (BPS) as well as phthalates, dibutyl phthalate (DBP) and di-2-ethylhexyl phthalate (DEHP), are ubiquitous environmental contaminants linked to adverse health effects. Here, we combined computational and experimental methods to investigate the ability of montmorillonite clays to sorb these compounds. Molecular dynamics simulations predicted that parent, unamended, clay has higher binding propensity for BPA and BPS than for DBP and DEHP; carnitine-amended clay improved BPA and BPS binding, through carnitine simultaneously anchoring to the clay through its quaternary ammonium cation and forming hydrogen bonds with BPA and BPS. Experimental isothermal analysis confirmed that carnitine-amended clay has enhanced BPA binding capacity, affinity and enthalpy. Our studies demonstrate how computational and experimental methods, combined, can characterize clay binding and sorption of toxic compounds, paving the way for future investigation of clays to reduce BPA and BPS exposure.

Enhanced Fluorescence for Bioassembly by Environment-Switching Doping of Metal Ions.

The self-assembly of cyclodipeptides composed of natural aromatic amino acids into supramolecular structures of diverse morphologies with intrinsic emissions in the visible light region is demonstrated. The assembly process can be halted at the initial oligomerization by coordination with zinc ions, with the most prominent effect observed for cyclo-dihistidine (cyclo-HH). This process is mediated by attracting and pulling of the metal ions from the solvent into the peptide environment, rather than by direct interaction in the solvent as commonly accepted, thus forming an "environment-switching" doping mechanism. The doping induces a change of cyclo-HH molecular configurations and leads to the formation of pseudo "core/shell" clusters, comprising peptides and zinc ions organized in ordered conformations partially surrounded by relatively amorphous layers, thus significantly enhancing the emissions and allowing the application of the assemblies for ecofriendly color-converted light emitting diodes. These findings shed light into the very initial coordination procedure and elucidate an alternative mechanism of metal ions doping on biomolecules, thus presenting a promising avenue for integration of the bioorganic world and the optoelectronic field.

High-Efficiency Fluorescence through Bioinspired Supramolecular Self-Assembly.

Peptide self-assembly has attracted extensive interest in the field of eco-friendly optoelectronics and bioimaging due to its inherent biocompatibility, intrinsic fluorescence, and flexible modulation. However, the practical application of such materials was hindered by the relatively low quantum yield of such assemblies. Here, inspired by the molecular structure of BFPms1, we explored the "self-assembly locking strategy" to design and manipulate the assembly of metal-stabilized cyclic(l-histidine-d-histidine) into peptide material with the high-fluorescence efficiency. We used this bioorganic material as an emissive layer in photo- and electroluminescent prototypes, demonstrating the feasibility of utilizing self-assembling peptides to fabricate a biointegrated microchip that incorporates eco-friendly and tailored optoelectronic properties. We further employed a "self-encapsulation" strategy for constructing an advanced nanocarrier with integrated in situ monitoring. The strategy of the supramolecular capture of functional components exemplifies the use of bioinspired organic chemistry to provide frontiers of smart materials, potentially allowing a better interface between sustainable optoelectronics and biomedical applications.

Computational evolution of an RNA-binding protein towards enhanced oxidized-RNA binding.

The oxidation of RNA has been implicated in the development of many diseases. Among the four ribonucleotides, guanosine is the most susceptible to oxidation, resulting in the formation of 8-oxo-7,8-dihydroguanosine (8-oxoG). Despite the limited knowledge about how cells regulate the detrimental effects of oxidized RNA, cellular factors involved in its regulation have begun to be identified. One of these factors is polynucleotide phosphorylase (PNPase), a multifunctional enzyme implicated in RNA turnover. In the present study, we have examined the interaction of PNPase with 8-oxoG in atomic detail to provide insights into the mechanism of 8-oxoG discrimination. We hypothesized that PNPase subunits cooperate to form a binding site using the dynamic SFF loop within the central channel of the PNPase homotrimer. We evolved this site using a novel approach that initially screened mutants from a library of beneficial mutations and assessed their interactions using multi-nanosecond Molecular Dynamics simulations. We found that evolving this single site resulted in a fold change increase in 8-oxoG affinity between 1.2 and 1.5 and/or selectivity between 1.5 and 1.9. In addition to the improvement in 8-oxoG binding, complementation of K12 Δpnp with plasmids expressing mutant PNPases caused increased cell tolerance to H2O2. This observation provides a clear link between molecular discrimination of RNA oxidation and cell survival. Moreover, this study provides a framework for the manipulation of modified-RNA protein readers, which has potential application in synthetic biology and epitranscriptomics.

Interactions between Curcumin Derivatives and Amyloid-ß Fibrils: Insights from Molecular Dynamics Simulations.

