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Amyloid β (Aβ) aggregates are implicated in the pathology of several neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, and Parkinson's disease, and damage to membranes is considered one of the pathology-related effects of Aβ. Experiments in vitro indicate that Aβ can damage these membranes; however, such experiments were performed at Aβ concentrations in the micromolar range, several orders above the physiologically relevant conditions. Our studies with Aβ42 in the low nanomolar concentrations did not reveal any damage to the supported lipid bilayer, questioning this membrane damage mechanism of Aβ. However, the phospholipid composition can be a factor contributing to the interaction of Aβ with the membrane. Therefore, in this study, we investigated the interaction of 50 nM Aβ42 with supported lipid bilayers composed of equimolar ratios of POPS and POPC at phospholipid concentrations of 0.1 and 0.25 mg/mL. Using atomic force microscopy (AFM), we observed that Aβ42 induced damage to bilayers at 0.1 mg/mL, characterized by forming defects that grew in size and number over time. The defects penetrate only the upper leaflet of the bilayer, but no such defects were observed at 0.25 mg/mL phospholipid concentrations. We additionally determined Young's modulus of these bilayers as a measure of stiffness, and these values were 6.9 ± 3.6 MPa and 16.6 ± 5.3 MPa for the 0.1 mg/mL and the 0.25 mg/mL bilayers, respectively. These findings suggest that Aβ42's ability to induce bilayer damage depends on membrane stiffness, with softer bilayers (0.1 mg/mL) being more susceptible to Aβ42-induced damage. The results are discussed and compared with models in which Aβ42 oligomers create localized membrane damage. The implication of the results to the mechanisms of the Aβ42 oligomer pathology is discussed.

Over a century has passed since Alois Alzheimer first described Alzheimer's disease (AD), and since then, researchers have made significant strides in understanding its pathology. One key feature of AD is the presence of amyloid-β (Aβ) peptides, which form amyloid plaques, and therefore, it is a primary target for treatment studies. Naturally occurring peptides have garnered attention for their potential pharmacological benefits, particularly in the central nervous system. In this study, nine peptide derivatives of Crotamine, a polypeptide from Crotalus durissus terrificus Rattlesnake venom, as well as one d-enantiomer, were evaluated for their ability to modulate Aβ 42 aggregation through various assays such as ThT, QIAD, SPR, and sFIDA. All tested peptides were able to decrease Aβ 42 aggregation and eliminate Aβ 42 aggregates. Additionally, all of the peptides showed an affinity for Aβ 42 . This study is the first to describe the potential of crotamine derivative peptides against Aβ 42 aggregation and to identify a promising d-peptide that could be used as an effective pharmacological tool against AD in the future.

Aβ oligomers are being investigated as cytotoxic agents in Alzheimer's disease (AD). Because of their transient nature and conformational heterogeneity, the relationship between the structure and activity of these oligomers is still poorly understood. Hence, methods for stabilizing Aβ oligomeric species relevant to AD are needed to uncover the structural determinants of their cytotoxicity. Here, we build on the observation that metal ions and metabolites have been shown to interact with Aβ, influencing its aggregation and stabilizing its oligomeric species. We thus developed a method that uses zinc ions, Zn(II), to stabilize oligomers produced by the 42-residue form of Aβ (Aβ 42 ), which is dysregulated in AD. These Aβ 42 -Zn(II) oligomers are small in size, spanning the 10-30 nm range, stable at physiological temperature, and with a broad toxic profile in human neuroblastoma cells. These oligomers offer a tool to study the mechanisms of toxicity of Aβ oligomers in cellular and animal AD models.

