scholarly journals Deletion of Serf2 shifts amyloid conformation in an Aβ amyloid mouse model

2021 ◽  
Author(s):  
E. Stroo ◽  
L. Janssen ◽  
O. Sin ◽  
W. Hogewerf ◽  
M. Koster ◽  
...  
Keyword(s):  
Mouse Model ◽  
Amyloid Fibrils ◽  
Mammalian Brain ◽  
Amyloid Formation ◽  
List Type ◽  
Lung Maturation ◽  
Disease Manifestation ◽  
Aβ Amyloid ◽  
Perinatal Lethality

AbstractNeurodegenerative diseases like Alzheimer, Parkinson and Huntington disease are characterized by aggregation-prone proteins that form amyloid fibrils through a nucleation process. Despite the shared β-sheet structure, recent research has shown that structurally different polymorphs exist within fibrils of the same protein. These polymorphs are associated with varying levels of toxicity and different disease phenotypes. MOAG-4 and its human orthologs SERF1 and SERF2 have previously been shown to modify the nucleation and drive amyloid formation and protein toxicity in vitro and in C. elegans. To further explore these findings, we generated a Serf2 knockout (KO) mouse model and crossed it with the APPPS1 mouse model for Aβ amyloid pathology. Full-body KO of Serf2 resulted in a developmental delay and perinatal lethality due to insufficient lung maturation. Therefore, we proceeded with a brain-specific Serf2 KO, which was found to be viable. We examined the Aβ pathology at 1 and 3 months of age, which is before and after the start of amyloid deposition. We show that SERF2 deficiency does not affect the production and overall Aβ levels. Serf2 KO-APPPS1 mice displayed an increased intracellular Aβ accumulation at 1 month and a higher number of Aβ deposits compared to APPPS1 mice with similar Aβ levels. Moreover, conformation-specific dyes and electron microscopy revealed a difference in the structure and amyloid content of these Aβ deposits. Together, our results reveal that SERF2 causes a structural shift in Aβ aggregation in a mammalian brain. These findings indicate that a single endogenous factor may contribute to amyloid polymorphisms, allowing for new insights into this phenomenon’s contribution to disease manifestation.HighlightsLoss of SERF2 slows embryonic development and causes perinatal lethalitySERF2 affects proliferation in a cell-autonomous fashionBrain-specific Serf2 knockout does not affect viability or Aβ productionBrain deletion of Serf2 shifts the amyloid conformation of Aβ

scholarly journals αB-crystallin affects the morphology of Aβ(1-40) aggregates

2021 ◽  
Author(s):  
Henrik Müller ◽  
David M. Dias ◽  
Anna van der Zalm ◽  
Andrew J. Baldwin
Keyword(s):  
Amyloid Fibrils ◽  
Recombinant Expression ◽  
Small Heat Shock Protein ◽  
Primary Component ◽  
Amyloid Formation ◽  
List Type ◽  
Native Sequence ◽  
Αb Crystallin ◽  
Aggregate Morphology

SummaryαB-crystallin (ABC) is a human small heat shock protein that is strongly linked to Alzheimer’s disease (AD). In vitro, it can inhibit the aggregation and amyloid formation of a range of proteins including Aβ(1-40), a primary component of AD amyloid plaques. Despite the strong links, the mechanism by which ABC inhibits amyloid formation has remained elusive, in part due to the notorious irreproducibility of aggregation assays involving preparations of Aβ-peptides of native sequence. Here, we present a recombinant expression protocol to produce native Aβ(1-40), devoid of any modifications or exogenous residues, with yields up to 4 mg/L E. coli. This material provides highly reproducible aggregation kinetics and, by varying the solution conditions, we obtain either highly ordered amyloid fibrils or more disordered aggregates. Addition of ABC slows the aggregation of Aβ(1-40), and interferes specifically with the formation of ordered amyloid fibrils, favouring instead the more disordered aggregates. Solution-state NMR spectroscopy reveals that the interaction of ABC with Aβ(1-40) depends on the specific aggregate morphology. These results provide mechanistic insight into how ABC inhibits the formation of amyloid fibrils.HighlightsProtocol for production of native recombinant Aβ(1-40)Amyloid formation under physiological conditions is highly reproducibleBoth ordered fibrils and disordered aggregates can be reliably formedαB-crystallin specifically inhibits amyloid fibril assembling, favouring disordered aggregateseTOC blurbMüller et al. introduce a protocol for the highly reproducible production of amyloid from native Aβ(1-40) and determine that the human chaperone ABC specifically destabilises them in favour of disordered aggregates. NMR shows that ABC can distinguish between aggregate morphologies.Graphical Abstract

scholarly journals Cryo-EM structure and polymorphism of Aβ amyloid fibrils purified from Alzheimer’s brain tissue

