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Protein misfolding and aggregation have been associated with the onset of neurodegenerative disorders. Recent studies demonstrate that the aggregation process can result in a high diversity of protein conformational states, however the identity of the specific species responsible for the cellular damage is still unclear. Here, we use yeast as a model to systematically analyse the intracellular effect of expressing 21 variants of the amyloid-ß-peptide, engineered to cover a continuous range of intrinsic aggregation propensities. We demonstrate the existence of a striking negative correlation between the aggregation propensity of a given variant and the oxidative stress it elicits. Interestingly, each variant generates a specific distribution of protein assemblies in the cell. This allowed us to identify the aggregated species that remain diffusely distributed in the cytosol and are unable to coalesce into large protein inclusions as those causing the highest levels of oxidative damage. Overall, our results indicate that the formation of large insoluble aggregates may act as a protective mechanism to avoid cellular oxidative stress., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

The complex pathogenic mechanisms of Alzheimer's disease (AD) include the aggregation of β-amyloid peptides (Aβ) into oligomers or fibrils as well as Aβ-mediated oxidative stress, which require comprehensive treatment. Therefore, the inhibition of Aβ aggregation and free-radical scavenging are essential for the treatment of AD. Nanoparticles (NPs) have been found to influence Aβ aggregation process in vitro. Herein, we report the inhibition effects of molybdenum disulfide (MoS 2 ) NPs on Aβ aggregation. Polyvinylpyrrolidone-functionalized MoS 2 NPs were fabricated by a pulsed laser ablation method. We find that MoS 2 NPs exhibit multifunctional effects on Aβ peptides: inhibiting Aβ aggregation, destabilizing Aβ fibrils, alleviating Aβ-induced oxidative stress, as well as Aβ-mediated cell toxicity. Moreover, we show that MoS 2 NPs can block the formation of the Ca 2+ channel induced by Aβ fibrils in the cell membrane for the first time. Thus, these observations suggest that MoS 2 NPs have great potential for a multifunctional therapeutic agent against amyloid-related diseases.

Fibrillar deposits of the human islet amyloid polypeptide (hIAPP) are considered as a root of Type II diabetes mellitus. Fluorinated graphene quantum dots (FGQDs) are new carbon nanomaterials with unique physicochemical properties containing highly electronegative F atoms. Herein we report a single step synthesis method of FGQDs with an inhibitory effect on aggregation and cytotoxicity of hIAPP in vitro. Highly fluorescent and water dispersible FGQDs, less than 3 nm in size, were synthesized by the microwave-assisted hydrothermal method. Efficient inhibition capability of FGQDs to amyloid aggregation was demonstrated. The morphologies of hIAPP aggregates were observed to change from the entangled long fibrils to short thin fibrils and amorphous aggregates in the presence of FGQDs. In thioflavin T fluorescence analysis, inhibited aggregation with prolonged lag time and reduced fluorescence intensity at equilibrium were observed when hIAPP was incubated together with FGQDs. Circular dichroism spectrum results reveal that FGQDs could inhibit conformational transition of the peptide from native structure to β-sheets. FGQDs could also rescue the cytotoxicity of INS-1 cells induced by hIAPP in a dose dependent manner. This study could be beneficial for design and preparation of inhibitors for amyloids, which is important for prevention and treatment of amyloidosis.

Alzheimer's disease (AD) is a multifaceted disease that is hard to treat by single-modal treatment. AD starts with amyloid peptides, mitochondrial dysfunction, and oxidative stress and later is accompanied with chronic endoplasmic reticulum (ER) stress and autophagy dysfunction, resulting in more complicated pathogenesis. Currently, few treatments can modify the complicated pathogenic progress of AD. Compared to the treatment with exogenous antioxidants, the activation of global antioxidant defense system via Nrf2 looks more promising in attenuating oxidative stress in AD brains. Accompanying the activation of the Nrf2-mediated antioxidant defense system that reduce the AD-causative factor, oxidative stress, it is also necessary to activate the neurotrophic signaling pathway that replaces damaged organelles and molecules with new ones. Thus, the dual actions to activate both the Nrf2 antioxidant system and neurotrophic signaling pathway are expected to provide a better strategy to modify AD pathogenesis. Here, we review the current understanding of AD pathogenesis and neuronal defense systems and discuss a possible way to co-activate the Nrf2 antioxidant system and neurotrophic signaling pathway with the hope of helping to find a better strategy to slow AD., Competing Interests: The authors declare no conflict of interest.