ADAM 17, a disintegrin and metalloproteinase 17 belonging to the adamalysin protein family, is a Zn 2+ -dependent type-I transmembrane α-secretase protein. As a major sheddase, ADAM 17 acts as an indispensable regulator of chief cellular events and controls diverse cytokines, adhesion molecules, and growth factors. The signal peptide (residues 1-17) of ADAM 17 targets the protein to the secretory pathway and gets cleaved off afterward. No other function is documented for the ADAM 17 signal peptide (ADAM 17-SP) inside the cells. Here, we have taken a reductionist approach to understand the biophysical properties of ADAM 17-SP. Aiming to understand the possibility of aggregation, we found several aggregation-prone segments in the signal peptide. We performed in vitro experiments to show that the signal peptide forms amyloid-like aggregates in buffered conditions. We also studied its aggregation in the presence of sodium tripolyphosphate and heparin to correlate with the cellular conditions, as these biomolecules are naturally present inside cells. Further, we performed seeding experiments to observe the possibility of ADAM 17-SP aggregate interaction with the Aβ42 peptide. The results suggest that its seeds escalate the aggregation kinetics of the Aβ42 peptide and form heteromeric aggregates with it. We believe this finding could further intensify the aggregation studies on other signal peptides and shed light on the potential role of these segments other than signaling.

An amyloid precursor protein inhibitor (APPI) and amyloid beta 42 (Aβ42) are both subdomains of the human transmembrane amyloid precursor protein (APP). In the brains of patients with Alzheimer's disease (AD), Aβ42 oligomerizes into aggregates of various sizes, with intermediate, low-molecular-weight Aβ42 oligomers currently being held to be the species responsible for the most neurotoxic effects associated with the disease. Strategies to ameliorate the toxicity of these intermediate Aβ42 oligomeric species include the use of short, Aβ42-interacting peptides that either inhibit the formation of the Aβ42 oligomeric species or promote their conversion to high-molecular-weight aggregates. We therefore designed such an Aβ42-interacting peptide that is based on the β-hairpin amino acid sequence of the APPI, which exhibits high similarity to the β-sheet-like aggregation site of Aβ42. Upon tight binding of this 20-mer cyclic peptide to Aβ42 (in a 1:1 molar ratio), the formation of Aβ42 aggregates was enhanced, and consequently, Aβ42-mediated cell toxicity was ameliorated. We showed that in the presence of the cyclic peptide, interactions of Aβ42 with both plasma and mitochondrial membranes and with phospholipid vesicles that mimic these membranes were inhibited. Specifically, the cyclic peptide inhibited Aβ42-mediated mitochondrial membrane depolarization and reduced Aβ42-mediated apoptosis and cell death. We suggest that the cyclic peptide modulates Aβ42 aggregation by enhancing the formation of large aggregates─as opposed to low-molecular-weight intermediates─and as such has the potential for further development as an AD therapeutic.

The recent approval of antibody-based therapy for targeting the clearance of amyloid plaques fuels the research in designing small molecules and peptide inhibitors to target the aggregation of Aβ-peptides. Here, we report that the 15-residue ααγ-hybrid peptide not only inhibits the aggregation of soluble Aβ 42 into fibrils but also disintegrates the aggregated Aβ 42 fibrils into smaller assemblies. Further, the hybrid peptide completely rescues neuronal cells from the toxicity of Aβ 42 at equimolar concentrations. The shorter 10- and 12-mer peptides showed weak aggregation inhibition activity, while the fully hydrophobic 15-mer ααγ-hybrid peptide analogue showed no aggregation inhibition activity. Further, the 15-mer ααγ-hybrid peptide showed resistance against trypsin digestion and also nontoxic to the neuronal cells. The CD revealed that the peptide upon interaction induces a helix-type conformation in the Aβ 42 . This is in sharp contrast to the β-sheet conformation of Aβ 42 upon incubation. The two-dimensional-NMR (2D-NMR) analysis revealed a large perturbation in the chemical shifts of residues at the N-terminus. The presence of 15-mer peptide at an equimolar concentration of Aβ 42 showed less tendency for aggregation and also exhibited nontoxicity to the neuronal cells. The results reported here may be useful in designing new therapeutics for Alzheimer's disease.