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Marius Kollmer ◽  
William Close ◽  
Leonie Funk ◽  
Jay Rasmussen ◽  
Aref Bsoul ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Brain Tissue ◽  
Amyloid Fibrils ◽  
Structural Basis ◽  
Amyloid Angiopathy ◽  
Cryo Electron Microscopy ◽  
Aβ Amyloid ◽  
Cerebral Amyloid ◽  
Aβ Fibrils

Abstract The formation of Aβ amyloid fibrils is a neuropathological hallmark of Alzheimer’s disease and cerebral amyloid angiopathy. However, the structure of Aβ amyloid fibrils from brain tissue is poorly understood. Here we report the purification of Aβ amyloid fibrils from meningeal Alzheimer’s brain tissue and their structural analysis with cryo-electron microscopy. We show that these fibrils are polymorphic but consist of similarly structured protofilaments. Brain derived Aβ amyloid fibrils are right-hand twisted and their peptide fold differs sharply from previously analyzed Aβ fibrils that were formed in vitro. These data underscore the importance to use patient-derived amyloid fibrils when investigating the structural basis of the disease.

scholarly journals Desmin forms toxic, seeding-competent amyloid aggregates that persist in muscle fibers

2019 ◽  
Vol 116 (34) ◽  
pp. 16835-16840 ◽  
Author(s):  
Niraja Kedia ◽  
Khalid Arhzaouy ◽  
Sara K. Pittman ◽  
Yuanzi Sun ◽  
Mark Batchelor ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Therapeutic Potential ◽  
Epigallocatechin Gallate ◽  
Amyloid Formation ◽  
Myofibrillar Myopathy ◽  
Protein Aggregate ◽  
Disease Mutations ◽  
Amyloid Aggregates ◽  
Associated Proteins

Desmin-associated myofibrillar myopathy (MFM) has pathologic similarities to neurodegeneration-associated protein aggregate diseases. Desmin is an abundant muscle-specific intermediate filament, and disease mutations lead to its aggregation in cells, animals, and patients. We reasoned that similar to neurodegeneration-associated proteins, desmin itself may form amyloid. Desmin peptides corresponding to putative amyloidogenic regions formed seeding-competent amyloid fibrils. Amyloid formation was increased when disease-associated mutations were made within the peptide, and this conversion was inhibited by the anti-amyloid compound epigallocatechin-gallate. Moreover, a purified desmin fragment (aa 117 to 348) containing both amyloidogenic regions formed amyloid fibrils under physiologic conditions. Desmin fragment-derived amyloid coaggregated with full-length desmin and was able to template its conversion into fibrils in vitro. Desmin amyloids were cytotoxic to myotubes and disrupted their myofibril organization compared with desmin monomer or other nondesmin amyloids. Finally, desmin fragment amyloid persisted when introduced into mouse skeletal muscle. These data suggest that desmin forms seeding-competent amyloid that is toxic to myofibers. Moreover, small molecules known to interfere with amyloid formation and propagation may have therapeutic potential in MFM.

scholarly journals Amyloid conformers of the FXR1 protein prevent mRNA degradation in cortical neurons

2019 ◽  
Author(s):  
J.V. Sopova ◽  
E. I Koshel ◽  
T.A. Belashova ◽  
S.P. Zadorsky ◽  
A.V. Sergeeva ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Large Scale ◽  
Amyloid Fibrils ◽  
Mammalian Species ◽  
Rnase A ◽  
Mammalian Brain ◽  
Polarization Microscopy ◽  
Proteomic Approach ◽  
Functional Amyloids

