Shear flow during reciprocal shaking does not disrupt protein films at the solid-liquid interface.

Recent advances in small molecules for amyloid fibril inhibition: chemical strategies and molecular mechanistic insights from lysozyme and insulin models.

Genetic transformation of rice (Oryza sativa L.) plants with the Hsp101 genomic fragment containing the coding sequence and its natural, heat-inducible promoter produced transgenics with enhanced heat tolerance.

cGAS-STING activation in Parkinson's Disease: From mechanisms to Disease-Modifying therapeutic strategies.

3-Deoxyanthocyanidins inhibit β-amyloid aggregation, toxicity, and mitochondrial dysfunction: Evidence from MC-65 cells and molecular dynamics simulations.

Charge-dependent mechanisms of ionic polysaccharides modulating myofibrillar protein gelation: A dual-system evaluation from minced chicken gel to protein system.

Two curcumin derivatives, structurally related to the CRANAD family of compounds, were investigated for their theranostic properties in Alzheimer's disease. They exhibited fluorescence emission in the NIR region (650-690nm) with a significant Stokes' shift (70-90nm). Their affinity for Aβ aggregates, oligomers and monomers allows to detect, differentiate and map in vitro the various amyloid species within and around the plaques by fluorescence microscopy. Fluorescence lifetime microscopy, a robust and sensitive technique allowing to visualize biomolecules with high spatial resolution at nanomolar level, was employed to discriminate the less soluble and more toxic species from the more soluble ones by determination of fluorescence lifetime values at the core and the periphery of the β-amyloid plaques, without the need for the use of antibodies. In vivo brain images show that the fluorescence signals of the sensors are 5-6 times higher for transgenic mice with aberrant proteins than wild type mice after intraperitoneal injection, differentiating plaques of amyloid beta (Aβ) protein in real samples in vivo. These experiments also showed a good blood brain barrier penetration of the sensors, which remain in the brain for 90-120min, opening up the possibility of their therapeutic use. In vitro studies showed a good activity of both compounds as inhibitors of Aβ aggregation into small soluble oligomers and large insoluble aggregates and also the inhibition of tau protein aggregation, both in a dose-dependent manner. These studies confirm that both compounds have an unprecedented profile that justifies their further study as small-molecule theranostic agents in AD.

Effects of plasma-activated water rinsing on gastric emptying and the digestion kinetics of surimi myofibrillar proteins under dynamic in vitro digestion.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by the progressive loss of motor neurons. ALS often overlaps clinically and pathologically with frontotemporal dementia (FTD), the second most common form of dementia. Like many neurodegenerative disorders, both ALS and FTD share a crucial pathological hallmark, the aggregation of misfolded proteins into insoluble inclusions in degenerating neurons. This process is referred to as proteinopathy. This review focuses on the proteinopathies associated with ALS, including aggregates of TDP-43, SOD1, FUS, and CHCHD10, which disrupt critical cellular processes such as RNA metabolism, mitochondrial function, and protein homeostasis. The review highlights to the identification of new types of mitochondrial and cytosolic aggregates linked to CHCHD10-related ALS. Although the precise pathological mechanisms remain to be fully elucidated, strategies aimed at restoring proteostasis and reducing protein aggregation may be promising therapeutic approaches for treating ALS, as they directly target fundamental pathogenic mechanisms.

Nucleophagy: The guardian of genome stability - from molecular mechanisms to disease associations.

Transthyretin (TTR) is a tetrameric transporter of retinol and thyroxine that aggregates in the central and peripheral nervous system upon a severe pathology known as transthyretin amyloidosis. Although small molecular weight drugs can stabilize TTR preventing its aggregation, molecular mechanisms of transthyretin amyloidosis remain poorly understood. Accumulating evidence indicates that lipids can alter TTR stability by facilitating protein aggregation into toxic oligomers and fibrils. Consequently, pathological changes in the lipid composition of plasma membranes can be responsible for the onset and progression of transthyretin amyloidosis. In this study, we investigated the role of concentration-dependent changes in phosphatidylcholine (PC), one of the most abundant phospholipids in the plasma membrane, on the rate of TTR aggregation. For this, TTR was exposed to large unilamellar vesicles (LUVs) composed of 30%, 35%, and 40% PC. We found that a decrease in the concentration of PC from 40% to 35% drastically accelerated TTR aggregation. We also observed an increase in the cytotoxicity of TTR aggregates formed in the presence of 35% PC compared to TTR fibrils grown in the presence of LUVs with 40% PC. These results indicate that changes in the concentration of PC in the plasma membrane could trigger amyloid formation that leads to transthyretin amyloidosis.

