Search

Supramolecular helical structures formed by the assembly of biological and bio-inspired building blocks (typically amino acids and peptides) are an exciting class of materials with prospective applications in biomedical technologies and electronics.

Flexible and biocompatible metal peptide frameworks (MPFs) derived from short and ultra-short peptides have been explored for the storage of greenhouse gases, molecular recognition, and chiral transformations. In addition to short flexible peptides, peptides with specifically folded conformations have recently been utilized to fabricate a variety of metal helix frameworks (MHFs). The secondary structures of the peptides govern the structure-assembly relationship and thereby control the formation of three-dimensional (3D)-MHFs. Particularly, the hierarchical structural organization of peptide-based MHFs has not yet been discussed in detail. Here, we describe the recent progress of metal-driven folded peptide assembly to construct 3D porous structures for use in future energy storage, chiral recognition, and biomedical applications, which could be envisioned as an alternative to the conventional metal-organic frameworks (MOFs).

Carbon quantum dots (CDs) are a class of emerging carbonaceous nanomaterials that have received considerable attention due to their excellent fluorescent properties, extremely small size, ability to penetrate cells and tissues, ease of synthesis, surface modification, low cytotoxicity, and superior water dispersion. In light of these properties, CDs are extensively investigated as candidates for bioimaging probes, efficient drug carriers, and disease diagnostics. Functionalized CDs represent a promising therapeutic candidate for ocular diseases. Here, this work reviews the potential use of functionalized CDs in the diagnosis and treatment of eye-related diseases, including the treatment of macular and anterior segment diseases, as well as targeting Aβ amyloids in the retina.

Protein misfolding and aggregation are associated with human diseases and aging. However, microorganisms widely exploit the self-propagating properties of misfolded infectious protein particles, prions, as epigenetic information carriers that drive various phenotypic adaptations and encode molecular information. Microbial prion research has faced a paradigm shift in recent years, with breakthroughs that demonstrate the great functional and structural diversity of these agents. Here, we outline unorthodox examples of microbial prions in yeast and other microorganisms, focusing on their noncanonical functions. We discuss novel molecular mechanisms for the inheritance of conformationally-encoded epigenetic information and the evolutionary advantages they confer. Lastly, in light of recent advancements in the field of molecular self-assembly, we present a hypothesis regarding the existence of non-proteinaceous prion-like entities.

The apparent piezoelectricity of biological materials is not yet fully understood at the molecular level. In particular, dynamic noncovalent interactions, such as host-guest binding, are not included in the classical piezoelectric model, which limits the rational design of eco-friendly piezoelectric supramolecular materials. Here, inspired by the conformation-dependent mechanoresponse of the Piezo channel proteins, we show that guest-host interactions can amplify the electromechanical response of a conformationally mobile peptide metal-organic framework (MOF) based on the endogenous carnosine dipeptide, demonstrating a new type of adaptive piezoelectric supramolecular material. Density functional theory (DFT) predictions validated by piezoresponse force microscopy (PFM) measurements show that directional alignment of the guest molecules in the host carnosine-zinc peptide MOF channel determines the macroscopic electromechanical properties. We produce stable, robust 1.4 V open-circuit voltage under applied force of 25 N with a frequency of 0.1 Hz. Our findings demonstrate that the regulation of host-guest interactions could serve as an efficient method for engineering sustainable peptide-based power generators.