AbstractFunctional amyloids regulate vital processes in a variety of organisms from bacteria to higher eukaryotes. The development of methods enabling large-scale screening for amyloids opens up opportunity for systemic analysis of the prevalence of amyloids in nature. Using an original proteomic approach, we identified several proteins forming amyloid-like detergent-resistant aggregates in the rat brain. One of them is the FXR1 protein, which is known to regulate memory and emotions (1, 2). We demonstrated that in brain FXR1 forms amyloid oligomers and insoluble detergent-resistant aggregates that strongly colocalize with amyloid-specific dye Thioflavin S and bind mRNA molecules. Moreover, we demonstrated that mRNAs colocalized with FXR1 amyloid particles are completely resistant to treatment with RNAse A. Taking into consideration that the members of ribonuclease A superfamily function in neurons (3) we can conclude that amyloid conformers of FXR1 control RNA stability in brain. Thus, in contrast to pathological amyloids that cause neurodegeneration, FXR1 is the functional amyloid in forebrain. We showed that amyloid properties of FXR1 depend on its N-terminal part from 1 to 379 amino acids. This fragment forms amyloid fibrils in vitro that bind Congo red and manifest apple-green birefringence when assayed by polarization microscopy. The amyloid-forming region of FXR1 is highly conserved in mammals. These data suggest that the ability of amyloid conformers of FXR1 to protect mRNAs is characteristic of different mammalian species, including humans.Significance StatementAmyloids are highly ordered cross-β sheet protein fibrils associated with many neurodegenerative diseases including Alzheimer’s disease. However, some amyloid proteins regulate vital processes. We identified a set of proteins that form amyloid-like aggregates in the brain of healthy rats. One of them - the FXR1 protein is known to regulate memory and emotions. FXR1 forms amyloid fibrils that bind RNA molecules and prevent their degradation in brain cortex neurons. Amyloid-forming sequence of FXR1 is highly conserved across mammals including human. Discovery of functional amyloids in mammalian brain shows that strategy aimed at the development of universal anti-amyloid drugs is unpromising. Such potential drugs should prevent or suppress formation of pathological aggregates of a certain protein, but not affect functional amyloids.

scholarly journals Comparative study of the stabilities of synthetic in vitro and natural ex vivo transthyretin amyloid fibrils

2020 ◽  
Vol 295 (33) ◽  
pp. 11379-11387 ◽  
Author(s):  
Sara Raimondi ◽  
P. Patrizia Mangione ◽  
Guglielmo Verona ◽  
Diana Canetti ◽  
Paola Nocerino ◽  
...  
Keyword(s):  
Thermodynamic Stability ◽  
Amyloid Fibrils ◽  
Current Knowledge ◽  
Ex Vivo ◽  
Biochemical Properties ◽  
Amyloid Formation ◽  
Free Energies

Systemic amyloidosis caused by extracellular deposition of insoluble fibrils derived from the pathological aggregation of circulating proteins, such as transthyretin, is a severe and usually fatal condition. Elucidation of the molecular pathogenic mechanism of the disease and discovery of effective therapies still represents a challenging medical issue. The in vitro preparation of amyloid fibrils that exhibit structural and biochemical properties closely similar to those of natural fibrils is central to improving our understanding of the biophysical basis of amyloid formation in vivo and may offer an important tool for drug discovery. Here, we compared the morphology and thermodynamic stability of natural transthyretin fibrils with those of fibrils generated in vitro either using the common acidification procedure or primed by limited selective cleavage by plasmin. The free energies for fibril formation were −12.36, −8.10, and −10.61 kcal mol−1, respectively. The fibrils generated via plasmin cleavage were more stable than those prepared at low pH and were thermodynamically and morphologically similar to natural fibrils extracted from human amyloidotic tissue. Determination of thermodynamic stability is an important tool that is complementary to other methods of structural comparison between ex vivo fibrils and fibrils generated in vitro. Our finding that fibrils created via an in vitro amyloidogenic pathway are structurally similar to ex vivo human amyloid fibrils does not necessarily establish that the fibrillogenic pathway is the same for both, but it narrows the current knowledge gap between in vitro models and in vivo pathophysiology.

scholarly journals Designing transthyretin mutants affecting tetrameric structure: implications in amyloidogenicity

2000 ◽  
Vol 348 (1) ◽  
pp. 167-172 ◽  
Author(s):  
Clara REDONDO ◽  
Ana M. DAMAS ◽  
Maria João M. SARAIVA
Keyword(s):  
Molecular Mechanisms ◽  
Amyloid Fibrils ◽  
Hydrophobic Interactions ◽  
Fibril Formation ◽  
Amyloid Formation ◽  
Structural Determinants ◽  
Tetrameric Structure ◽  
Structural Subunit ◽  
The Stability

