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Confirmation of extracellular amyloid deposition across various animal species and tissue types has been a long-standing challenge in veterinary diagnostic pathology. Transmission electron microscopy (TEM) has historically been used to advance the understanding of amyloid fibril morphology and confirm amyloid fibril deposition when histologic methods provide unclear results. We assessed the feasibility of utilizing TEM for routine confirmation of amyloidosis as an addition to histology. We analyzed ex situ amyloid fibrils with direct, negative-contrast TEM and in situ amyloid fibrils with aldehyde-fixed, plastic-embedding TEM to confirm amyloidosis in a variety of cases in which amorphous extracellular amyloid deposits had been identified by H&E and Congo red staining. We compared the 2 TEM methods and documented amyloid fibril morphology and morphometry in 7 species (goat, guinea pig, kudu, fox, sheep, flamingo, and duck). Ex situ fibrils had helical morphology and widths of 15-18 nm across all species. Fibril crossover distances had more interspecies variation of 60-130 nm, and species could be grouped based on pitch (twist size). Twisting patterns of in situ fibrils could not be visualized, but in situ widths of 10 nm were measured across all species. In 4 different chicken cases, fibrils differing morphologically from amyloid were consistently detected via both TEM methods, suggesting the possibility of a non-amyloid deposit that is commonly diagnosed as amyloidosis based on its histologic appearance. When available, we recommend routine confirmation of amyloid fibril deposition by TEM.

Protein misfolding, aggregation, and aberrant aggregate accumulation play a central role in neurodegenerative disease progression. The proteotoxic factors also govern the aging process to a large extent. Molecular chaperones modulate proteostasis and thereby impact aberrant-protein-induced proteotoxicity. These chaperones have a diverse functional spectrum, including nascent protein folding, misfolded protein sequestration, refolding, or degradation. Small heat shock proteins (sHsps) possess an ATP-independent chaperone-like activity that prevents protein aggregation by keeping target proteins in a folding-competent state to be refolded by ATP-dependent chaperones. Due to their near-universal upregulation and presence in sites of proteotoxic stress like diseased brains, sHsps were considered pathological. However, gene knockdown and overexpression studies have established their protective functions. This review provides an updated overview of the sHsp role in protein aggregation amelioration and highlights evidence for sHsp modulation of neurodegenerative disease-related protein aggregation and aging. [ABSTRACT FROM AUTHOR]

The effect of sodium alginate on the denaturation and aggregation behavior of bovine serum albumin and hen egg-white lysozyme was studied. Large amounts of polysaccharide increase the thermal stability of albumin due to the weak binding interactions. At the same time, sodium alginate can reduce the quantity of amyloid fibrils formed by albumin under denaturing conditions, which is a consequence of the stabilization of the native protein form by glycan binding. In the case of lysozyme, the polysaccharide has no influence on the thermal stability of the protein in 2 M guanidinium hydrochloride. However, the inhibition of fibril formation with an increase in the lag time was observed, which is explained by the binding of sodium alginate to lysozyme fibrils, but not to the protein monomer. The molecular nature of the binding interactions between alginate and the studied proteins was elucidated using molecular docking and known experimental structures of glycan–protein complexes.

The effect of sodium alginate on the denaturation and aggregation behavior of bovine serum albumin and hen egg-white lysozyme was studied. Large amounts of polysaccharide increase the thermal stability of albumin due to the weak binding interactions. At the same time, sodium alginate can reduce the quantity of amyloid fibrils formed by albumin under denaturing conditions, which is a consequence of the stabilization of the native protein form by glycan binding. In the case of lysozyme, the polysaccharide has no influence on the thermal stability of the protein in 2 M guanidinium hydrochloride. However, the inhibition of fibril formation with an increase in the lag time was observed, which is explained by the binding of sodium alginate to lysozyme fibrils, but not to the protein monomer. The molecular nature of the binding interactions between alginate and the studied proteins was elucidated using molecular docking and known experimental structures of glycan–protein complexes. [ABSTRACT FROM AUTHOR]

Amyloid fibrils from Alzheimer's amyloid-beta peptides (Aβ) are found to be polymorphic. So far, 14 Aβ40 fibril structures have been determined. The mechanism of why one particular protein sequence adopts so many different three-dimensional structures is yet not understood. In this work, we describe the assignment of the NMR chemical shifts of two Alzheimer's disease fibril polymorphs, P1 and P2, which are formed by the amyloid-beta peptide Aβ40. The assignment is based on 13C-detected 3D NCACX and NCOCX experiments MAS solid-state NMR experiments. The fibril samples are prepared using an extensive seeding protocol in the absence and presence of the small heat shock protein αB-crystallin. In addition to manual assignments, we obtain chemical shift assignments using the automation software ARTINA. We present an analysis of the secondary chemical shifts and a discussion on the differences between the manual and automated assignment strategies. [ABSTRACT FROM AUTHOR]

The α-synuclein (α-syn) amyloid fibrils are involved in various neurogenerative diseases. Solid-state NMR (ssNMR) has been showed as a powerful tool to study α-syn aggregates. Here, we report the 1H, 13C and 15N back-bone chemical shifts of a new α-syn polymorph obtained using proton-detected ssNMR spectroscopy under fast (95 kHz) magic-angle spinning conditions. The manual chemical shift assignments were cross-validated using FLYA algorithm. The secondary structural elements of α-syn fibrils were calculated using 13C chemical shift differences and TALOS software. [ABSTRACT FROM AUTHOR]

