Carbon dot/polylactic acid nanofibrous membranes for solar-mediated oil absorption/separation: Performance, environmental sustainability, ecotoxicity and reusability.

Carbon dots (CDs) are promising photothermal nanoparticles that can be utilized in environmental treatments. They exhibit favorable physicochemical properties, including low toxicity, physical and chemical stability, photo-dependant reversible behaviour, and environmentally friendly synthesis using benign building blocks. Here, we synthesized innovative CDs/polylactic acid (PLA) electrospun composite membranes for evaluating the removal of hydrophobic compounds like long-chain hydrocarbons or oils in biphasic mixtures with water. The ultimate goal was to develop innovative and sustainable solar-heated oil absorbents. Specifically, we fabricated PLA membranes with varying CD contents, characterized their morphology, thermal, and mechanical properties, and assessed the environmental impact of membrane production according to ISO 14040 and 14044 standards in a preliminary "cradle-to-gate" life cycle assessment study. Solar radiation experiments demonstrated that the CDs/PLA composites exhibited greater uptake of hydrophobic compounds compared to pure PLA membranes, ascribable to the CDs-induced photothermal effect. The adsorption and regeneration capacity of the new CDs/PLA membrane was demonstrated through multiple uptake/release cycles. Ecotoxicity analyses confirmed the safety profile of the new adsorbent system towards freshwater microalgae, further emphasizing its potential as an environmentally friendly solution for the removal of hydrophobic compounds in water treatment processes.

Self-assembly at the interface of λ-carrageenan and amphiphilic and cationic peptides: More than meets the eye.

Self-assembly of macroscopic membranes at the interface between self-assembling peptides and aqueous polymer solutions of opposite charge has been explored mostly due to the membranes' unique hierarchical structure of three distinct regions, including a layer of perpendicular fibers. We report here on the formation and characterization of self-assembled membranes made with λ-carrageenan and the cationic ß-sheet peptides, Pro-Lys-(Phe-Lys)5-Pro (PFK). Using SAXS, SEM, ITC, and rheology, we compared these membranes' morphology and physical properties to membranes made with alginate. We recognized that the polysaccharide's single chain conformation, its solution's viscosity, the potential of hydrogen bonding and electrostatic interactions between the polysaccharides and the peptides charged groups, and the strength of these interactions all affect the properties of the resulting membranes. As a result, we identified that an interplay between the polymer-peptide strength of interactions and the stiffness of the polysaccharide's single chain could be used as a route to control the structure-function relationship of the membranes. These results provide valuable information for creating guidelines to design self-assembly membranes with specific properties.

Real-time monitoring of phase transitions in π-SnS nanoparticles.

While the new cubic phase of tin monosulfide, π-SnS, shows potential for various applications, not much work was focused on the phase transitions, thermal stability, and thermal properties of π-SnS. In this work, we addressed these issues using temperature-resolved in situ X-ray diffraction combined with thermo-gravimetric differential scanning calorimetry and thermo-gravimetric infrared spectroscopy. The cubic π-SnS phase nanoparticles capped with polyvinylpyrrolidone were proven stable for 12 hours at 400 °C, pointing out the possible utilization of this new cubic phase at elevated temperatures. At the same time, heating above this temperature resulted in a phase transition to the high-temperature orthorhombic ß-SnS phase. Subsequent cooling to room temperature led to an additional phase transition to the stable orthorhombic α-SnS phase. Interestingly, heating-induced phase transformation of π-SnS nanoparticles always resulted in ß-SnS, even at temperatures below the α- to ß-SnS equilibrium transition temperature. It was shown that surfactant decomposition and evaporation triggers the phase transition. Several thermal parameters were calculated, including the phase transition activation energy and the thermal expansion of the unit cell parameter of π-SnS.

Scavenging neurotoxic aldehydes using lysine carbon dots.

Reactive aldehydes generated in cells and tissues are associated with adverse physiological effects. Dihydroxyphenylacetaldehyde (DOPAL), the biogenic aldehyde enzymatically produced from dopamine, is cytotoxic, generates reactive oxygen species, and triggers aggregation of proteins such as α-synuclein implicated in Parkinson's disease. Here, we demonstrate that carbon dots (C-dots) prepared from lysine as the carbonaceous precursor bind DOPAL molecules through interactions between the aldehyde units and amine residues on the C-dot surface. A set of biophysical and in vitro experiments attests to attenuation of the adverse biological activity of DOPAL. In particular, we show that the lysine-C-dots inhibit DOPAL-induced α-synuclein oligomerization and cytotoxicity. This work underlines the potential of lysine-C-dots as an effective therapeutic vehicle for aldehyde scavenging.