The formation of ordered protein and peptide assemblies is a phenomenon related to a wide range of human diseases. However, the mechanism of assembly at the molecular level remains largely unknown. Minimal models enable the exploration of the underlying interactions that are at the core of such self-assembly processes. In particular, the ability of phenylalanine, a single aromatic amino acid, to form an amyloid-like structure has challenged the previous dogma viewing a peptide backbone as a prerequisite for assembly. The driving forces controlling the nucleation and assembly in the absence of a peptide backbone remain to be identified. Here, aiming to unravel these forces, we explored the kinetics and thermodynamics of three phenylalanine-containing molecules during their assembly process: the amino acid phenylalanine, which accumulates in phenylketonuria patients, the diphenylalanine core-motif of the amyloid β peptide related to Alzheimer's disease, and the extended triphenylalanine peptide which forms a range of distinct nanostructuresiin vitro/i. We found that the aggregation propensity, regarding the critical monomer concentration, strongly increases with size, with triphenylalanine being the most aggregation-prone species under our experimental conditions. In the context of classical nucleation theory, this increase in aggregation propensity can be attributed to the larger free energy decrease upon aggregation of larger peptides and is not due to the presence/absence of a peptide bondiper se/i. Taken together, this work provides insights into the aggregation processes of chemically simple systems and suggests that both backbone-containing peptides and backbone-lacking amino acids assemble through a similar mechanism, thus supporting the classification of amino acids in the continuum of amyloid-forming building blocks.

The formation of metabolite fibrillar assemblies represents a paradigm shift in the study of human metabolic disorders. Yet, direct clinical relevance has been attributed only to metabolite crystals. A notable example for metabolite crystallization is calcium oxalate crystals observed in various diseases, including primary hyperoxaluria. We unexpectedly observed retinal damage among young hyperoxaluria patients in the absence of crystals. Exploring the possible formation of alternative supramolecular organizations and their biological role, here we show that oxalate can form ordered fibrils with no associated calcium. These fibrils inflict intense retinal cytotoxicity in cultured cells. A rat model injected with oxalate fibrils recaptures patterns of retinal dysfunction observed in patients. Antibodies purified from hyperoxaluria patient sera recognize oxalate fibrils regardless of the presence of calcium. These findings highlight a new molecular basis for oxalate-associated disease, and to our knowledge provide the first direct clinical indication for the pathogenic role of metabolite fibrillar assemblies. Primary hyperoxaluria is a rare genetic condition which involves elevated levels of oxalate in the body, but its role in causing symptoms is not well understood. Here self-assembled oxalate fibrils are implicated in the mechanism of retinopathy in primary hyperoxaluria in humans.

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 β-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.

Phenylalanine was the minimalistic and first of numerous nonproteinaceous building blocks to be demonstrated to form amyloid-like fibrils. This unexpected organization of such a simple building block into canonical architecture, which was previously observed only with proteins and peptides, has numerous implications for medicine and supramolecular chemistry. However, the morphology of phenylalanine fibrils and their mechanical properties was never characterized in solutions. Here, using electron and atomic force microscopy, we analyze the morphological and mechanical properties of phenylalanine fibrils in both air and fluids. The fibrils demonstrate an exceptionally high Young's modulus (up to 30 GPa) and are found to be composed of intertwined protofilaments in a helical or twisted ribbon morphology. In addition, X-ray scattering experiments provide convincing evidence of an amyloidal cross-β-like secondary structure within the nanoassemblies. Furthermore, increasing the phenylalanine concentration results in the formation of highly homogenous, noncrystalline, self-healing hydrogels that display storage and loss moduli significantly higher than similar noncovalently cross-linked biomolecular nanofibrillar scaffolds. These remarkably stiff nanofibrillar hydrogels can be harnessed for various technological and biomedical applications, such as self-healing, printable, structural, load-bearing 3D scaffolds. The properties of this simple but quite remarkable hydrogel open a possibility to utilize it in the biomaterial industry.

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.

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.