[Display omitted] • Modular bioengineered nanofibrils neutralize SARS-CoV-2 virus-like particles. • Recombinant protein subunits spontaneously assemble into functional fibrils. • The assembled amyloid-like nanofibrils are biocompatible. • Mesoscopic structure and activity of the protein assembly are easily modulated. • Surfaces coated with nanofibrils capture SARS-CoV-2 Spike in wet environments. The recent coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spurred intense research efforts to develop new materials with antiviral activity. In this study, we genetically engineered amyloid-based nanofibrils for capturing and neutralizing SARS-CoV-2. Building upon the amyloid properties of a short Sup35 yeast prion sequence, we fused it to SARS-CoV-2 receptor-binding domain (RBD) capturing proteins, LCB1 and LCB3. By tuning the reaction conditions, we achieved the spontaneous self-assembly of the Sup35-LCB1 fusion protein into a highly homogeneous and well-dispersed amyloid-like fibrillar material. These nanofibrils exhibited high affinity for the SARS-CoV-2 RBD, effectively inhibiting its interaction with the angiotensin-converting enzyme 2 (ACE2) receptor, the primary entry point for the virus into host cells. We further demonstrate that this functional nanomaterial entraps and neutralizes SARS-CoV-2 virus-like particles (VLPs), with a potency comparable to that of therapeutic antibodies. As a proof of concept, we successfully fabricated patterned surfaces that selectively capture SARS-CoV-2 RBD protein on wet environments. Collectively, these findings suggest that these protein-only nanofibrils hold promise as disinfecting coatings endowed with selective SARS-CoV-2 neutralizing properties to combat viral spread or in the development of sensitive viral sampling and diagnostic tools. [ABSTRACT FROM AUTHOR]

It is assumed that the stability and bioaccessibility of iron ions in iron–pea protein fibril (Fe-Fib PP) nanocomposite can be remarkably enhanced, and Fe-Fib PP exhibits great potential as an effective iron fortificant. Fe-Fib PP, a stable and effective iron supplement, was fabricated based on the reducing property of pea protein fibrils, derived from pea protein through thermal treatment at pH 2.0. The results demonstrated that the reducing power of iron was remarkably affected by fibril concentration and fibrillization degree. The reducing power of pea protein fibrils gradually enhanced from 0.31 to 0.92 with the increase in incubation time from 0 to 48 h. Compared with iron nanoparticles (Fe–Nano), Fe-Fib PP possessed much higher dispersibility. Additionally, the stability of iron in Fe-Fib PP was significantly higher than that in Fe–Nano under different storage conditions. X-ray photoelectron spectroscopy (XPS) outcomes revealed Fe (II) content in Fe-Fib PP (70.75 ± 0.65%) was remarkably higher than that of Fe–Nano (56.05 ± 0.50%). In addition, the bioaccessibility of Fe (II) dramatically improved from 42.7% to 62.8% using PP fibrils as carriers. The findings suggest that Fe-Fib PP is an effective iron nutrition enhancer. [ABSTRACT FROM AUTHOR]

Background/Objectivities: The presence of beta-amyloid plaques is a part of the pathogenesis of Alzheimer's disease, but there is currently no universally accepted method for magnetic resonance (MR) imaging of the disease. However, it is known that magnetic nanoparticles (MNPs) can improve the T2 contrast in MR images of various targets. Methods: We used cubic MNPs, which were produced by thermal decomposition and then it was covalently bonded to a modified fluorescently labeled tetrapeptide, HAEE-Cy5, for visualizing beta-amyloid plaques. The interaction of MNPs-HAEE-Cy5 and beta-amyloid was determinate by confocal microscopy using SH-SY5Y cell line. Results: MNPs exhibit relatively high relaxivity (approximately 200 mM−1s−1), which is crucial for enhancing target visibility in MR imaging. HAEE provides targeted delivery of MNPs by specifically interacting with beta-amyloid, while the fluorescent label Cy5 enables monitoring the efficacy of the interaction through confocal microscopy. Conclusions: The MNPs modified with HAEE-Cy5 demonstrated excellent binding to beta-amyloid plaques in vitro, as shown by experiments on the SH-SY5Y cell line. These results suggest that the proposed method has potential for use in future MR imaging studies of Alzheimer's disease. [ABSTRACT FROM AUTHOR]

Background/Objectives: Antibody light chains form amyloid fibrils that lead to progressive tissue damage in amyloid light chain (AL) amyloidosis. The properties of each patient's unique light chain appear to determine its propensity to form amyloid. One factor is N-glycosylation, which is more frequent in amyloid-associated light chains than in light chains from the normal immune repertoire. However, the mechanisms underlying this association are unknown. Here, we investigate the frequency and position within the light chain sequence of the N-glycosylation sequence motif, or sequon. Methods: Monoclonal light chains from AL amyloidosis and multiple myeloma were identified from the AL-Base repository. Polyclonal light chains were obtained from the Observed Antibody Space resource. We compared the fraction of light chains from each group harboring an N-glycosylation sequon, and the positions of these sequons within the sequences. Results: Sequons are enriched among AL-associated light chains derived from a subset of precursor germline genes. Sequons are observed at multiple positions, which differ between the two types of light chains, κ and λ, but are similar between light chains from AL amyloidosis and multiple myeloma. Positions of sequons map to residues with surface-exposed sidechains that are compatible with the folded structures of light chains. Within the known structures of λ AL amyloid fibrils, many residues where sequons are observed are buried, inconsistent with N-glycosylation. Conclusions: There is no clear structural rationale for why N-glycosylation of κ light chains is associated with AL amyloidosis. A better understanding of the roles of N-glycosylation in AL amyloidosis is required before it can be used as a marker for disease risk. [ABSTRACT FROM AUTHOR]