Hydrothermal carbonization reaction severity as an indicator of human-excreta-derived hydrochar properties and it's combustion.

Hydrothermal carbonization (HTC) is an emerging technology that may potentially address sanitation problems and energy scarcity. However, the significance of the parameters that govern HTC (e.g., temperature and time) is not fully understood, in particular for human excreta. A simplified coalification model was used to describe the 'strength' of thermal reactions by combining temperature and time into a single parameter, the severity factor. This study is the first to assess the extent to which a severity coalification model can predict the properties of human-excreta-derived hydrochar for a given severity with different combinations of reaction time and temperature. HTC experiments with raw human excreta were undertaken with 50 mL batch reactors at five different severities. Severity was established with different combinations of temperature (180 °C, 210 °C, and 240 °C) and reaction time based on the severity-factor equation. The resulting hydrochars were tested for combustion properties, and the respective gas emission as well as, physicochemical and surface area parameters. Significant correlations were found between severity and yield (R2 = 0.88), carbon content (R2 = 0.85), and calorific value (R2 = 0.90), with the properties being similar for a given severity but varying with different severities. Hydrochar's contact angle increased from 53.1° to 81.3° with increasing SF, while surface area remained low, ranging from <1 to 5.1 m2g-1, with no definite correlation to SF. Combustion profiles for a given severity were generally similar, but the ignition, peak, and burnout temperatures differed between severities. Gram-Schmidt curves indicated that gas emission profiles are similar for a given severity but vary with different severities. The main gases emitted in combustion were virtually identical in all treatments, and included CO2, alkenes (C9, C10), CH4, and H2O. It is concluded that many properties of hydrochar can be inferred from the severity factor.

High-Resolution Cryo-Electron Microscopy Reveals the Unique Striated Hollow Structure of Photocatalytic Macrocyclic Polydiacetylene Nanotubes.

High-resolution structures are crucial for understanding the functional properties of nanomaterials. We applied single-particle cryo-electron microscopy (cryo-EM), a method traditionally used for structure determination of biological macromolecules, to obtain high-resolution structures of synthetic non-biological filaments formed by photopolymerization of macrocyclic diacetylene (MDA) amphiphilic monomers. Tomographic analysis showed that the MDA monomers self-assemble into hollow nanotubes upon dispersion in water. Single-particle analysis revealed tubes consisting of six pairs of covalently bonded filaments held together by hydrophobic interactions, where each filament is composed of macrocyclic rings stacked in parallel "chair" conformations. The hollow MDA nanotube structures we found may account for the efficient scavenging of amphiphilic pollutants in water and subsequent photodegradation of the guest species.

Native Glucagon Amyloids Catalyze Key Metabolic Reactions.

Glucagon is a prominent peptide hormone, playing central roles in the regulation of glucose blood-level and lipid metabolism. Formation of glucagon amyloid fibrils has been previously reported, although no biological functions of such fibrils are known. Here, we demonstrate that glucagon amyloid fibrils catalyze biologically important reactions, including esterolysis, lipid hydrolysis, and dephosphorylation. In particular, we found that glucagon fibrils catalyze dephosphorylation of adenosine triphosphate (ATP), a core metabolic reaction in cell biology. Comparative analysis of several glucagon variants allowed mapping the catalytic activity to an enzymatic pocket-like triad formed at the glucagon fibril surface, comprising the histidyl-serine domain at the N-terminus of the peptide. This study may point to previously unknown physiological roles and pathological consequences of glucagon fibrillation and supports the hypothesis that catalytic activities of native amyloid fibrils play functional roles in human physiology and disease.

Inhibition of Staphylococcus aureus biofilm-forming functional amyloid by molecular tweezers.