Protein aggregation is a central challenge in protein science and biopharmaceutical formulation, yet its reproduction by all-atom molecular dynamics (MD) simulations remains difficult within accessible computational limits. Here, we demonstrate that explicit control of the denaturation extent of initial protein structures enables reproducible thermal aggregation in MD simulations. Using hen egg white lysozyme (LYZ) as a model system, we generated unfolded conformational ensembles by high-temperature MD (300-700 K) and examined their aggregation behavior. Two factors proved critical for reproducing LYZ aggregation: a sufficiently high initial denaturation extent and direct initiation of simulations without an additional structural relaxation step. Under these conditions, 100 ns simulations successfully reproduced experimentally established additive effects: arginine suppressed aggregation only at high concentration (1 M), while sodium chloride promoted aggregation via nonspecific ionic effects. This proof-of-concept study establishes denaturation-controlled MD as a practical framework for computational evaluation of protein aggregation and stabilizer effects.

Like-charge attraction has been observed in processes like protein aggregation, colloidal stability, and surface wettability. That this attractive force between two like-charged objects at close separation arises from ionic electrostatic correlations was elucidated decades ago. However, how those correlations (and other energetic components of the electrochemical potential) ultimately translate into a drop in the force between two like-charged objects has not been fully explored. In order to gain a deeper insight into this process, we systematically analyze like-charge attraction between two parallel walls over a wide range of wall surface charge densities and electrolyte properties (ion concentration, valence, and size). We do this with over 70,000 classical density functional theory calculations of primitive model charged, hard sphere electrolytes. We find that, as the slit narrows, the ions undergo substantial rearrangements as their electrochemical potential's energetic components grow and decline nonmonotonically. This initially leads to relatively high co-ion concentration, but later causes the co-ions to be expelled from the slit. In turn, counterions shift to the middle of the pore, decreasing their edge concentrations, causing a decrease in the disjoining pressure. Our approach also provides more clarity about the extent to which charge inversion and like-charge attraction are related.

Impact of starch surface hydrophobicity on gluten aggregation behavior in dough-like model systems.

The ubiquitin-proteasome system (UPS) is important for podocyte health, but the specific UPS proteins involved in podocyte injury of diabetic nephropathy (DN) are not well known. Patients with DN have lower expression of USP46 in podocytes, which is linked to higher proteinuria. Deleting the Usp46 gene in podocytes of mice (Usp46PKO mice) led to spontaneous albuminuria and worsened podocyte injury and glomerular lesions under diabetic conditions. Mechanically, loss of USP46 caused cytosolic translocation and aggregation of TAR DNA binding protein 43 (TDP-43) in podocytes. Here, we identified acarbose as an agonist of USP46. Treatment with acarbose reduced TDP-43 aggregation in podocytes, prevented podocyte loss, and mitigated albuminuria in diabetic mice; the therapeutic efficacy of acarbose was abolished in Usp46PKO mice. This research elucidates the role of USP46 in podocyte homeostasis and injury in DN and indicates a potential therapeutic impact for acarbose in DN beyond the regulation of blood glucose concentrations through its activation of USP46.

Proteins exhibit complex phase behavior as they convert between the native state, the liquid condensate (or droplet) state and the solid condensate (or amyloid) state. To facilitate the study of these processes, we describe the FuzDrop method of predicting the condensation propensity of proteins to undergo liquid-liquid phase separation and to subsequently form amyloid aggregates. The method is based on the principle that liquid condensations reflect a balance between enthalpic and entropic contributions; FuzPred is an algorithm that provides sequence-based estimates for these contributions in stoichiometric complexes ( https://fuzpred.bio.unipd.it/predictor ). FuzDrop extends this algorithm to protein condensates, and enables prediction of the propensity for amyloid formation within liquid condensates, known as the condensation pathway to protein aggregation ( https://fuzdrop.bio.unipd.it/predictor ). This prediction is based on the principle that the sequence regions that promote aggregation within liquid condensates have a multiplicity of binding modes, because they have a strong propensity for both entropic-driven interactions to stabilize the droplet state and enthalpic-driven interactions to stabilize the amyloid state. The time required for FuzDrop predictions on the web server scales linearly with protein length and is typically ~30 s for a protein of 500 residues. By enabling predictions of protein phase behavior, FuzDrop may facilitate experimental studies directed at the development of therapies for protein condensation diseases.