Significance: Recently, chronic degenerative diseases have become one of the main health problems worldwide. That is the case of Alzheimer's disease (AD) and metabolic syndrome (MetS), whose expression can be influenced by different risk factors. Recent Advances: In recent decades, it has been widely described that MetS increases the risk of cognitive impairment and dementia. MetS pathogenesis involves several vascular risk factors such as diabetes, dyslipidemia, hypertension, and insulin resistance (I/R)., Critical Issues: Reported evidence shows that vascular risk factors are associated with AD, particularly in the development of protein aggregation, inflammation, oxidative stress, neuronal dysfunction, and disturbances in signaling pathways, with insulin receptor signaling being a common alteration between MetS and AD., Future Directions: Insulin signaling has been involved in tau phosphorylation and amyloid β (Aβ) metabolism. However, it has also been demonstrated that Aβ oligomers can bind to insulin receptors, triggering their internalization, decreasing neuron responsiveness to insulin, and promoting insulin I/R. Thus, it could be argued that Aβ could be a convergent factor in the development of both pathologies. Antioxid. Redox Signal. 26, 542-560.

Cerebrovascular insufficiency appears years prior to clinical symptoms in Alzheimer's disease. The soluble, highly toxic amyloid-β species, generated from the amyloidogenic processing of amyloid precursor protein, are known instigators of the chronic cerebrovascular insufficiency observed in both Alzheimer's disease patients and transgenic mouse models. We have previously demonstrated that pioglitazone potently reverses cerebrovascular impairments in a mouse model of Alzheimer's disease overexpressing amyloid-β. In this study, we sought to characterize the effects of amyloid-β overproduction on the cerebrovascular proteome; determine how pioglitazone treatment affected the altered proteome; and analyze the relationship between normalized protein levels and recovery of cerebrovascular function. Three-month-old wildtype and amyloid precursor protein mice were treated with pioglitazone- (20 mg/kg/day, 14 weeks) or control-diet. Cerebral arteries were surgically isolated, and extracted proteins analyzed by gel-free and gel-based mass spectrometry. 193 cerebrovascular proteins were abnormally expressed in amyloid precursor protein mice. Pioglitazone treatment rescued a third of these proteins, mainly those associated with oxidative stress, promotion of cerebrovascular vasocontractile tone, and vascular compliance. Our results demonstrate that amyloid-β overproduction perturbs the cerebrovascular proteome. Recovery of cerebrovascular function with pioglitazone is associated with normalized levels of key proteins in brain vessel function, suggesting that pioglitazone-responsive cerebrovascular proteins could be early biomarkers of Alzheimer's disease.

The amyloid precursor protein (APP) is a membrane protein engaged in complex proteolytic pathways. APP and its derivatives have been shown to play a central role in Alzheimer's disease (AD), a progressive neurodegenerative disease characterized by memory decline. Despite a huge effort from the research community, the primary cause of AD remains unclear, making it crucial to better understand the physiological role of the APP pathway in brain plasticity and memory. Drosophila melanogaster is a model system well-suited to address this issue. Although relatively simple, the fly brain is highly organized, sustains several forms of learning and memory, and drives numerous complex behaviors. Importantly, molecules and mechanisms underlying memory processes are conserved from flies to mammals. The fly encodes a single non-essential APP homolog named APP-Like (APPL). Using in vivo inducible RNA interference strategies, it was shown that APPL knockdown in the mushroom bodies (MB)-the central integrative brain structure for olfactory memory-results in loss of memory. Several APPL derivatives, such as secreted and full-length membrane APPL, may play different roles in distinct types of memory phases. Furthermore, overexpression of Drosophila amyloid peptide exacerbates the memory deficit caused by APPL knockdown, thus potentiating memory decline. Data obtained in the fly support the hypothesis that APP acts as a transmembrane receptor, and that disruption of its normal function may contribute to cognitive impairment during early AD.