Biofilms are rigid and largely impenetrable three-dimensional matrices constituting virulence determinants of various pathogenic bacteria. Here, we demonstrate that molecular tweezers, unique supramolecular artificial receptors, modulate biofilm formation of Staphylococcus aureus. In particular, the tweezers affect the structural and assembly properties of phenol-soluble modulin α1 (PSMα1), a biofilm-scaffolding functional amyloid peptide secreted by S. aureus. The data reveal that CLR01, a diphosphate tweezer, exhibits significant S. aureus biofilm inhibition and disrupts PSMα1 self-assembly and fibrillation, likely through inclusion of lysine side chains of the peptide. In comparison, different peptide binding occurs in the case of CLR05, a tweezer containing methylenecarboxylate units, which exhibits lower affinity for the lysine residues yet disrupts S. aureus biofilm more strongly than CLR01. Our study points to a possible role for molecular tweezers as potent biofilm inhibitors and antibacterial agents, particularly against untreatable biofilm-forming and PSM-producing bacteria, such as methicillin-resistant S. aureus.

Mitochondria membrane transformations in colon and prostate cancer and their biological implications.

Mitochondria have emerged as important determinants in cancer progression and malignancy. However, the role of mitochondrial membranes in cancer onset and progression has not been thoroughly investigated. This study compares the structural and functional properties of mitochondrial membranes in prostate and colon cancer cells in comparison to normal mitochondria, and possible therapeutic implications of these membrane changes. Specifically, isolation of cell mitochondria and preparation of inverted sub-mitochondrial particles (SMPs) illuminated significant cancer-induced modulations of membrane lipid compositions, fluidity, and activity of cytochrome c oxidase, one of the key mitochondrial enzymes. The experimental data further show that cancer-associated membrane transformations may account for mitochondria targeting by betulinic acid and resveratrol, known anti-cancer molecules. Overall, this study probes the relationship between cancer and mitochondrial membrane transformations, underlying a potential therapeutic significance for mitochondrial membrane targeting in cancer.

The cation diffusion facilitator protein MamM's cytoplasmic domain exhibits metal-type dependent binding modes and discriminates against Mn2.

Cation diffusion facilitator (CDF) proteins are a conserved family of divalent transition metal cation transporters. CDF proteins are usually composed of two domains: the transmembrane domain, in which the metal cations are transported through, and a regulatory cytoplasmic C-terminal domain (CTD). Each CDF protein transports either one specific metal or multiple metals from the cytoplasm, and it is not known whether the CTD takes an active regulatory role in metal recognition and discrimination during cation transport. Here, the model CDF protein MamM, an iron transporter from magnetotactic bacteria, was used to probe the role of the CTD in metal recognition and selectivity. Using a combination of biophysical and structural approaches, the binding of different metals to MamM CTD was characterized. Results reveal that different metals bind distinctively to MamM CTD in terms of their binding sites, thermodynamics, and binding-dependent conformations, both in crystal form and in solution, which suggests a varying level of functional discrimination between CDF domains. Furthermore, these results provide the first direct evidence that CDF CTDs play a role in metal selectivity. We demonstrate that MamM's CTD can discriminate against Mn2+, supporting its postulated role in preventing magnetite formation poisoning in magnetotactic bacteria via Mn2+ incorporation.

The metal binding site composition of the cation diffusion facilitator protein MamM cytoplasmic domain impacts its metal responsivity.

The cation diffusion facilitator (CDF) is a conserved family of divalent d-block metal cation transporters that extrude these cations selectively from the cytoplasm. CDF proteins are composed of two domains: the transmembrane domain, through which the cations are transported, and a regulatory cytoplasmic C-terminal domain (CTD). It was recently shown that the CTD of the CDF protein MamM from magnetotactic bacteria has a role in metal selectivity, as binding of different metal cations exhibits distinctive affinities and conformations. It is yet unclear whether the composition of the CTD binding sites can impact metal selectivity and if we can manipulate the CTD to response to other non-native metals in CDF proteins. Here we performed a mutational study of the model protein MamM CTD, where we exchanged the native metal binding residues with different metal-binding amino acids. Using X-ray crystallography and Trp-fluorescence spectrometry, we studied the impact of these mutations on the CTD conformation in the presence of non-native metals. Our results reveal that the incorporation of such mutations alters the domain response to metals in vitro, as mutant forms of the CTD bind metals differently in terms of the composition of the binding sites and the CTD conformation. Therefore, the results demonstrate the direct influence of the CTD binding site composition on CDF proteins structure and hence, function, and constitute a first step for rational design of MamM for transporting different metals in vivo.