The molecular mechanisms that convert soluble transthyretin (TTR) tetramers into insoluble amyloid fibrils are still unknown; dissociation of the TTR tetramer is a pre-requisite for amyloid formation in vitro and involvement of monomers and/or dimers in fibril formation has been suggested by structural studies. We have designed four mutated proteins with the purpose of stabilizing [Ser117 → Cys (S117C) and Glu92 → Cys (E92C)] or destabilizing [Asp18 → Asn (D18N) and Leu110 → Ala (D110A)] the dimer/tetramer interactions in TTR, aiming at elucidating structural determinants in amyloidogenesis. The resistance of the mutated proteins to dissociation was analysed by HPLC studies of diluted TTR preparations. Both ‘stabilized’ mutants migrated as tetramers and, upon dilution, no other TTR species was observed, confirming the increased resistance to dissociation. For the ‘destabilized’ mutants, a mixture of tetrameric and monomeric forms co-existed at low dilution and the latter increased upon 10-fold dilution. Both of the destabilizing mutants formed amyloid in vitro when acidified. This result indicated that both the AB loop of TTR, destabilized in D18N, and the hydrophobic interactions affecting the dimer-dimer interfaces in L110A are implicated in the stability of the tetrameric structure. The stabilized mutants, which were dimeric in nature through disulphide bonding, were unable to polymerize into amyloid, even at pH 3.2. When the amyloid formation assay was repeated in the presence of 2-mercaptoethanol, upon disruption of the S-S bridges of these stable dimers, amyloid fibril formation was observed. This experimental evidence suggests that monomers, rather than dimers, are the repeating structural subunit comprising the amyloid fibrils.

scholarly journals Barriers to Small Molecule Drug Discovery for Systemic Amyloidosis

Molecules ◽  
2021 ◽  
Vol 26 (12) ◽  
pp. 3571
Author(s):  
Gareth J. Morgan
Keyword(s):  
Small Molecules ◽  
Light Chain ◽  
Amyloid Fibrils ◽  
Systemic Amyloidosis ◽  
Light Chains ◽  
Al Amyloidosis ◽  
Amyloid Formation ◽  
Amyloid Fibril Formation ◽  
Amyloid Light Chain

Inhibition of amyloid fibril formation could benefit patients with systemic amyloidosis. In this group of diseases, deposition of amyloid fibrils derived from normally soluble proteins leads to progressive tissue damage and organ failure. Amyloid formation is a complex process, where several individual steps could be targeted. Several small molecules have been proposed as inhibitors of amyloid formation. However, the exact mechanism of action for a molecule is often not known, which impedes medicinal chemistry efforts to develop more potent molecules. Furthermore, commonly used assays are prone to artifacts that must be controlled for. Here, potential mechanisms by which small molecules could inhibit aggregation of immunoglobulin light-chain dimers, the precursor proteins for amyloid light-chain (AL) amyloidosis, are studied in assays that recapitulate different aspects of amyloidogenesis in vitro. One molecule reduced unfolding-coupled proteolysis of light chains, but no molecules inhibited aggregation of light chains or disrupted pre-formed amyloid fibrils. This work demonstrates the challenges associated with drug development for amyloidosis, but also highlights the potential to combine therapies that target different aspects of amyloidosis.

scholarly journals Conjugates of neuroprotective chaperone L-PGDS provide MRI contrast for detection of amyloid β-rich regions in live Alzheimer’s Disease mouse model brain

2020 ◽  
Author(s):  
Bhargy Sharma ◽  
Joanes Grandjean ◽  
Margaret Phillips ◽  
Ambrish Kumar ◽  
Francesca Mandino ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Mouse Model ◽  
Mouse Brain ◽  
Amyloid Fibrils ◽  
Amyloid Β ◽  
Brain Regions ◽  
Mri Contrast ◽  
Neuroprotective Function

AbstractEndogenous brain proteins can recognize the toxic oligomers of amyloid-β (Aβ) peptides implicated in Alzheimer’s disease (AD) and interact with them to prevent their aggregation. Lipocalin-type Prostaglandin D Synthase (L-PGDS) is a major Aβ-chaperone protein in the human cerebrospinal fluid. Here we demonstrate that L-PGDS detects amyloids in diseased mouse brain. Conjugation of L-PGDS with magnetic nanoparticles enhanced the contrast for magnetic resonance imaging. We conjugated the L-PGDS protein with ferritin nanocages to detect amyloids in the AD mouse model brain. We show here that the conjugates administered through intraventricular injections co-localize with amyloids in the mouse brain. These conjugates can target the brain regions through non-invasive intranasal administration, as shown in healthy mice. These conjugates can inhibit the aggregation of amyloids in vitro and show potential neuroprotective function by breaking down the mature amyloid fibrils.