The aggregation of amyloid-ß (Aß) peptides into senile plaques is a hallmark of Alzheimer's disease (AD) and is hypothesized to be the primary cause of AD related neurodegeneration. Previous studies have shown the ability of curcumin to both inhibit the aggregation of Aß peptides into oligomers or fibrils and reduce amyloids in vivo. Despite the promise of curcumin and its derivatives to serve as diagnostic, preventative, and potentially therapeutic AD molecules, the mechanism by which curcumin and its derivatives bind to and inhibit Aß fibrils' formation remains elusive. Here, we investigated curcumin and a set of curcumin derivatives in complex with a hexamer peptide model of the Aß1-42 fibril using nearly exhaustive docking, followed by multi-ns molecular dynamics simulations, to provide atomistic-detail insights into the molecules' binding and inhibitory properties. In the vast majority of the simulations, curcumin and its derivatives remain firmly bound in complex with the fibril through primarily three different principle binding modes, in which the molecules interact with residue domain 17LVFFA21, in line with previous experiments. In a small subset of these simulations, the molecules partly dissociate the outermost peptide of the Aß1-42 fibril by disrupting ß-sheets within the residue domain 12VHHQKLVFF20. A comparison between binding modes leading or not leading to partial dissociation of the outermost peptide suggests that the latter is attributed to a few subtle key structural and energetic interaction-based differences. Interestingly, partial dissociation appears to be either an outcome of high affinity interactions or a cause leading to high affinity interactions between the molecules and the fibril, which could partly serve as a compensation for the energy loss in the fibril due to partial dissociation. In conjunction with this, we suggest a potential inhibition mechanism of Αß1-42 aggregation by the molecules, where the partially dissociated 16KLVFF20 domain of the outermost peptide could either remain unstructured or wrap around to form intramolecular interactions with the same peptide's 29GAIIG33 domain, while the molecules could additionally act as a patch against the external edge of the second outermost peptide's 16KLVFF20 domain. Thereby, individually or concurrently, these could prohibit fibril elongation.

Molecular Mechanism for Attractant Signaling to DHMA by E. coli Tsr.

The attractant chemotaxis response of Escherichia coli to norepinephrine requires that it be converted to 3,4-dihydroxymandelic acid (DHMA) by the monoamine oxidase TynA and the aromatic aldehyde dehydrogenase FeaB. DHMA is sensed by the serine chemoreceptor Tsr, and the attractant response requires that at least one subunit of the periplasmic domain of the Tsr homodimer (pTsr) has an intact serine-binding site. DHMA that is generated in vivo by E. coli is expected to be a racemic mixture of the (R) and (S) enantiomers, so it has been unclear whether one or both chiral forms are active. Here, we used a combination of state-of-the-art tools in molecular docking and simulations, including an in-house simulation-based docking protocol, to investigate the binding properties of (R)-DHMA and (S)-DHMA to E. coli pTsr. Our studies computationally predicted that (R)-DHMA should promote a stronger attractant response than (S)-DHMA because of a consistently greater-magnitude piston-like pushdown of the pTsr α-helix 4 toward the membrane upon binding of (R)-DHMA than upon binding of (S)-DHMA. This displacement is caused primarily by interaction of DHMA with Tsr residue Thr156, which has been shown by genetic studies to be critical for the attractant response to L-serine and DHMA. These findings led us to separate the two chiral species and test their effectiveness as chemoattractants. Both the tethered cell and motility migration coefficient assays validated the prediction that (R)-DHMA is a stronger attractant than (S)-DHMA. Our study demonstrates that refined computational docking and simulation studies combined with experiments can be used to investigate situations in which subtle differences between ligands may lead to diverse chemotactic responses.

Designer Amyloid Cell-Penetrating Peptides for Potential Use as Gene Transfer Vehicles.

Cell-penetrating peptides are used extensively to deliver molecules into cells due to their unique characteristics such as rapid internalization, charge, and non-cytotoxicity. Amyloid fibril biomaterials were reported as gene transfer or retroviral infection enhancers; no cell internalization of the peptides themselves is reported so far. In this study, we focus on two rationally and computationally designed peptides comprised of ß-sheet cores derived from naturally occurring protein sequences and designed positively charged and aromatic residues exposed at key residue positions. The ß-sheet cores bestow the designed peptides with the ability to self-assemble into amyloid fibrils. The introduction of positively charged and aromatic residues additionally promotes DNA condensation and cell internalization by the self-assembled material formed by the designed peptides. Our results demonstrate that these designer peptide fibrils can efficiently enter mammalian cells while carrying packaged luciferase-encoding plasmid DNA, and they can act as a protein expression enhancer. Interestingly, the peptides additionally exhibited strong antimicrobial activity against the enterobacterium Escherichia coli.