Diphenylalanine-Derivative Peptide Assemblies with Increased Aromaticity Exhibit Metal-like Rigidity and High Piezoelectricity.

Diphenylalanine (FF) represents the simplest peptide building block that self-assembles into ordered nanostructures with interesting physical properties. Among self-assembled peptide structures, FF nanotubes display notable stiffness and piezoelectric parameters (Young's modulus = 19-27 GPa, strain coefficient d33 = 18 pC/N). Yet, inorganic alternatives remain the major materials of choice for many applications due to higher stiffness and piezoelectricity. Here, aiming to broaden the applications of the FF motif in materials chemistry, we designed three phenyl-rich dipeptides based on the ß,ß-diphenyl-Ala-OH (Dip) unit: Dip-Dip, cyclo-Dip-Dip, and tert-butyloxycarbonyl (Boc)-Dip-Dip. The doubled number of aromatic groups per unit, compared to FF, produced a dense aromatic zipper network with a dramatically improved Young's modulus of ∼70 GPa, which is comparable to aluminum. The piezoelectric strain coefficient d33 of ∼73 pC/N of such assembly exceeds that of poled polyvinylidene-fluoride (PVDF) polymers and compares well to that of lead zirconium titanate (PZT) thin films and ribbons. The rationally designed π-π assemblies show a voltage coefficient of 2-3 Vm/N, an order of magnitude higher than PVDF, improved thermal stability up to 360 °C (∼60 °C higher than FF), and useful photoluminescence with wide-range excitation-dependent emission in the visible region. Our data demonstrate that aromatic groups improve the rigidity and piezoelectricity of organic self-assembled materials for numerous applications.

Peptide Self-Assembly Is Linked to Antibacterial, but Not Antifungal, Activity of Histatin 5 Derivatives.

The rise of multidrug-resistant pathogens has awakened interest in new drug candidates such as antimicrobial peptides and their derivatives. Recent work suggests that some antimicrobial peptides have the ability to self-assemble into ordered amyloid-like nanostructures which facilitate their antibacterial activity. Here, we evaluate a histatin-based antimicrobial peptide, and its self-assembling derivative, in the interplay between self-assembly, membrane interactions, and antibacterial and antifungal activities. We demonstrate substantial membrane targeting by both peptides, as well as mechanistic insights into this mode of action, which correlates to their antifungal activity and is not affected by their self-assembling state. The ability to self-assemble does, however, significantly affect peptide antibacterial activity against both Gram-negative and Gram-positive bacteria. These results are surprising and hint at important distinctions between antifungal and antibacterial peptide activities in prokaryotes and eukaryotic microbes.IMPORTANCE Antimicrobial peptides are important modulators of host defense against bacterial, fungal, and viral pathogens in humans and other multicellular organisms. Two converging paradigms point to a link between antimicrobial peptides that self-assemble into amyloid-like nanoassemblies and classical amyloidogenic peptides that often have potent broad-spectrum antimicrobial activity, suggesting that antimicrobial and amyloidogenic peptides may represent two sides of the same coin. Here, we asked if the ability of an antifungal peptide to self-assemble affects its antifungal or antibacterial activity. We found that modifications of classical antifungal peptide derivative allowed it to self-assemble and did not alter its antifungal activity, and yet self-assembly substantially increased the antibacterial activity of the peptide. These results support the idea that peptide self-assembly can enhance antibacterial activities and emphasize a distinction between the action of antifungal peptides and that of antibacterial peptides. Accordingly, we suggest that the possible generality of this distinction should be widely tested.

Imaging Flow Cytometry Illuminates New Dimensions of Amyloid Peptide-Membrane Interactions.