Protein aggregation into amyloid fibrils underlies numerous human diseases, yet the most widely used fluorescent probe, Thioflavin T (ThT), offers an incomplete picture of the process and fails to detect certain fibril structures. Here, we introduce and characterize the photophysical properties of DANIR-2b(2OH), a water-soluble push-pull dye that overcomes these limitations. It successfully binds early prefibrillar aggregates and small fibrils of the human Islet Amyloid Polypeptide that elude detection by ThT, which we confirm by time-resolved cryo-electron microscopy of aliquots taken during the kinetic assays. We further demonstrate that DANIR-2b(2OH) can also track the aggregation of other amyloid proteins, such as insulin and Aβ1-42. The protein-dye interaction was characterized via steady-state and time-resolved fluorescent spectroscopy. DANIR-2b(2OH) features environment-sensitive emission, high photostability, and a straightforward synthesis. Critically, it provides a substantially lower noise level in standard plate-reader assays, allowing the tracking of aggregation processes that are not visible in standard ThT measurements. This establishes DANIR-2b(2OH) as a highly sensitive and broadly applicable probe for real-time amyloid aggregation measurements and imaging.

Surface-catalyzed secondary nucleation is well-known in protein aggregation but underexplored in supramolecular polymerization of π-systems. Herein, we report a unique example of multistep secondary nucleation-dominated controlled supramolecular polymerization of an amide-functionalized naphthalene monoimide derivative (O-NMI-1) in a methylcyclohexane:decane mixture, undergoing a stepwise transformation from a metastable particle to crystalline nanorods via intermediate nanotapes over 48 h. The resulting hydrogen-bonded nanorods display enhanced emission, a high quantum yield (34%) and strong circularly polarized luminescence (CPL), revealing an unprecedented correlation between their structural evolution and the time-dependent increment in the CPL intensity, reaching an appreciably high luminescence dissymmetry factor (glum) of ∼0.088 in the film state. Contrastingly, the seeded polymerization of O-NMI-1 follows a fragmentation-dominated mechanism, yielding unexpected fragmented nanorods in multi-seeding cycles. Control experiments with a structurally similar analogue (O-NMI-2), lacking a methylene spacer, revealed no CPL, highlighting the spacer's crucial role in controlling the excited-state chiral emission. The contrasting behavior between the two monomers enabled hetero-seeded polymerization of O-NMI-1 nanorods from O-NMI-2 fibrillar seeds, producing CPL-active unique heterostructures under kinetic control from two initial CPL-silent components. The present findings unveil the overlooked impact of secondary nucleation in temporal control of supramolecular chirality and CPL-activity in homo- and heterostructures of supramolecular polymers.

The transcriptional repressor Fli1 inhibits proteostasis during nutrient stress to limit NK cell persistence in solid tumors.

Aggregation and deposition of TAR DNA-binding protein 43 (TDP-43) is a salient pathological signature of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration-TDP (FTLD-TDP). TDP-43 proteostasis and aggregation are controlled by several posttranslational modifications, including ubiquitination. While multiple E3 ubiquitin ligases are known to facilitate TDP-43 clearance, little is known about the role of deubiquitinases (DUBs) in controlling TDP-43 proteostasis. Through an unbiased discovery screen of DUBs, here we identify and demonstrate using in vitro and in vivo models, as well as human brain tissue, that ubiquitin-specific peptidase-19 (USP19) acts as a TDP-43-directed DUB that removes K48- and K63-linked ubiquitin conjugates from TDP-43 and preferentially promotes cytoplasmic aggregation of TDP-43 C-terminal fragments (TDP-CTFs) through its catalytic activity. Specifically, the endoplasmic reticulum (ER)-anchored USP19 isoform (USP19-ER) exhibits superior activity in deubiquitinating TDP-CTFs, enhancing its phase separation and aggregation, compared to its cytosolic isoform (USP19-Cyto). Furthermore, as TDP-CTFs are generated at the ER, USP19 acts to couple the aggregation of TDP-CTFs to ER stress (ATF6, ATF4, IRE1, & CHOP). In humans, USP19 protein levels increase in FTLD-TDP brains, which extensively colocalize with cytoplasmic phospho-TDP-43 (pTDP-43) pathology. Importantly, we demonstrate in vivo that genetic reduction of usp19 mitigates pTDP-43 pathology, astrogliosis, and ER stress while reversing long-term potentiation (LTP) and motor deficits in a mouse model of TDP-43 pathogenesis (TAR4 mice). These findings establish a critical role of USP19 at the nexus of TDP-43 proteostasis and ER stress, implicating its pathogenic role in FTLD-TDP and ALS.