Background: For long time Alzheimer's disease has been attributed to a cholinergic deficit. More recently, it has been considered dependent on the accumulation of the amyloid beta peptide (Aβ), which promotes neuronal loss and impairs neuronal function. Results/methodology: In the present study, using biophysical and biochemical experiments we tested the hypothesis that in addition to its role as a neurotransmitter, acetylcholine may exert its action as an anti-Alzheimer agent through a direct interaction with Aβ., Conclusion: Our data provide evidence that acetylcholine favors the soluble peptide conformation and exerts a neuroprotective effect against the neuroinflammatory and toxic effects of Aβ. The present paper paves the way toward the development of new polyfunctional anti-Alzheimer therapeutics capable of intervening on both the cholinergic transmission and the Aβ aggregation.

APPswe/PS1dE9 and Tg2576 are very common transgenic mouse models of Alzheimer's disease (AD), used in many laboratories as tools to research the mechanistic process leading to the disease. In order to augment our knowledge about the amyloid-β (Aβ) isoforms present in both transgenic mouse models, we have developed two chromatographic methods, one acidic and the other basic, for the characterization of the Aβ species produced in the brains of the two transgenic mouse models. After immunoprecipitation and micro-liquid chromatography-electrospray ionization mass spectrometry/mass spectrometry, 10 species of Aβ, surprisingly all of human origin, were detected in the brain of Tg2576 mouse, whereas 39 species, of both murine and human origin, were detected in the brain of the APP/PS1 mouse. To the best of our knowledge, this is the first study showing the identification of such a high number of Aβ species in the brain of the APP/PS1 transgenic mouse, whereas, in contrast, a much lower number of Aβ species were identified in the Tg2576 mouse. Therefore, this study brings to light a relevant phenotypic difference between these two popular mice models of AD.

Background: Alzheimer's disease (AD) is the most common form of neurodegenerative disorder that leads to a decline in cognitive function. In AD, aggregates of amyloid β peptide precede the accumulation of neurofibrillary tangles, both of which are hallmarks of the disease. The great majority (>90 %) of the AD cases are not originated from genetic defects, therefore supporting the central roles of epigenetic modifications that are acquired progressively during the life span. Strong evidences have indicated the implication of epigenetic modifications, including histone modification and DNA methylation, in AD. Recent studies revealed that 5-hydroxymethylcytosine (5hmC) is dynamically regulated during neurodevelopment and aging., Results: We show that amyloid peptide 1-42 (Aβ1-42) could significantly reduce the overall level of 5hmC in vitro. We found that the level of 5hmC displayed differential response to the pathogenesis in different brain regions, including the cortex, cerebellum, and hippocampus of APP-PSEN1 double transgenic (DTg) mice. We observed a significant decrease of overall 5hmC in hippocampus, but not in cortex and cerebellum, as the DTg mice aged. Genome-wide profiling identified differential hydroxymethylation regions (DhMRs) in DTg mice, which are highly enriched in introns, exons and intergenic regions. Gene ontology analyses indicated that DhMR-associated genes are highly enriched in multiple signaling pathways involving neuronal development/differentiation and neuronal function/survival., Conclusions: 5hmC-mediated epigenetic regulation could potentially be involved in the pathogenesis of AD.

Currently, deficit of amyloid β-peptide (Aβ) clearance from the brain is considered as one of the possible causes of amyloid accumulation and neuronal death in the sporadic form of Alzheimer's disease (AD). Aβ clearance can involve either specific proteases present in the brain or Aβ-binding/transport proteins. Among amyloid-degrading enzymes the most intensively studied are neprilysin (NEP) and insulin-degrading enzyme (IDE). Since ageing and development of brain pathologies is often accompanied by a deficit in the levels of expression and activity of these enzymes in the brain, there is an urgent need to understand the mechanisms involved in their regulation. We have recently reported that NEP and also an Aβ-transport protein, transthyretin are epigenetically co-regulated by the APP intracellular domain (AICD) and this regulation depends on the cell type and APP695 isoform expression in a process that can be regulated by the tyrosine kinase inhibitor, Gleevec. We have now extended our work and shown that, unlike NEP, another amyloid-degrading enzyme, IDE, is not related to over-expression of APP695 in neuroblastoma SH-SY5Y cells but is up-regulated by APP751 and APP770 isoforms independently of AICD but correlating with reduced HDAC1 binding to its promoter. Studying the effect of the nuclear retinoid X receptor agonist, bexarotene, on NEP and IDE expression, we have found that both enzymes can be up-regulated by this compound but this mechanism is not APP-isoform specific and does not involve AICD but, on the contrary, affects HDAC1 occupancy on the NEP gene promoter. These new insights into the mechanisms of NEP and IDE regulation suggest possible pharmacological targets in developing AD therapies.