Alzheimer's disease (AD) is one of the world's most pressing health crises. AD is an incurable disease affecting more than 6.5 million Americans, predominantly the elderly, and in its later stages, leads to memory loss, dementia, and death. Amyloid β (Aβ) protein aggregates have been one of the pathological hallmarks of AD since its initial characterization. The early stages of Aβ accumulation and aggregation involve the formation of oligomers, which are considered neurotoxic and play a key role in further aggregation into fibrils that eventually appear in the brain as amyloid plaques. We have recently shown by combining ion mobility mass spectrometry (IM-MS) and atomic force microscopy (AFM) that Aβ42 rapidly forms dodecamers (12-mers) as the terminal oligomeric state, and these dodecamers seed the early formation of Aβ42 protofibrils. The link between soluble oligomers and fibril formation is one of the essential aspects for understanding the root cause of the disease state and is critical to developing therapeutic interventions. Utilizing a joint pharmacophore space (JPS) method, potential drugs have been designed specifically for amyloid-related diseases. These small molecules were generated based on crucial chemical features necessary for target selectivity. In this paper, we utilize our combined IM-MS and AFM methods to investigate the impact of three second-generation JPS small-molecule inhibitors, AC0201, AC0202, and AC0203, on dodecamer as well as fibril formation in Aβ42. Our results indicate that AC0201 works well as an inhibitor and remodeler of both dodecamers and fibril formation, AC0203 behaves less efficiently, and AC0202 is ineffective.

It is currently believed that molecular agents that specifically bind to and neutralize the toxic proteins/peptides, amyloid β (Aβ42), tau, and the tau-derived peptide PHF6, hold the key to attenuating the progression of Alzheimer's disease (AD). We thus tested our previously developed nonaggregating Aβ42 double mutant (Aβ42 DM ) as a multispecific binder for three AD-associated molecules, wild-type Aβ42, the tau K174Q mutant, and a synthetic PHF6 peptide. Aβ42 DM acted as a functional inhibitor of these molecules in in vitro assays and in neuronal cell-based models of AD. The double mutant bound both cytotoxic tau K174Q and synthetic PHF6 and protected neuronal cells from the accumulation of tau in cell lysates and mitochondria. Aβ42 DM also reduced toxic intracellular levels of calcium and the overall cell toxicity induced by overexpressed tau, synthetic PHF6, Aβ42, or a combination of PHF6and Aβ42. Aβ42 DM inhibited PHF6-induced overall mitochondrial dysfunction: In particular, Aβ42 DM inhibited PHF6-induced damage to submitochondrial particles (SMPs) and suppressed PHF6-induced elevation of the ζ-potential of inverted SMPs (proxy for the inner mitochondrial membrane, IMM). PHF6 reduced the lipid fluidity of cardiolipin/DOPC vesicles (that mimic the IMM) but not DOPC (which mimics the outer mitochondrial membrane), and this effect was inhibited by Aβ42 DM . This inhibition may be explained by the conformational changes in PHF6 induced by Aβ42 DM in solution and in membrane mimetics. On this basis, the paper presents a mechanistic explanation for the inhibitory activity of Aβ42 DM against Aβ42- and tau-induced membrane permeability and cell toxicity and provides confirmatory evidence for its protective function in neuronal cells.

Amyloid-β (Aβ) aggregates are believed to be one of the main causes of Alzheimer's disease. Aβ peptides form fibrils having cross β-sheet structures mainly through primary nucleation, secondary nucleation, and elongation. In particular, self-catalyzed secondary nucleation is of great interest. Here, we investigate the adsorption of Aβ42 peptides to the Aβ42 fibril to reveal a role of adsorption as a part of secondary nucleation. We performed extensive molecular dynamics simulations based on replica exchange with solute tempering 2 (REST2) to two systems: a monomeric Aβ42 in solution and a complex of an Aβ42 peptide and Aβ42 fibril. Results of our simulations show that the Aβ42 monomer is extended on the fibril. Furthermore, we find that the hairpin structure of the Aβ42 monomer decreases but the helix structure increases by adsorption to the fibril surface. These structural changes are preferable for forming fibril-like aggregates, suggesting that the fibril surface serves as a catalyst in the secondary nucleation process. In addition, the stabilization of the helix structure of the Aβ42 monomer on the fibril indicates that the strategy of a secondary nucleation inhibitor design for Aβ40 can also be used for Aβ42.