Conformational transition of proteins and peptides into highly stable, β-sheet-rich structures is observed in many amyloid-associated neurodegenerative disorders, yet the precise mechanism of amyloid formation at the molecular level remains poorly understood due to the complex molecular structures. Short peptides provide simplified models for studying the molecular basis of the assembly mechanism that governs β-sheet fibrillation processes underlying the formation and inhibition of amyloid-like structures. Herein, we report a supramolecular coassembly strategy for the inhibition and transformation of stable β-sheet-rich amyloid-derived dipeptide self-assemblies into adaptable secondary structural fibrillar assemblies by mixing with bipyridine derivatives. The interplay between the type and mixing ratio of bipyridine derivatives allowed the variable coassembly process with stimuli-responsive functional properties, studied by various experimental characterizations and computational methods. Furthermore, the resulting coassemblies showed functional redox- and photoresponsive properties, making them promising candidates for controllable drug release and fluorescent imprint. This work presents a coassembly strategy not only to explore the mechanism of amyloid-like structure formation and inhibition at the molecular level but also to manipulate amyloid-like structures into responsive supramolecular coassemblies for material science and biotechnology applications.

Metabolites are essential for the normal operation of cells and fulfill various physiological functions. It was recently found that in several metabolic disorders, the associated metabolites could self-assemble to generate amyloid-like structures, similar to canonical protein amyloids that have a role in neurodegenerative disorders. Yet, assemblies with typical amyloid characteristics are also known to have physiological function. In addition, many non-natural proteins and peptides presenting amyloidal properties have been used for the fabrication of functional nanomaterials. Similarly, functional metabolite assemblies are also found in nature, demonstrating various physiological roles. A notable example is the structural color formed by guanine crystals or fluorescent crystals in feline eyes responsible for enhanced night vision. Moreover, some metabolites have been used for the in vitro fabrication of functional materials, such as glycine crystals presenting remarkable piezoelectric properties or indigo films used to assemble organic semiconductive electronic devices. Therefore, we believe that the study of metabolite assemblies is not only important in order to understand their role in normal physiology and in pathology, but also paves a new route in exploring the fabrication of organic, bio-compatible materials.

The formation of ordered nanostructures by metabolites is gaining increased interest due to the simplicity of the building blocks and their natural occurrence. Specifically, aromatic amino acids possess the ability to form ordered supramolecular interactions due to their limited solubility in aqueous solution. Unexpectedly, l-tyrosine (l-Tyr) is almost 2 orders of magnitude less soluble in water compared to l-phenylalanine (l-Phe). However, the underlying mechanism is not fully understood as l-Tyr is more polar. Here, we explore the utilization of insoluble tyrosine assemblies for technological applications and their molecular basis by manipulating the basic building blocks of tightly packed dimers. We show that the addition of an amyloid inhibition agent increases l-Tyr solubility due to the disruption of the dimer formation. The molecular organization grants the l-Tyr crystal higher thermal stability and mechanical properties between three amino acids. Additionally, l-Tyr crystals are shown to generate high and stable piezoelectric power outputs under mechanical pressure in a sandwich device. By incorporating the rigid l-Tyr crystals into a soft polymer, a mechano-responsive bending composite was fabricated. Furthermore, the l-Tyr crystalline needles exhibit an active photowaveguiding property, making them promising candidates for the generation of photonic biomaterial-based devices. The present work exemplifies a feasible strategy to explore physical properties of supramolecular self-assemblies comprises minimalistic naturally occurring building blocks and their applications in energy harvesting, photonic devices, stretchable electronics, and soft robotics.