Self-assembly and aggregation of amyloid peptides, such as Aβ(1-40) and Aβ(1-42), lead to the development of Alzheimer disease and similar neurodegenerative disorders associated with protein aggregation. The structures of large aggregates, specifically fibrils, are well characterized. However, our understanding about the structure of oligomeric forms of amyloids is incomplete and needs to be expanded, particularly given the finding that oligomeric rather than fibrillar amyloid morphologies are neurotoxic. This lack of knowledge is primarily due to the existence of transient oligomeric forms that require the use of non-traditional approaches capable of probing transiently existing amyloid forms. We have recently developed the Single-Molecule Force Spectroscopy (SMFS) approach enabling us to probe dimeric forms of amyloids. These studies suggest that the assembly of amyloid proteins into dimers leads to extremely stabilized amyloids in non-native, misfolded states [1]. Herein, we applied the SMFS approach to probe amyloid trimers. We used the Aβ(14-23) segment of Aβ42 protein that is responsible for full-size protein aggregation. The dimerization of this peptide was recently characterized [2]. The dimeric form of Aβ (14-23) was assembled by the use of a tandem Aβ(14-23)-YNGK-Aβ(14-23), in which the YNGK motif between the two Aβ(14-23) monomers makes a β turn to form a hairpin loop with an antiparallel arrangement of Aβ(14-23) monomers[3]. The Aβ(14-23) monomer was tethered to the AFM tip, and trimers were formed by approaching the tip to the mica surface on which the Aβ(14-23)-YNGK-Aβ(14-23) dimer was immobilized via a polyethylene glycol tether. We identified trimers by rupture forces that were considerably larger than those for dimers. Models for the trimer assembly process are discussed.

Oxidative stress is considered as an important factor and an early event in the etiology of Alzheimer's disease (AD). Cu bound to the peptide amyloid-β (Aβ) is found in AD brains, and Cu-Aβ could contribute to this oxidative stress, as it is able to produce in vitro H2O2 and HO˙ in the presence of oxygen and biological reducing agents such as ascorbate. The mechanism of Cu-Aβ-catalyzed H2O2 production is however not known, although it was proposed that H2O2 is directly formed from O2 via a 2-electron process. Here, we implement an electrochemical setup and use the specificity of superoxide dismutase-1 (SOD1) to show, for the first time, that H2O2 production by Cu-Aβ in the presence of ascorbate occurs mainly via a free O2˙(-) intermediate. This finding radically changes the view on the catalytic mechanism of H2O2 production by Cu-Aβ, and opens the possibility that Cu-Aβ-catalyzed O2˙(-) contributes to oxidative stress in AD, and hence may be of interest., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The growing interest for peptide therapeutics calls for new strategies to determine the physico-chemical properties responsible for the interactions of peptides with the environment. This study reports about the lipophilicity of two fragments of the amyloid β-peptide, Aβ 25-35 and Aβ 12-28. Firstly, computational studies showed the limits of log D(7.4)oct in describing the lipophilicity of medium-sized peptides. Chromatographic lipophilicity indexes (expressed as log k', the logarithm of the retention factor) were then measured in three different systems to highlight the different skills of Aβ 25-35 and Aβ 12-28 in giving interactions with polar and apolar environments. CD studies were also performed to validate chromatographic experimental conditions. Results show that Aβ 12-28 has a larger skill in promoting hydrophobic and electrostatic interactions than Aβ 25-35. This finding proposes a strategy to determine the lipophilicity of peptides for drug discovery purposes but also gives insights in unraveling the debate about the aminoacidic region of Aβ responsible for its neurotoxicity., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The plasma membrane, which encapsulates human cells, is composed of a complex mixture of lipids and embedded proteins. Emerging knowledge points towards the lipids as having a regulating role in protein function. Furthermore, insight from protein crystallography has revealed several different types of lipids intimately bound to membrane proteins and peptides, hereby possibly pointing to a site of action for the observed regulation. Cholesterol is among the lipid membrane constituents most often observed to be co-crystallized with membrane proteins, and the cholesterol levels in cell membranes have been found to play an essential role in health and disease. Remarkably little is known about the mechanism of lipid regulation of membrane protein function in health as well as in disease. Herein, we review molecular dynamics simulation studies aimed at investigating the effect of cholesterol on membrane protein and peptide properties. This article is part of a Special Issue entitled: Lipid-protein interactions., (Copyright © 2015. Published by Elsevier B.V.)