Amyloid‐beta aggregation is considered one of the factors influencing the onset of the Alzheimer's disease. Early prevention of such aggregation should alleviate disease condition by applying small molecule compounds that shift the aggregation equilibrium toward the soluble form of the peptide or slow down the process. We have discovered that fluorinated benzenesulfonamides of particular structure slowed the amyloid‐beta peptide aggregation process by more than three‐fold. We synthesized a series of ortho‐para and meta‐para double‐substituted fluorinated benzenesulfonamides that inhibited the aggregation process to a variable extent yielding a detailed picture of the structure‐activity relationship. Analysis of compound chemical structure effect on aggregation in artificial cerebrospinal fluid showed the necessity to arrange the benzenesulfonamide, hydrophobic substituent, and benzoic acid in a particular way. The amyloid beta peptide aggregate fibril structures varied in cross‐sectional height depending on the applied inhibitor indicating the formation of a complex with the compound. Application of selected inhibitors increased the survivability of cells affected by the amyloid beta peptide. Such compounds may be developed as drugs against Alzheimer's disease. [ABSTRACT FROM AUTHOR]

Förster resonance energy transfer (FRET) between the membrane fluorescent probes pyrene and TDV was employed to investigate the modulation of amyloid-lipid interactions by polyphenols. The effects of various polyphenols, including quercetin, curcumin, gallic and salicylic acids, on the complexation between the amyloid fibrils derived from N-terminal fragment of apolipoprotein A-I (ApoA-IF) and insulin (InsF), and liposomes composed of phosphatidylcholine (PC) and its mixtures with cardiolipin (CL), cholesterol (Chol), or phosphatidylglycerol (PG) were investigated. The incorporation of polyphenols resulted in decreased energy transfer efficiency, indicating a significant alteration in the spatial relationship between amyloid fibrils and lipid membranes. The magnitude of this effect was found to be dependent on lipid bilayer composition, the chemical nature of the polyphenols, and the type of amyloidogenic protein. Notably, curcumin exhibited the most pronounced impact across all systems, with a particularly strong effect on ApoA-IF compared to InsF. This differential response suggests protein-specific mechanisms of interaction and highlights the potential for targeted therapeutic approaches. Our findings provide novel insights into the intricate interplay between polyphenols, amyloid fibrils, and lipid membranes, contributing to the fundamental understanding of amyloid-related pathologies and opening new avenues for the development of polyphenol-based therapeutic strategies in amyloid-associated disorders. [ABSTRACT FROM AUTHOR]

The present study employs molecular dynamics simulations to investigate the interactions between quercetin, amyloid fibrils, and POPC lipid bilayers. The results demonstrate that quercetin does not significantly affect the molecular organization of the bilayer, while IAPP fibrils induce substantial structural changes, particularly in the outer monolayer. Quercetin mitigates these effects by reducing the impact on headgroup and glycerol regions and causing a more superficial positioning of IAPP. Additionally, quercetin slightly decreases the order of sn-2 acyl chains, indicating a disordering effect. In a ternary system with POPC, quercetin, and IAPP, the reduction in the deuterium order parameter of sn-2 acyl chains is less pronounced, underscoring quercetin's protective role. Unlike IAPP, ApoAI and insulin fibrils undergo significant structural reorganization in the membrane-bound state. Quercetin attenuative effects are observed only with ApoAI, highlighting its potential as a protective agent against amyloid-induced membrane disruption. These findings provide valuable insights into the interactions between polyphenols, amyloid fibrils, and lipid membranes, contributing to the understanding of membrane-associated amyloid pathologies.

Abstract Self-assembly is a key process in living systems—from the microscopic biological level (e.g. assembly of proteins into fibrils within biomolecular condensates in a human cell) through to the macroscopic societal level (e.g. assembly of humans into common-interest communities across online social media platforms). The components in such systems (e.g. macromolecules, humans) are highly diverse, and so are the self-assembled structures that they form. However, there is no simple theory of how such structures assemble from a multi-species pool of components. Here we provide a very simple model which trades myriad chemical and human details for a transparent analysis, and yields results in good agreement with recent empirical data. It reveals a new inhibitory role for biomolecular condensates in the formation of dangerous amyloid fibrils, as well as a kinetic explanation of why so many diverse distrust movements are now emerging across social media. The nonlinear dependencies that we uncover suggest new real-world control strategies for such multi-species assembly.