The unique physiochemical properties and multiscale organization of layered materials draw the attention of researchers across a wide range of scientific disciplines. Layered structures are commonly found in diverse biological systems where they fulfill various functions. A prominent example of layered biological materials is the organization of proteins and polypeptides into the archetypal aggregated amyloidal structures. While the organization of proteins into amyloid structures was initially associated with various degenerative disorders, it was later revealed that proteins not related to any disease could also form identical layered assemblies. Thus, it appears that the ability of peptides and proteins to produce amyloid-like aggregates represents a generic property of polyamides to assemble into higher order fibrillar structures. In the aggregated state, the peptide backbone forms β-sheet structures which are further organized into layered arrangements. We have recently extended the identified amyloidogenic building blocks to include not only peptides or proteins, but also single amino acids and other metabolites. High resolution spectroscopy and crystallography analyses confirm the clear potential of amino acids and other metabolites to form layered amyloid-like aggregates showing biophysical and biochemical properties similar to protein amyloids. Therefore, the generic propensity of peptides and proteins backbones to assemble into layered organizations may emanate from their basic building block, the amino acid. In this Account, we aim to introduce the concept of supramolecular β-sheet organization of single amino acids and to present an analysis of their layered-structure organization based on single crystal structures. We demonstrate that, despite the different side-chains that considerably vary in their chemical properties, all coded amino acids display a layer-like assembly stabilized by α-amine to α-carboxyl H-bonds, resembling supramolecular β-sheet structures, while the side-chains determine the higher order organization of the layers. Our work presents the first analysis of the β-sheet propensity of single amino acids in their unbound form, indicating an evolutionary predisposition. We classify the amino acids β-sheet propensity on the basis of the interlayer separation distance in the crystal packing, which correlates well with previously reported classifications based on various criteria, such as hydrophobicity, steric bulkiness, and folding. In addition, we demonstrate that the relative direction of α-amine to α-carboxyl H-bonding pattern provides critical insights regarding the stabilization of parallel versus antiparallel β-sheet structures by the various amino acids. Taken together, our analysis of amino acid crystals provides substantial information regarding protein folding and dynamics and could serve as basic rules set for the design of potential building blocks for molecular self-assembly to produce functional materials of tunable properties, an important objective of bottom-up nanotechnology.

The COP9 signalosome (CSN) is an essential eight-subunit repressor of light-regulated development in Arabidopsis. This complex has also been identified in animals, though its developmental role remains obscure. CSN subunits have been implicated in various cellular processes, suggesting a possible role for the CSN as an integrator of multiple signaling pathways. In order to elucidate the function of the CSN in animals, a Drosophila model system has previously been established. Gel-filtration analysis with antibodies against CSN subunits 4, 5 and 7 revealed that these proteins act as a complex in Drosophila that is similar in size to the plant and mammalian complexes. Null mutations in either one of two subunits, CSN4 or CSN5, are larval lethal. Successful embryogenesis appears to be a consequence of maternal contribution of the complex. Biochemical analysis indicates that the different subunits are found in both CSN-dependent and CSN-independent forms, and that these forms are differentially affected by the mutations. Phenotypic characterization of these two mutants indicates that they show both shared and unique phenotypes, which suggest specific roles for each subunit. Both mutants have defective oocyte and embryo patterning, and defects in response to DNA damage, while csn5 mutants develop melanotic tumors and csn4 mutants have phenotypes reminiscent of defects in ecdysone signaling.

During organogenesis, PAX6 is required for establishment of various progenitor subtypes within the central nervous system, eye and pancreas. PAX6 expression is maintained in a variety of cell types within each organ, although its role in each lineage and how it acquires cell-specific activity remain elusive. Herein, we aimed to determine the roles and the hierarchical organization of the PAX6-dependent gene regulatory network during the differentiation of the retinal pigmented epithelium (RPE). Somatic mutagenesis of Pax6 in the differentiating RPE revealed that PAX6 functions in a feed-forward regulatory loop with MITF during onset of melanogenesis. PAX6 both controls the expression of an RPE isoform of Mitf and synergizes with MITF to activate expression of genes involved in pigment biogenesis. This study exemplifies how one kernel gene pivotal in organ formation accomplishes a lineage-specific role during terminal differentiation of a single lineage., Author Summary It is currently poorly understood how a single developmental transcription regulator controls early specification as well as a broad range of highly specialized differentiation schemes. PAX6 is one of the most extensively investigated factors in central nervous system development, yet its role in execution of lineage-specific programs remains mostly elusive. Here, we directly investigated the involvement of PAX6 in the differentiation of one lineage, the retinal pigmented epithelium (RPE), a neuroectodermal-derived tissue that is essential for retinal development and function. We revealed that PAX6 accomplishes its role through a unique regulatory interaction with the transcription factor MITF, a master regulator of the pigmentation program. During the differentiation of the RPE, PAX6 regulates the expression of an RPE-specific isoform of Mitf and importantly, at the same time, PAX6 functions together with MITF to directly activate the expression of downstream genes required for pigment biogenesis. These findings provide comprehensive insight into the gene hierarchy that controls RPE development: from a kernel gene (a term referring to the upper-most gene in the gene regulatory network) that is broadly expressed during CNS development through a lineage-specific transcription factor that together with the kernel gene creates cis-regulatory input that contributes to transcriptionally activate a battery of terminal differentiation genes.