Neurons have developed elaborate mechanisms for sorting of proteins to their destination in dendrites and axons as well as dynamic local trafficking. Recent evidence suggests that polarized axonal sorting of β-site converting enzyme 1 (BACE1), a type I transmembrane aspartyl protease involved in Alzheimer's disease (AD) pathogenesis, entails an unusual journey. In hippocampal neurons, BACE1 internalized from dendrites is conveyed in recycling endosomes via unidirectional retrograde transport towards the soma and sorted to axons where BACE1 becomes enriched. In comparison to other transmembrane proteins that undergo transcytosis or elimination in somatodendritic compartment, vectorial transport of internalized BACE1 in dendrites is unique and intriguing. Dysfunction of protein transport contributes to pathogenesis of AD and other neurodegenerative diseases. Therefore, characterization of BACE1 transcytosis is an important addition to the multiple lines of evidence that highlight the crucial role played by endosomal trafficking pathway as well as axonal sorting mechanisms in AD pathogenesis., (© 2015 WILEY Periodicals, Inc.)

Protein misfolding and abnormal assembly could lead to aggregates such as oligomer, proto-fibril, mature fibril, and senior amyloid plaques, which are associated with the pathogenesis of many amyloid diseases. These irreversible amyloid aggregates typically form in vivo and researchers have been endeavoring to find new modulators to invert the aggregation propensity in vitro, which could increase understanding in the mechanism of the aggregation of amyloid protein and pave the way to potential clinical treatment. Graphene oxide (GO) was shown to be a good modulator, which could strongly control the amyloidosis of Aβ (33-42). In particular, quartz crystal microbalance (QCM), circular dichroism (CD) spectroscopy, and atomic force microscopy (AFM) measurements revealed the size-dependent manner of GO on modulating the assembly of amyloid peptides, which could be a possible way to regulate the self-assembled nanostructure of amyloid peptide in a predictable manner., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Mast cells (MCs) store an array of proinflammatory mediators in secretory granules that are rapidly released upon activation by diverse conditions including amyloid beta (Aβ) peptides. In the present work, we found a rapid degranulation of cultured MCs through a pannexin1 hemichannel (Panx1 HC)-dependent mechanism induced by Aβ25-35 peptide. Accordingly, Aβ25-35 peptide also increased membrane current and permeability, as well as intracellular Ca(2+) signal, mainly via Panx1 HCs because all of these responses were drastically inhibited by Panx1 HC blockers and absent in the MCs of Panx1(-/-) mice. Moreover, in acute coronal brain slices of control mice, Aβ25-35 peptide promoted both connexin 43 (Cx43)- and Panx1 HC-dependent MC dye uptake and histamine release, responses that were only Cx43 HC dependent in Panx1(-/-) mice. Because MCs have been found close to amyloid plaques of patients with Alzheimer's disease (AD), their distribution in brain slices of APPswe/PS1dE9 mice, a murine model of AD, was also investigated. The number of MCs in hippocampal and cortical areas increased drastically even before amyloid plaque deposits became evident. Therefore, MCs might act as early sensors of amyloid peptide and recruit other cells to the neuroinflammatory response, thus playing a critical role in the onset and progression of AD., (Copyright © 2015 the authors 0270-6474/15/359526-13$15.00/0.)

In the past years the self-assembly of amyloid-like peptides has attracted increasing attentions, because it is highly related to neurodegenerative diseases and has a potential for serving as nanomaterial to fabricate novel and useful nanostructures. In this paper, we focused on the role of interfacial conditions in the self-assembly of an amyloid-like peptide, termed Pep11. It was found that, when dissolved in bulk solutions, Pep11 formed into β-sheet structures and assembled into long filaments in several hours, as revealed by Thioflavin T fluorescence and transmission electron microscopy (TEM) morphology characterization, respectively. When the peptide solution was added onto a mica/HOPG substrate, peptide filaments with three preferred orientations with an angle of 60° to each other were formed immediately, as imaged in situ by atomic force microscopy (AFM). However, the kinetics in filament formation and the morphologies of the formed beta sheet either on HOPG and mica or in bulk solutions were quite different. These results indicate that the interfacial properties dramatically affect the peptide self-assembly process., (© 2015 Wiley Periodicals, Inc.)