Liquid-like protein condensates have recently attracted much attention due to their critical roles in biological phenomena. They typically show high fluidity and reversibility for exhibiting biological functions, while occasionally serving as sites for the formation of amyloid fibrils. To comprehend the properties of protein condensates that underlie biological function and pathogenesis, it is crucial to study them at the single-condensate level; however, this is currently challenging due to a lack of applicable methods. Here, we demonstrate that optical trapping is capable of inducing the formation of a single liquid-like condensate of α-synuclein in a spatiotemporally controlled manner. The irradiation of tightly focused near-infrared laser at an air/solution interface formed a condensate under conditions coexisting with polyethylene glycol. The fluorescent dye-labeled imaging showed that the optically induced condensate has a gradient of protein concentration from the center to the edge, suggesting that it is fabricated through optical pumping-up of the a-synuclein clusters and the expansion along the interface. Furthermore, Raman spectroscopy and thioflavin T fluorescence analysis revealed that continuous laser irradiation induces structural transition of protein molecules inside the condensate to β-sheet rich structure, ultimately leading to the condensate deformation and furthermore, the formation of amyloid fibrils. These observations indicate that optical trapping is a powerful technique for examining the microscopic mechanisms of condensate appearance and growth, and furthermore, subsequent aging leading to amyloid fibril formation. [ABSTRACT FROM AUTHOR]

We recently reported on 8 different pentameric oligothiophenes, denoted proteophenes, with different amino acid substitution patterns along the conjugated thiophene backbone. The proteophenes were forming chiral both molecular (solvent dissolved) and self‐assembled nano‐architectonic structures, the latter having approximately 5–6 times greater ICD responses. Herein, two proteophenes, HS‐84‐E−E and HS‐84‐K−K, were further investigated in terms of binding to two chiral variants of insulin amyloid fibrils. Binding curves based on fluorescence revealed strong primary binding sites for both proteophenes and more unspecific secondary binding for especially the latter. Interestingly, their induced circular dichroism (ICD) responses were similar to or stronger than for their solvent form and showed a complicated alteration upon binding suggesting that the microstructure at the binding site governs the helical structure of the ligand. Hyperspectral fluorescence microscopy and FLIM revealed more details on the molecular binding in terms of proteophene emission decay‐times. Vibrational circular dichroism (VCD) analysis showed that VCD signal of amyloid fibrils was enhanced upon binding of proteophenes, suggesting changes in the amyloid microstructures. [ABSTRACT FROM AUTHOR]

Protein aggregation correlates with many human diseases. Protein aggregates differ in structure and shape. Strategies to develop effective aggregation inhibitors that reach the clinic failed so far. Here, we developed a family of peptides targeting early aggregation stages for both amorphous and fibrillar aggregates of proteins unrelated in sequence and structure. They act on dynamic precursors before mechanistic differentiation takes place. Using peptide arrays, we first identified peptides inhibiting the amorphous aggregation of a molten globular, aggregation‐prone mutant of the Axin tumor suppressor. Optimization revealed that the peptides activity did not depend on their sequences but rather on their molecular determinants: a composition of 20–30 % flexible, 30–40 % aliphatic and 20–30 % aromatic residues, a hydrophobicity/hydrophilicity ratio close to 1, and an even distribution of residues of different nature throughout the sequence. The peptides also suppressed fibrillation of Tau, a disordered protein that forms amyloids in Alzheimer's disease, and slowed down that of Huntingtin Exon1, an amyloidogenic protein in Huntington's disease, both entirely unrelated to Axin. Our compounds thus target early stages of different aggregation mechanisms, inhibiting both amorphous and amyloid aggregation. Such cross‐mechanistic, multi‐targeting aggregation inhibitors may be lead compounds for developing drug candidates against various protein aggregation diseases. [ABSTRACT FROM AUTHOR]

[Display omitted] Carbon dots (CDs) are carbon nano materials (CNMs) that find use across several biological applications because of their water solubility, biocompatible nature, eco-friendliness, and ease of synthesis. Additionally, their physiochemical properties can be chemically tuned for further optimization towards specific applications. Here, we investigate the efficacy of C70-derived Graphene Acid Quantum Dots (GAQDs) in mitigating the transformation of soluble, monomeric Hen Egg-White Lysozyme (HEWL) to mature fibrils during its amyloidogenic trajectory. Our findings reveal that GAQDs exhibit dose-dependent inhibition of HEWL fibril formation (up to 70 % at 5 mg/mL) without affecting mitochondrial membrane potential or inducing apoptosis at the same density. Furthermore, GAQDs scavenged reactive oxygen species (ROS); achieving a 50 % reduction in ROS levels at a mere 100 µg/mL when exposed to a standard free radical generator. GAQDs were not only found to be biocompatible with a human neuroblastoma-derived SHSY-5Y cell line but also rescued the cells from rotenone-induced apoptosis. The GAQD-tolerance of SHSY-5Y cells coupled with their ability to restitute cells from rotenone-dependent apoptosis, when taken in conjunction with the biocompatibility data, indicate that GAQDs possess neuroprotective potential. The data position this class of CNMs as promising candidates for resolving aberrant cellular outputs that associate with the advent and progress of multifactorial neurodegenerative disorders including Parkinson's (PD) and Alzheimer's diseases (AD) wherein environmental causes are implicated (95 % etiology). The data suggest that GAQDs are a multifunctional carbon-based sustainable nano-platform at the intersection of nanotechnology and neuroprotection for advancing green chemistry-derived, sustainable healthcare solutions. [ABSTRACT FROM AUTHOR]