Membrane interactions of amyloidogenic proteins constitute central determinants both in protein aggregation as well as in amyloid cytotoxicity. Most reported studies of amyloid peptide-membrane interactions have employed model membrane systems combined with application of spectroscopy methods or microscopy analysis of individual binding events. Here, we applied for the first time, to our knowledge, imaging flow cytometry for investigating interactions of representative amyloidogenic peptides, namely, the 106-126 fragment of prion protein (PrP(106-126)) and the human islet amyloid polypeptide (hIAPP), with giant lipid vesicles. Imaging flow cytometry was also applied to examine the inhibition of PrP(106-126)-membrane interactions by epigallocatechin gallate, a known modulator of amyloid peptide aggregation. We show that imaging flow cytometry provided comprehensive population-based statistical information upon morphology changes of the vesicles induced by PrP(106-126) and hIAPP. Specifically, the experiments reveal that both PrP(106-126) and hIAPP induced dramatic transformations of the vesicles, specifically disruption of the spherical shapes, reduction of vesicle circularity, lobe formation, and modulation of vesicle compactness. Interesting differences, however, were apparent between the impact of the two peptides upon the model membranes. The morphology analysis also showed that epigallocatechin gallate ameliorated vesicle disruption by PrP(106-126). Overall, this study demonstrates that imaging flow cytometry provides powerful means for disclosing population-based morphological membrane transformations induced by amyloidogenic peptides and their inhibition by aggregation modulators.

Tyrosine carbon dots inhibit fibrillation and toxicity of the human islet amyloid polypeptide.

Misfolding and aggregation of the human islet amyloid polypeptide (hIAPP) are believed to play key roles in the pathophysiology of type-II diabetes. Here, we demonstrate that carbon dots (C-dots) prepared from the amino acid tyrosine inhibit fibrillation of hIAPP, reduce hIAPP-induced cell toxicity and block membrane disruption by the peptide. The pronounced inhibitory effect is traced to the display of ubiquitous aromatic residues upon the C-dots' surface, mimicking the anti-fibril and anti-toxic activity of natural polyphenolic compounds. Notably, spectroscopy and thermodynamics analysis demonstrated different hIAPP interactions and fibril inhibition effects induced by tyrosine-C-dots displaying phenolic residues and C-dots prepared from phenylalanine which exhibited phenyl units on their surface, underscoring the significance of hydrogen bonding mediated by the phenolic hydroxide moieties for the fibril modulation activity. The presented experiments attest to the potential of tyrosine-C-dots as a therapeutic vehicle for protein misfolding diseases, interfering in both π-π interactions as well as hydrogen bonding involving aromatic residues of amyloidogenic peptides.

Ultrashort Cell-Penetrating Peptides for Enhanced Sonophoresis-Mediated Transdermal Transport.

The skin is a key site for drug administration because of its large surface area and noninvasive accessibility. However, the dermal architecture serves as an excellent barrier, protecting from external mechanical, chemical, microbial, and physical perturbations. Most drugs display poor permeability through this barrier, thus making dermal and subdermal delivery challenging. Cell-penetrating peptides (CPPs), a diverse group of relatively short cationic and amphipathic membrane-interacting peptides, are fast becoming an important class of drug carriers and could potentially be developed for the dermal delivery of active molecules. However, the mechanism of CPP transdermal delivery is not fully understood, and there is a genuine need for a minimal model to understand this important phenomenon. Here, we demonstrate the potent membrane interactions of a minimal four-amino-acid-long CPP as well as the significance of guanidinium patterning and cationic nature of this palindromic peptide on its bioactivity. Furthermore, we demonstrate the biocompatibility of this peptide as well as its rapid cellular uptake and endosomal distribution. Finally, by utilizing a porcine full-thickness skin model, we demonstrate the substantial independent dermal and sonophoresis-based transdermal penetration of this minimal model. These results provide a minimal model for CPPs which can be easily manipulated for further biophysical and biochemical evaluations as well as a potent functional CPP with excellent skin permeability, which can be utilized for a wide variety of cosmetic and medical applications.

Mechanically rigid supramolecular assemblies formed from an Fmoc-guanine conjugated peptide nucleic acid.

The variety and complexity of DNA-based structures make them attractive candidates for nanotechnology, yet insufficient stability and mechanical rigidity, compared to polyamide-based molecules, limit their application. Here, we combine the advantages of polyamide materials and the structural patterns inspired by nucleic-acids to generate a mechanically rigid fluorenylmethyloxycarbonyl (Fmoc)-guanine peptide nucleic acid (PNA) conjugate with diverse morphology and photoluminescent properties. The assembly possesses a unique atomic structure, with each guanine head of one molecule hydrogen bonded to the Fmoc carbonyl tail of another molecule, generating a non-planar cyclic quartet arrangement. This structure exhibits an average stiffness of 69.6 ± 6.8 N m-1 and Young's modulus of 17.8 ± 2.5 GPa, higher than any previously reported nucleic acid derived structure. This data suggests that the unique cation-free "basket" formed by the Fmoc-G-PNA conjugate can serve as an attractive component for the design of new materials based on PNA self-assembly for nanotechnology applications.