In order to confirm the neurotoxicity of bifunctional chelators containing hydrophobic groups and metal chelating moiety, the interaction of di(picolyl)amine (dpa) derivatives toward Aβ42 peptide was investigated. Fluorescence titration reveals that a hydrophobic chelator (such as BODIPY) shows high binding affinity to amyloid Aβ42. Circular dichroism (CD) spectra confirm that the hydrophobic bifunctional chelator can decrease α-helix fraction and increase the β-sheet fraction of amyloid Aβ42. In particular, experimental results indicate that a bifunctional chelator can assemble with Cu(II)-Aβ42 forming chelator-Cu(II)-Aβ42 nanospheres, which are toxic to SH-S5Y5 cells. The hydrophobic interaction between the chelator and the amyloid peptide (Aβ42) has great contribution to the formation of neurotoxic chelator-Cu(II)-Aβ42 nanospheres. This work gives a general guide to the development of low cytotoxic inhibitors of Aβ42 aggregation., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Several commercial immobilized metal affinity chromatography sorbents were evaluated in this study for the analysis of two small peptide fragments of the amyloid β-protein (Aβ) (Aβ(1-15) and Aβ(10-20) peptides) by on-line immobilized metal affinity SPE-CE (IMA-SPE-CE). The performance of a nickel metal ion (Ni(II)) sorbent based on nitrilotriacetic acid as a chelating agent was significantly better than two copper metal ion (Cu(II)) sorbents based on iminodiacetic acid. A BGE of 25 mM phosphate (pH 7.4) and an eluent of 50 mM imidazole (in BGE) yielded a 25-fold and 5-fold decrease in the LODs by IMA-SPE-CE-UV for Aβ(1-15) and Aβ(10-20) peptides (0.1 and 0.5 μg/mL, respectively) with regard to CE-UV (2.5 μg/mL for both peptides). The phosphate BGE was also used in IMA-SPE-CE-MS, but the eluent needed to be substituted by a 0.5% HAc v/v solution. Under optimum preconcentration and detection conditions, reproducibility of peak areas and migration times was acceptable (23.2 and 12.0%RSD, respectively). The method was more sensitive for Aβ(10-20) peptide, which could be detected until 0.25 μg/mL. Linearity for Aβ(10-20) peptide was good in a narrow concentration range (0.25-2.5 μg/mL, R(2) = 0.93). Lastly, the potential of the optimized Ni(II)-IMA-SPE-CE-MS method for the analysis of amyloid peptides in biological fluids was evaluated by analyzing spiked plasma and serum samples., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Although the N-terminal region of Amyloid β (Aβ) peptides plays dual roles as metal-coordinating sites and conformational modulator, few studies have been performed to explore the effects of mutations at this region on the overall conformational ensemble of Aβ and the binding propensity of metal ions. In this work, we focus on how three familial Alzheimer's disease mutations (D7H, D7N, and H6R) alter the structural characteristics and thermodynamic stabilities of Aβ42 using molecular dynamics simulations. We observe that each mutation displays increased β-sheet structures in both N and C termini. In particular, both the N terminus and central hydrophobic region of D7H can form stable β-hairpin structures with its C terminus. The conserved turn structure at Val²⁴-Lys²⁸ in all peptides and Zn²⁺-bound Aβ42 is confirmed as the common structural motif to nucleate folding of Aβ. Each mutant can significantly increase the solvation free energy and thus enhance the aggregation of Aβ monomers. The correlation dynamics between Aβ(1-16) and Aβ(17-42) fragments are elucidated by linking the domain motions with the corresponding structured conformations. We characterize the different populations of correlated domain motions for each mutant from a more macroscopic perspective, and unexpectedly find that Zn²⁺-bound Aβ42 ensemble shares the same populations as Aβ42, indicating that the binding of Zn²⁺ to Aβ follows the conformational selection mechanism, and thus is independent of domain motions, even though the structures of Aβ have been modified at a residue level., (© 2014 Wiley Periodicals, Inc.)