The pathological process of prion diseases implicates that the normal physiological cellular prion protein (PrPC) converts into misfolded abnormal scrapie prion (PrPSc) through post-translational modifications that increase β-sheet conformation. We recently demonstrated that HuPrP(90–231) thermal unfolding is partially irreversible and characterized by an intermediate state (β-PrPI), which has been revealed to be involved in the initial stages of PrPC fibrillation, with a seeding activity comparable to that of human infectious prions. In this study, we report the thermal unfolding characterization, in cell-mimicking conditions, of the truncated (HuPrP(90–231)) and full-length (HuPrP(23–231)) human prion protein by means of CD and NMR spectroscopy, revealing that HuPrP(90–231) thermal unfolding is characterized by two successive transitions, as in buffer solution. The amyloidogenic propensity of HuPrP(90–231) under crowded conditions has also been investigated. Our findings show that although the prion intermediate, structurally very similar to β-PrPI, forms at a lower temperature compared to when it is dissolved in buffer solution, in cell-mimicking conditions, the formation of prion fibrils requires a longer incubation time, outlining how molecular crowding influences both the equilibrium states of PrP and its kinetic pathways of folding and aggregation. [ABSTRACT FROM AUTHOR]

Cardiac amyloidosis is an infiltrative disease that causes progressive myocardial impairment secondary to amyloid fibril deposition in the extracellular space of the myocardium. Many amyloid precursors, including transthyretin protein, are known to determine cardiac damage by aggregating and precipitating in cardiac tissue. Transthyretin cardiac amyloidosis may be either caused by rare genetic mutations of the transthyretin gene in the hereditary variant, or may arise as a consequence of age-related mechanisms in the acquired form. Although it has been labeled as a rare disease, in recent years, transthyretin cardiac amyloidosis has stood out as an emerging cause of aortic stenosis, unexplained left ventricular hypertrophy and heart failure with preserved ejection fraction, particularly in the elderly. Indeed, the integration of data deriving from both in vivo imaging techniques (whose advancement in the last years has allowed to achieve an easier and more accessible non-invasive diagnosis) and forensic studies (showing a prevalence of amyloid deposition in cardiac tissue of elderly patients up to 29%) suggests that cardiac amyloidosis is a more common disease than traditionally considered. Thanks to all the improvements in non-invasive diagnostic techniques, along with the development of efficacious therapies offering improvements in survival rates, transthyretin cardiac amyloidosis has been transformed from an incurable and infrequent condition to a relatively more diffuse and treatable disease, which physicians should take into consideration in the differential diagnostic processes in daily clinical practice. [ABSTRACT FROM AUTHOR]

The effect of binding of divalent metal cations (Ca2+, Cu2+, Mg2+, Mn2+, Zn2+) on the kinetics of fibril formation of bovine α-lactalbumin at acidic conditions is considered. The kinetic parameters of the process were determined using a thioflavin T fluorescence assay. The DSC thermograms of bovine α-lactalbumin in the presence and absence of cations were recorded. The duration of the lag period correlates with the changes in the thermal stability of the molten globule of the protein in the presence of cations. The final thioflavin T fluorescence intensity after formation of the mature fibrils decreases under the influence of calcium ions which strongly bind to the monomeric protein, and increases in solutions containing copper and especially zinc. These ions seem to accelerate secondary nucleation processes and change the fibril morphology, which was confirmed by atomic force microscopy imaging. [ABSTRACT FROM AUTHOR]

Amyloid fibrils are supramolecular systems showing distinct chirality at different levels of their complex multilayered architectures. Due to the regular long‐range chiral organization, amyloid fibrils exhibit the most intense Vibrational Optical Activity (VOA) signal observed up to now, making VOA techniques: Vibrational Circular Dichroism (VCD) and Raman Optical Activity (ROA) very promising tools to explore their structures, handedness and intricate polymorphism. This concept article reviews up‐to‐date experimental studies on VOA applications to investigate amyloid fibrils highlighting its future potential in analyzing of these unique supramolecular systems, in particular in the context of biomedicine and nanotechnology. [ABSTRACT FROM AUTHOR]