Real-Time In-Situ Monitoring of a Tunable Pentapeptide Gel-Crystal Transition.

Supramolecular gels often become destabilized by the transition of the gelator into a more stable crystalline phase, but often the long timescale and sporadic localization of the crystalline phase preclude a persistent observation of this process. We present a pentapeptide gel-crystal phase transition amenable for continuous visualization and quantification by common microscopic methods, allowing the extraction of kinetics and visualization of the dynamics of the transition. Using optical microscopy and microrheology, we show that the transition is a sporadic event in which gel dissolution is associated with microcrystalline growth that follows a sigmoidal rate profile. The two phases are based on ß-sheets of similar yet distinct configuration. We also demonstrate that the transition kinetics and crystal morphology can be modulated by extrinsic factors, including temperature, solvent composition, and mechanical perturbation. This work introduces an accessible model system and methodology for studying phase transitions in supramolecular gels.

Structure and membrane-targeting of a Bordetella pertussis effector N-terminal domain.

BteA, a 69-kDa cytotoxic protein, is a type III secretion system (T3SS) effector in the classical Bordetella, the etiological agents of pertussis and related mammalian respiratory diseases. Like other cytotoxicity-mediating effectors, BteA uses its multifunctional N-terminal domain to target phosphatidylinositol (PI)-rich microdomains in the host membrane. Despite their structural similarity, T3SS effectors exhibit a variable range of membrane interaction modes, and currently only limited structural information is available for the BteA membrane-targeting domain and the molecular mechanisms underlying its function. Employing a synergistic combination of structural methods, here we determine the structure of this functional domain and uncover key molecular determinants mediating its interaction with membranes. Residues 29-121 of BteA form an elongated four-helix bundle packed against two shorter perpendicular helices, the second of which caps the domain in a critical 'tip motif'. A flexible region preceding the BteA helical bundle contains the characteristic ß-motif required for binding its cognate chaperone BtcA. We show that BteA targets PI(4,5)P2-containing lipoprotein nanodiscs and binds a soluble PI(4,5)P2 analog via an extensive positively charged surface spanning its first two helices, and that this interaction is weaker for PI(3,5)P2 and abolished for PI(4)P. We confirmed this model of membrane-targeting by observation of BteA-induced changes in the structure of PI(4,5)P2-containing phospholipid bilayers using small-angle X-ray scattering (SAXS). We also extended these results to a larger BteA domain (residues 1-287), confirming its interaction with bilayers using calorimetry, fluorescence and SAXS methods. This novel view of the structural underpinnings of membrane targeting by BteA is an important step towards a comprehensive understanding of cytotoxicity in Bordetella, as well as interactions of a broad range of pathogens with their respective hosts.

Metal binding to the dynamic cytoplasmic domain of the cation diffusion facilitator (CDF) protein MamM induces a 'locked-in' configuration.

Cation diffusion facilitator (CDF) proteins are a conserved family of transmembrane transporters that ensure cellular homeostasis of divalent transition metal cations. Metal cations bind to CDF protein's cytoplasmic C-terminal domain (CTD), leading to closure from its apo open V-shaped dimer to a tighter packed structure, followed by a conformational change of the transmembrane domain, thus enabling transport of the metal cation. By implementing a comprehensive range of biochemical and biophysical methods, we studied the molecular mechanism of metal binding to the magnetotactic bacterial CDF protein MamM CTD. Our results reveal that the CTD is rather dynamic in its apo form, and that two dependent metal-binding sites, a single central binding site and two symmetrical, peripheral sites, are available for metal binding. However, only cation binding to the peripheral sites leads to conformational changes that lock the protein in a compact state. Thus, this work reveals how metal binding is regulating the sequential uptakes of metal cations by MamM, and extends our understanding of the complex regulation mechanism of CDF proteins. DATABASE: Structural data are available in RCSB Protein Data Bank under the accession numbers: 6G64, 6G55, 6G5E and 6G6I (for CS, C267S, CS-C267S and W247A, respectively).