The β-amyloid peptide (Aβ) (1–40) is one of the major components that form Alzheimer's amyloid deposits. Studies of the membrane insertion of amyloid showed that amyloid is surface active and can insert into lipid monolayers [Ji et al. (2002). Biochemistry (Moscow), 67, 1283–1288; Ege & Lee (2004). Biophys. J. 87, 1732–1740]. The interaction between the peptide and a lipid monolayer or bilayer is critical to the understanding of the formation of amyloid peptide deposits on the membrane. In this paper, we have studied the structural transition of 1,2-dipalmitoyl-sn- glycero-3-phosphocholine (DPPC) multibilayers with the insertion of amyloid peptide 1–40 by grazing-incidence small-angle X-ray scattering at different bilayer hydration levels (changing the relatively humidity). We mixed the DPPC and amyloid peptide in an organic solvent at a 10 to 1 weight ratio, then cast it onto a silicon wafer to form the mixed lipid multibilayer film. In this study of the pure DPPC multibilayer film and the DPPC multibilayer film inserted with amyloid peptide, it was found that the hydration process was bimodal with a better-hydrated top layer and a less-hydrated bottom layer. The gel-to-ripple phase transition suffers a strong confinement effect due to the presence of the solid substrate. With the insertion of the amyloid peptide, the ripple phase of the membrane bilayers was suppressed at high humidity and the whole film can be swollen more uniformly at lower incubation time than the pure DPPC film supported on a silicon wafer. This means water vapor can penetrate more easily into the DPPC bilayers inserted with Aβ than into the pure DPPC bilayers. Amyloid peptides were found to form clusters in the bilayer and possess in-plane correlation. From analyzing the diffuse scattering around the Bragg peak in the lateral direction, the amyloid peptides are found to form clusters in the bilayer with a radius of about 9 nm. It is also estimated that the number of Aβ molecules in one cluster is about 12 and on average each Aβ molecule occupies an interface area of about 22 nm2. As the relative humidity exceeds about 94%, the Aβ clusters seem to develop an ordered structure with a spacing of about 300 Å. [ABSTRACT FROM AUTHOR]

Amyloid peptides are considered to be the main contributor for the membrane disruption related to the pathogenesis of degenerative diseases. The variation of amino acids at the carboxylic terminus of amyloid peptide has revealed significant effects on the modulation of abnormal assemblies of amyloid peptides. In this work, molecular binding agents were tethered to the C-terminus of β-amyloid peptide 1-42 (Aβ42). The molecular interaction between Aβ42 and molecule tethers was identified at single molecule level by using scanning tunneling microscopy (STM). The mechanistic insight into the feature variation of the self-assembly of Aβ42 peptide caused by molecular tethering at C-terminus was clearly revealed, which could appreciably affect the nucleation of amyloid peptide, thus reducing the membrane disruptions.

Abnormal elevation of amyloid β-peptide (Aβ) levels in the brain is the primary trigger for neuronal cell death specific to Alzheimer's disease (AD). It is now evident that Aβ levels in the brain are manipulable due to a dynamic equilibrium between its production from the amyloid precursor protein (APP) and removal by amyloid clearance proteins. Clearance can be either enzymic or non-enzymic (binding/transport proteins). Intriguingly several of the main amyloid-degrading enzymes (ADEs) are members of the M13 peptidase family (neprilysin (NEP), NEP2 and the endothelin converting enzymes (ECE-1 and -2)). A distinct metallopeptidase, insulin-degrading enzyme (IDE), also contributes to Aβ degradation in the brain. The ADE family currently embraces more than 20 members, both membrane-bound and soluble, and of differing cellular locations. NEP plays an important role in brain function terminating neuropeptide signals. Its decrease in specific brain areas with age or after hypoxia, ischaemia or stroke contribute significantly to the development of AD pathology. The recently discovered mechanism of epigenetic regulation of NEP (and other genes) by the APP intracellular domain (AICD) and its dependence on the cell type and APP isoform expression suggest possibilities for selective manipulation of NEP gene expression in neuronal cells. We have also observed that another amyloid-clearing protein, namely transthyretin (TTR), is also regulated in the neuronal cell by a mechanism similar to NEP. Dependence of amyloid clearance proteins on histone deacetylases and the ability of HDAC inhibitors to up-regulate their expression in the brain opens new avenues for developing preventive strategies in AD.