The Drosophila LIM-only (LMO) protein DLMO functions as a negative regulator of transcription during development of the fly wing. Here we report a novel role of DLMO as a positive regulator of transcription during the development of thoracic sensory bristles. We isolated new dlmo mutants, which lack some thoracic dorsocentral (DC) bristles. This phenotype is typical of malfunction of a thoracic multiprotein transcription complex, composed of CHIP, PANNIER (PNR), ACHAETE (AC), and DAUGHTERLESS (DA). Genetic interactions reveal that dlmo synergizes with pnr and ac to promote the development of thoracic DC bristles. Moreover, loss-of-function of dlmo reduces the expression of a reporter target gene of this complex in vivo. Using the GAL4-UAS system we also show that dlmo is spatially expressed where this complex is known to be active. Glutathione-S-transferase (GST)-pulldown assays showed that DLMO can physically bind CHIP and PNR through either of the two LIM domains of DLMO, suggesting that DLMO might function as part of this transcription complex in vivo. We propose that DLMO exerts its positive effect on DC bristle development by serving as a bridging molecule between components of the thoracic transcription complex.

The COP9 signalosome (CSN), an eight-subunit protein complex, is conserved in all higher eukaryotes. CSN intersects the ubiquitin–proteasome pathway, modulating signaling pathways controlling various aspects of development. We are using Drosophila as a model system to elucidate the function of this important complex. Transcriptome data were generated for four csn mutants, sampled at three developmental time points. Our results are highly reproducible, being confirmed using two different experimental setups that entail different microarrays and different controls. Our results indicate that the CSN acts as a transcriptional repressor during development of Drosophila, resulting in achronic gene expression in the csn mutants. ‘Time shift' analysis with the publicly available Drosophila transcriptome data indicates that genes repressed by the CSN are normally induced primarily during late embryogenesis or during metamorphosis. These temporal shifts are likely due to the roles of the CSN in regulating transcription factors. A null mutation in CSN subunit 4 and hypomorphic mutations in csn5 lead to more severe defects than seen in the csn5-null mutants strain, suggesting that CSN5 carries only some of the CSN function.

Background In Gaucher disease (GD), resulting from mutations in the GBA gene, mutant β-glucocerebrosidase (GCase) molecules are recognized as misfolded in the endoplasmic reticulum (ER). They are retrotranslocated to the cytoplasm, where they are ubiquitinated and undergo proteasomal degradation in a process known as the ER Associated Degradation (ERAD). We have shown in the past that the degree of ERAD of mutant GCase correlates with GD severity. Persistent presence of mutant, misfolded protein molecules in the ER leads to ER stress and evokes the unfolded protein response (UPR). Methods We investigated the presence of UPR in several GD models, using molecular and behavioral assays. Results Our results show the existence of UPR in skin fibroblasts from GD patients and carriers of GD mutations. We could recapitulate UPR in two different Drosophila models for carriers of GD mutations: flies heterozygous for the endogenous mutant GBA orthologs and flies expressing the human N370S or L444P mutant GCase variants. We encountered early death in both fly models, indicating the deleterious effect of mutant GCase during development. The double heterozygous flies, and the transgenic flies, expressing mutant GCase in dopaminergic/serotonergic cells developed locomotion deficit. Conclusion Our results strongly suggest that mutant GCase induces the UPR in GD patients as well as in carriers of GD mutations and leads to development of locomotion deficit in flies heterozygous for GD mutations.