Self-assembling peptide-based hydrogels have become a highly attractive scaffold for three-dimensional (3D) in vitro disease modeling as they provide a way to create tunable matrices that can resemble the extracellular matrix (ECM) of various microenvironments. Alzheimer's disease (AD) is an exceptionally complex neurodegenerative condition; however, our understanding has advanced due to the transition from two-dimensional (2D) to 3D in vitro modeling. Nonetheless, there is a current gap in knowledge regarding the role of amyloid structures, and previously developed models found long-term difficulty in creating an appropriate model involving the ECM and amyloid aggregates. In this report, we propose a multi-component self-assembling peptide-based hydrogel scaffold to mimic the amyloid-beta (β) containing microenvironment. Characterization of the amyloid-β-mimicking hydrogel (Col-HAMA-FF) reveals the formation of β-sheet structures as a result of the self-assembling properties of phenylalanine (Phe, F) through π−π stacking of the residues, thus mimicking the amyloid-β protein nanostructures. We investigated the effect of the amyloid-β-mimicking microenvironment on healthy neuronal progenitor cells (NPCs) compared to a natural-mimicking matrix (Col-HAMA). Our results demonstrated higher levels of neuroinflammation and apoptosis markers when NPCs were cultured in the amyloid-like matrix compared to a natural brain matrix. Here, we provided insights into the impact of amyloid-like structures on NPC phenotypes and behaviors. This foundational work, before progressing to more complex plaque models, provides a promising scaffold for future investigations on AD mechanisms and drug testing. In this study, we engineered two multi-component hydrogels: one to mimic the natural extracellular matrix (ECM) of the brain and one to resemble an amyloid-like microenvironment using a self-assembling peptide hydrogel. The self-assembling peptide mimics β-amyloid fibrils seen in amyloid-β protein aggregates. We report on the culture of neuronal progenitor cells within the amyloid-mimicking ECM scaffold to study the impact through marker expressions related to inflammation and DNA damage. This foundational work, before progressing to more complex plaque models, offers a promising scaffold for future investigations on AD mechanisms and drug testing. [Display omitted] [ABSTRACT FROM AUTHOR]

The spectral and luminescent properties of the styryl derivative of thioflavin T (ThT) 2-{(1E,3E)-4-[4-(dimethylamino)-2,6-dimethylphenyl]buta-1,3-dien-1-yl}-3-ethyl-1,3-benzothiazolium-3 tosylate (Th-C23) were studied. The fluorescence decay kinetics of Th-C23 incorporated into amyloid fibrils (AF) were not exponential and could be represented as a unimodal distribution of fluorophores over duration α(t). ThT and Th-C23 were shown to form an effective donor–acceptor (D–A) pair when incorporated into the AF structure. The degree of polarization of the acceptor fluorescence was high (~0.4) within the entire spectrum, despite the energy transfer, and was explained by the special orientation of D–A pairs, parallel and antiparallel. The measured fluorescence spectrum of the donor (ThT) and absorption spectrum of the acceptor (Th-C23) were used to calculate the critical energy-transfer radius R0, which could take values from 54 to 68 Å depending on the mutual orientation and quantum yield of the donor and acceptor. [ABSTRACT FROM AUTHOR]

The present study employs molecular dynamics simulations to investigate the interactions between quercetin, amyloid fibrils, and POPC lipid bilayers. The results demonstrate that quercetin does not significantly affect the molecular organization of the bilayer, while IAPP fibrils induce substantial structural changes, particularly in the outer monolayer. Quercetin mitigates these effects by reducing the impact on headgroup and glycerol regions and causing a more superficial positioning of IAPP. Additionally, quercetin slightly decreases the order of sn-2 acyl chains, indicating a disordering effect. In a ternary system with POPC, quercetin, and IAPP, the reduction in the deuterium order parameter of sn-2 acyl chains is less pronounced, underscoring quercetin's protective role. Unlike IAPP, ApoAI and insulin fibrils undergo significant structural reorganization in the membrane-bound state. Quercetin attenuative effects are observed only with ApoAI, highlighting its potential as a protective agent against amyloid-induced membrane disruption. These findings provide valuable insights into the interactions between polyphenols, amyloid fibrils, and lipid membranes, contributing to the understanding of membrane-associated amyloid pathologies. [ABSTRACT FROM AUTHOR]

Hydration plays a crucial role in the refolding of intrinsically disordered proteins into amyloid fibrils; however, the specific interactions between water and protein that may contribute to this process are still unknown. In our previous studies of alpha‐synuclein (aSyn), we have shown that waters confined in fibril cavities are stabilizing features of this pathological fold; and that amino acids that hydrogen bond with these confined waters modulate primary and seeded aggregation. Here, we extend our aSyn molecular dynamics (MD) simulations with three new polymorphs and correlate MD trajectory information with known post‐translational modifications (PTMs) and experimental data. We show that cavity residues are more evolutionarily conserved than non‐cavity residues and are enriched with PTM sites. As expected, the confinement within hydrophilic cavities results in more stably hydrated amino acids. Interestingly, cavity PTM sites display the longest protein‐water hydrogen bond lifetimes, three‐fold greater than non‐PTM cavity sites. Utilizing the deep mutational screen dataset by Newberry et al. and the Thioflavin T aggregation review by Pancoe et al. parsed using a fibril cavity/non‐cavity definition, we show that hydrophobic changes to amino acids in cavities have a larger effect on fitness and aggregation rate than residues outside cavities, supporting our hypothesis that these sites are involved in the inhibition of aSyn toxic fibrillization. Finally, we expand our study to include analysis of fibril structures of tau, FUS, TDP‐43, prion, and hnRNPA1; all of which contained hydrated cavities, with tau, FUS, and TDP‐43 recapitulating our PTM results in aSyn fibril cavities. [ABSTRACT FROM AUTHOR]

The elucidation of interactions between functional proteins and amyloid fibrils is crucial for understanding the molecular basis of amyloid diseases, which are characterized by protein misfolding and aggregation. Polyphenols, due to their diverse biological properties, have garnered attention for their potential to modulate these protein-fibril interactions, thereby influencing disease progression and offering therapeutic possibilities. In this study, we investigated the effects of quercetin and its binary combinations with other polyphenols on the binding affinity between cytochrome c, in both its reduced and oxidized forms, and amyloid fibrils of insulin and apolipoprotein A-I. Our results demonstrate that quercetin complexation with cytochrome c decreases the binding affinity of insulin fibrils for both forms of the protein, while increasing the affinity for apolipoprotein A-I fibrils. This modulation was attributed to competitive or allosteric effects exerted by quercetin on cytochrome c. Additionally, while binary combinations of quercetin with other polyphenols did not reduce the affinity of insulin fibrils for oxidized cytochrome c, they did decrease the affinity in the case of reduced counterpart. These findings highlight the selective and significant impact of polyphenolic compounds on the interactions between amyloid fibrils and functional proteins, suggesting potential pathways for therapeutic intervention in amyloid-related disorders.