scholarly journals Cholinesterase-inhibiting Potential and Anti-BACE1 Activities of Edible Leaves of Selected Thai Local Plants

2019 ◽  
Vol 16 (3) ◽  
pp. 155-163
Author(s):  
Uthaiwan SUTTISANSANEE ◽  
Kalyarat KRUAWAN
Keyword(s):  
Amyloid Fibrils ◽  
The Elderly ◽  
Amyloid Formation ◽  
Key Enzymes ◽  
Brain Functions ◽  
Polarity Index ◽  
Alzhiemer's Disease ◽  
Local Plants ◽  
Garcinia Cowa

Alzhiemer’s disease (AD) is common amongst the elderly and is associated with decline in brain functions in terms of memory and cognitive loss. The causes of the disease may occur through loss of presynaptic markers of cholinergic system and deposition of amyloid fibrils in the brain. Cholinesterases (ChEs) including acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are the key enzymes controlling degradation of neurotransmitters, acetylcholines (AChs), in cholinergic hypothesis. Whereas overproduction of b-secretase (BACE1) can generate insoluble b-amyloid peptides. Thus, retardation on enzyme reactions can lead to potential AD prevention. The aim of this research was to investigate in vitro anti-AD activity through key enzymes inhibitions from Thai local plants with edible sour leaves, including Garcinia cowa Roxb., Spondias pinnata (Linn.f.) Kurz, Syzygium gratum (Wight) S.N. Mitra., Tamarind indica L. and Cratoxylum formosum (Jack) Dyer. Leaves were extracted in organic solvents with different polarity index values (ethanol and hexane). As results, all plants possessed different degrees of anti-ChEs activity, in which ethanolic extracts of Spondias pinnata and Tamarind indica exhibited significantly higher ChEs inhibitory activities than Syzygium gratum, Garcinia cowa and Cratoxylum formosum, respectively. Interestingly, most hexane extracts exhibited higher anti-AChE activities than ethanol extracts, while the contrary results were observed in anti-BChE activity. Besides, only Cratoxylum formosum, Garcinia cowa and Tamarind indica extracts possessed anti-BACE1 activity. The information received from this study would be great support of future drug development or nutraceutical agents against AD occurrence regarding its cholinergic and b-amyloid formation hypotheses.

scholarly journals Amyloid formation of fish β-parvalbumin involves primary nucleation triggered by disulfide-bridged protein dimers

2020 ◽  
Vol 117 (45) ◽  
pp. 27997-28004
Author(s):  
Tony E. R. Werner ◽  
David Bernson ◽  
Elin K. Esbjörner ◽  
Sandra Rocha ◽  
Pernilla Wittung-Stafshede
Keyword(s):  
Disulfide Bonds ◽  
Amyloid Fibrils ◽  
Kinetic Mechanism ◽  
Amyloid Formation ◽  
Structural Conversion ◽  
Aggregation Mechanism ◽  
Primary Nucleation ◽  
Protein Dimers ◽  
Induced Aggregation

Amyloid formation involves the conversion of soluble protein species to an aggregated state. Amyloid fibrils of β-parvalbumin, a protein abundant in fish, act as an allergen but also inhibit the in vitro assembly of the Parkinson protein α-synuclein. However, the intrinsic aggregation mechanism of β-parvalbumin has not yet been elucidated. We performed biophysical experiments in combination with mathematical modeling of aggregation kinetics and discovered that the aggregation of β-parvalbumin is initiated by the formation of dimers stabilized by disulfide bonds and then proceeds via primary nucleation and fibril elongation processes. Dimer formation is accelerated by H2O2and hindered by reducing agents, resulting in faster and slower aggregation rates, respectively. Purified β-parvalbumin dimers readily assemble into amyloid fibrils with similar morphology as those formed when starting from monomer solutions. Furthermore, addition of preformed dimers accelerates the aggregation reaction of monomers. Aggregation of purified β-parvalbumin dimers follows the same kinetic mechanism as that of monomers, implying that the rate-limiting primary nucleus is larger than a dimer and/or involves structural conversion. Our findings demonstrate a folded protein system in which spontaneously formed intermolecular disulfide bonds initiate amyloid fibril formation by recruitment of monomers. This dimer-induced aggregation mechanism may be of relevance for human amyloid diseases in which oxidative stress is often an associated hallmark.

Sign in / Sign up

Export Citation Format

Share Document