Despite genetic variation has the potential to arise new protein functions, spontaneous mutations usually destabilize the native fold. Misfolded proteins tend to form cytotoxic intracellular aggregates, decreasing cell fitness and leading to degenerative disorders in humans. Therefore, it is thought that selection against protein misfolding and aggregation constrains the evolution of protein sequences. However, obtaining experimental data to validate this hypothesis has been traditionally difficult. Here we exploit bacteria as a model organism to address this question. Using variants of the Alzheimer's related Aβ42 peptide designed to exhibit different in vivo aggregation propensities we show here that, in cell competition experiments, the most aggregation-prone variants are always purged out from the growing population. Flow cytometry analysis of cellular metabolism and viability demonstrates that this purifying effect responds to a clear correlation between physiological burden and intrinsic aggregation propensity. Interestingly, the fitness cost of aggregation appears to be associated with aggregation rates rather than with overall protein solubility. Accordingly, we show that, by reducing in vivo aggregation rates, the model osmolyte proline is able to buffer the metabolic impact of protein aggregation. Overall, our data provide experimental support for the role of toxic protein aggregation on the cell fitness landscape and the evolution of natural protein sequences., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Protein aggregation is linked to many pathological conditions, including several neurodegenerative diseases. The aggregation propensities of proteins are thought to be controlled to a large extent by the physicochemical properties encoded in the primary sequence. We have previously exploited a set of amyloid β peptide (Aβ42) variants exhibiting a continuous gradient of intrinsic aggregation propensities to demonstrate that this rule applies in vivo in bacteria. In the present work we have characterized the behavior of these Aβ42 mutants when expressed in yeast. In contrast to bacteria, the intrinsic aggregation propensity is gated by yeast, in such a way that this property correlates with the formation of intracellular inclusions only above a specific aggregation threshold. Proteins displaying solubility levels above this threshold escape the inclusion formation pathway. In addition, the most aggregation-prone variants are selectively cleared by the yeast quality control degradation machinery. Thus, both inclusion formation and proteolysis target the same aggregation-prone variants and cooperate to minimize the presence of these potentially dangerous species in the cytosol. The demonstration that sorting to these pathways in eukaryotes is strongly influenced by protein primary sequence should facilitate the development of rational approaches to predict and hopefully prevent in vivo protein deposition., (© 2013.)

Exercise has been shown to enhance hippocampus-related cognition and slow the progression of Alzheimer's disease (AD). However, whether voluntary exercise directly decreases the neurotoxicity of amyloid peptide (Aβ) needs to be determined. In the present study, two-month old male C57bl/6 mice were intracerebroventricularly injected with Aβ25-35, and then allowed for voluntary exercise for 12 days. Y-maze test revealed that voluntary exercise mitigated spatial memory impairment induced by Aβ25-35. Consistently, Aβ25-35 treated mice with exercise showed reduced neuronal degeneration and synaptic protein loss in the hippocampus compared with sedentary controls. Moreover, voluntary exercise significantly ameliorated oxidative stress markers and increased vessel branches in the hippocampus of Aβ25-35 treated mice. Our results suggest that voluntary exercise counteracts the neurotoxicity of Aβ by reducing oxidative stress and increasing angiogenesis, which may underlie the beneficial effect of exercise on AD., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Surface-assisted self-assembly of amyloid-like peptides has received considerable interest in both amyloidosis research and nanotechnology in recent years. Despite extensive studies, some controlling factors, such as salts, are still not well understood, even though it is known that some salts can promote peptide self-assemblies through the so-called "salting-out" effect. However, they are usually noncontrollable, disordered, amorphous aggregates. Here, we show via a combined experimental and theoretical approach that a conserved consensus peptide NH2-VGGAVVAGV-CONH2 (GAV-9) (from representative amyloidogenic proteins) can self-assemble into highly ordered, multilayered nanofilaments, with surprising all-upright conformations, under high-salt concentrations. Our atomic force microscopy images also demonstrate that the vertical stacking of multiple layers is highly controllable by tuning the ionic strength, such as from 0 mM (monolayer) to 100 mM (mainly double layer), and to 250 mM MgCl2 (double, triple, quadruple, and quintuple layers). Our atomistic molecular dynamics simulations then reveal that these individual layers have very different internal nanostructures, with parallel β-sheets in the first monolayer but antiparallel β-sheets in the subsequent upper layers due to their different microenvironment. Further studies show that the growth of multilayered, all-upright nanostructures is a common phenomenon for GAV-9 at the mica/water interface, under a variety of salt types and a wide range of salt concentrations.