Transthyretin (TTR), also known as prealbumin, is a highly conserved protein found in several vertebrate species. Dissociation of TTR tetramer followed by aggregation and misfolding of the monomers causes genesis of insoluble amyloid fibrils, which might systematically lead to peripheral and/or autonomic neuropathy and other systemic manifestations. Structural insights into the conformational landscape of TTR monomers upon mutation are useful for understanding amyloid aggregation. In this study, we used all-atom molecular dynamics (MD) simulations to investigate the role of V30M mutation on the structure of TTR monomer. The MD simulations of the wild-type and V30M TTR monomeric showed that the V30M mutation leads to dissociation of one of the edge strands, strand C, from the core of the monomer and forms an ensemble of partially unfolded conformations. These changes can be linked to disruption in the inter-strand hydrogen bonding and the secondary structure of the monomeric state. Moreover, non-covalent interactions in the E–F loop region, which is important for aggregation propensity of TTR, are also perturbed. Overall, the MD simulations provide comparative structural insights into the dynamics of wild-type and V30M TTR monomers. [ABSTRACT FROM AUTHOR]

The elucidation of interactions between functional proteins and amyloid fibrils is crucial for understanding the molecular basis of amyloid diseases, which are characterized by protein misfolding and aggregation. Polyphenols, due to their diverse biological properties, have garnered attention for their potential to modulate these protein-fibril interactions, thereby influencing disease progression and offering therapeutic possibilities. In this study, we investigated the effects of quercetin and its binary combinations with other polyphenols on the binding affinity between cytochrome c, in both its reduced and oxidized forms, and amyloid fibrils of insulin and apolipoprotein A-I. Our results demonstrate that quercetin complexation with cytochrome c decreases the binding affinity of insulin fibrils for both forms of the protein, while increasing the affinity for apolipoprotein A-I fibrils. This modulation was attributed to competitive or allosteric effects exerted by quercetin on cytochrome c. Additionally, while binary combinations of quercetin with other polyphenols did not reduce the affinity of insulin fibrils for oxidized cytochrome c, they did decrease the affinity in the case of reduced counterpart. These findings highlight the selective and significant impact of polyphenolic compounds on the interactions between amyloid fibrils and functional proteins, suggesting potential pathways for therapeutic intervention in amyloid-related disorders. [ABSTRACT FROM AUTHOR]

Carbon nanotubes, an emerging class of carbon nanomaterials, possess tremendous potential for application in biotechnology and biomedicine particularly in neurological disorders. Carbon nanotubes owing to their fascinating properties have the potential to revolutionize medicine and technology, particularly in the realm of drug delivery, biosensing, bioimaging, and as therapeutic agents to tackle complex neurological disorders such as Alzheimer’s and Parkinson’s disease. In this review, a summary of the use of carbon nanotubes for neuropathological outcomes such as alleviating oxidative stress and amyloid formation, which are well-studied molecular outcomes associated with Alzheimer’s and Parkinson’s disease. In the end, challenges associated with the clinical testing of carbon nanotubes and possible ways to overcome them are highlighted.

Abstract Amyloid fibrils derived from different proteins have been proved as a promising material for adsorption of various pollutants from wastewater, which showed advantages of low cost and eco-friendliness. However, most of the amyloid fibrils derived from animal-based proteins with high environmental footprint, while more sustainable amyloid fibrils derived from plant materials are desirable. In this study, a plant-derived amyloid fibril was extracted from the commonly used wheat flour with a simple and scalable protein purification and fibrillization process. Interestingly, the amyloid fibrils showed good adsorption capacity towards typical organic dyes (Eosin Y (EY) and Congo red (CR)) from contaminated water. Adsorption kinetic analysis indicated the adsorption process to EY or CR by wheat flour amyloid well fitted with a pseudo-second-order model. The adsorption also followed a Langmuir isothermal model with adsorption capacities of 333 mg/g and 138 mg/g towards CR and EY, respectively. This work demonstrated the feasibility to utilize the plant-based amyloid fibril for organic dyes removal from contaminated water, which provided an affordable, sustainable and scalable tool for organic dyes removal from wastewater. Graphical Abstract

It is assumed that the stability and bioaccessibility of iron ions in iron–pea protein fibril (Fe-Fib PP) nanocomposite can be remarkably enhanced, and Fe-Fib PP exhibits great potential as an effective iron fortificant. Fe-Fib PP, a stable and effective iron supplement, was fabricated based on the reducing property of pea protein fibrils, derived from pea protein through thermal treatment at pH 2.0. The results demonstrated that the reducing power of iron was remarkably affected by fibril concentration and fibrillization degree. The reducing power of pea protein fibrils gradually enhanced from 0.31 to 0.92 with the increase in incubation time from 0 to 48 h. Compared with iron nanoparticles (Fe–Nano), Fe-Fib PP possessed much higher dispersibility. Additionally, the stability of iron in Fe-Fib PP was significantly higher than that in Fe–Nano under different storage conditions. X-ray photoelectron spectroscopy (XPS) outcomes revealed Fe (II) content in Fe-Fib PP (70.75 ± 0.65%) was remarkably higher than that of Fe–Nano (56.05 ± 0.50%). In addition, the bioaccessibility of Fe (II) dramatically improved from 42.7% to 62.8% using PP fibrils as carriers. The findings suggest that Fe-Fib PP is an effective iron nutrition enhancer.

Plant natriuretic peptides (PNPs) are hormones that have been extracted from many different species, with the Arabidopsis thaliana PNP (AtPNP-A) being the most studied among them. AtPNP-A is a signaling molecule that consists of 130 residues and is secreted into the apoplast, under conditions of biotic or abiotic stress. AtPNP-A has distant sequence homology with human ANP, a protein that forms amyloid fibrils in vivo. In this work, we investigated the amyloidogenic properties of a 34-residue-long peptide, located within the AtPNP-A sequence, in three different pH conditions, using transmission electron microscopy, X-ray fiber diffraction, ATR FT-IR spectroscopy, Congo red and Thioflavin T staining assays. We also utilize bioinformatics tools to study its association with known plant amyloidogenic proteins and other A. thaliana proteins. Our results reveal a new case of a pH-dependent amyloid forming peptide in A. thaliana, with a potential functional role.

Amyloidosis is a condition involving a disparate group of pathologies characterized by the extracellular deposition of insoluble fibrils composed of broken-down proteins. These proteins can accumulate locally, causing peculiar symptoms, or in a widespread way, involving many organs and. causing severe systemic failure. The damage that is created is related not only to the accumulation of. amyloid fibrils but above all to the precursor oligomers of the fibrils that manage to enter the cell in a very particular way. This article analyzes the current state of research related to the entry of these oligomers into the cell membrane and the theories related to their toxicity. The paper proposed here not only aims to review the contents in the literature but also proposes a new vision of amyloid toxicity. that could occur in a multiphase process catalyzed by the cell membrane itself. In this process, the denaturation of the lipid bilayer is followed by the stabilization of a pore through energetically favorable self-assembly processes which are achieved through particular oligomeric structures. [ABSTRACT FROM AUTHOR]

A progressive aggregation of misfolded proteins is a hallmark of numerous pathologies including diabetes Type 2, Alzheimer's disease, and Parkinson's disease. As a result, highly toxic protein aggregates, which are known as amyloid fibrils, are formed. A growing body of evidence suggests that phospholipids can uniquely alter the secondary structure and toxicity of amyloid aggregates. However, the role of phosphatidic acid (PA), a unique lipid that is responsible for cell signaling and activation of lipid‐gated ion channels, in the aggregation of amyloidogenic proteins remains unclear. In this study, we investigate the role of the length and degree of unsaturation of fatty acids (FAs) in PA in the structure and toxicity of lysozyme fibrils formed in the presence of this lipid. We found that both the length and saturation of FAs in PA uniquely altered the secondary structure of lysozyme fibrils. However, these structural differences in PA caused very little if any changes in the morphology of lysozyme fibrils. We also utilized cell toxicity assays to determine the extent to which the length and degree of unsaturation of FAs in PA altered the toxicity of lysozyme fibrils. We found that amyloid fibrils formed in the presence of PA with C18:0 FAs exerted significantly higher cell toxicity compared to the aggregates formed in the presence of PA with C16:0 and C18:1 FAs. These results demonstrated that PA can be an important player in the onset and spread of amyloidogenic diseases. [ABSTRACT FROM AUTHOR]

Aggregation of antibody light chain proteins is associated with the progressive disease light chain amyloidosis. Patient‐derived amyloid fibrils are formed from light chain variable domain residues in non‐native conformations, highlighting a requirement that light chains unfold from their native structures in order to aggregate. However, mechanistic studies of amyloid formation have primarily focused on the self‐assembly of natively unstructured peptides, and the role of native state unfolding is less well understood. Using a well‐studied light chain variable domain protein known as WIL, which readily aggregates in vitro under conditions where the native state predominates, we asked how the protein concentration and addition of pre‐formed fibril "seeds" alter the kinetics of aggregation. Monitoring aggregation with thioflavin T fluorescence revealed a distinctly non‐linear dependence on concentration, with a maximum aggregation rate observed at 8 μM protein. This behavior is consistent with formation of alternate aggregate structures in the early phases of amyloid formation. Addition of N‐ or C‐terminal peptide tags, which did not greatly affect the folding or stability of the protein, altered the concentration dependence of aggregation. Aggregation rates increased in the presence of pre‐formed seeds, but this effect did not eliminate the delay before aggregation and became saturated when the proportion of seeds added was greater than 1 in 1600. The complexity of aggregation observed in vitro highlights how multiple species may contribute to amyloid pathology in patients. [ABSTRACT FROM AUTHOR]