scholarly journals Structure and Aggregation Mechanisms in Amyloids

Molecules ◽  
2020 ◽  
Vol 25 (5) ◽  
pp. 1195 ◽  
Author(s):  
Zaida L. Almeida ◽  
Rui M. M. Brito
Keyword(s):  
Molecular Species ◽  
Molecular Mechanisms ◽  
Amyloid Fibrils ◽  
Current Knowledge ◽  
Point Of View ◽  
Resonance Spectroscopy ◽  
Amyloid Formation ◽  
Amyloid Fibril Formation ◽  
Misfolding Diseases ◽  
Amyloid Cascade

The aggregation of a polypeptide chain into amyloid fibrils and their accumulation and deposition into insoluble plaques and intracellular inclusions is the hallmark of several misfolding diseases known as amyloidoses. Alzheimer′s, Parkinson′s and Huntington’s diseases are some of the approximately 50 amyloid diseases described to date. The identification and characterization of the molecular species critical for amyloid formation and disease development have been the focus of intense scrutiny. Methods such as X-ray and electron diffraction, solid-state nuclear magnetic resonance spectroscopy (ssNMR) and cryo-electron microscopy (cryo-EM) have been extensively used and they have contributed to shed a new light onto the structure of amyloid, revealing a multiplicity of polymorphic structures that generally fit the cross-β amyloid motif. The development of rational therapeutic approaches against these debilitating and increasingly frequent misfolding diseases requires a thorough understanding of the molecular mechanisms underlying the amyloid cascade. Here, we review the current knowledge on amyloid fibril formation for several proteins and peptides from a kinetic and thermodynamic point of view, the structure of the molecular species involved in the amyloidogenic process, and the origin of their cytotoxicity.

scholarly journals Identification of Novel 1,3,5-Triphenylbenzene Derivative Compounds as Inhibitors of Hen Lysozyme Amyloid Fibril Formation

2019 ◽  
Vol 20 (22) ◽  
pp. 5558
Author(s):  
Hassan Ramshini ◽  
Reza Tayebee ◽  
Alessandra Bigi ◽  
Francesco Bemporad ◽  
Cristina Cecchi ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Inhibitory Effect ◽  
Fibril Formation ◽  
Amyloid Formation ◽  
Binding Assays ◽  
Egg White Lysozyme ◽  
Amyloid Fibril Formation ◽  
Activity Measurements ◽  
Model Protein

Deposition of soluble proteins as insoluble amyloid fibrils is associated with a number of pathological states. There is a growing interest in the identification of small molecules that can prevent proteins from undergoing amyloid fibril formation. In the present study, a series of small aromatic compounds with different substitutions of 1,3,5-triphenylbenzene have been synthesized and their possible effects on amyloid fibril formation by hen egg white lysozyme (HEWL), a model protein for amyloid formation, and of their resulting toxicity were examined. The inhibitory effect of the compounds against HEWL amyloid formation was analyzed using thioflavin T and Congo red binding assays, atomic force microscopy, Fourier-transform infrared spectroscopy, and cytotoxicity assays, such as the 3-(4,5-Dimethylthiazol)-2,5-Diphenyltetrazolium Bromide (MTT) reduction assay and caspase-3 activity measurements. We found that all compounds in our screen were efficient inhibitors of HEWL fibril formation and their associated toxicity. We showed that electron-withdrawing substituents such as –F and –NO2 potentiated the inhibitory potential of 1,3,5-triphenylbenzene, whereas electron-donating groups such as –OH, –OCH3, and –CH3 lowered it. These results may ultimately find applications in the development of potential inhibitors against amyloid fibril formation and its biologically adverse effects.

Sensing and modulation of amyloid fibrils by photo-switchable organic dots

Nanoscale ◽  
2020 ◽  
Vol 12 (32) ◽  
pp. 16805-16818
Author(s):  
Aslam Uddin ◽  
Bibhisan Roy ◽  
Gregor P. Jose ◽  
Sk Saddam Hossain ◽  
Partha Hazra
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Early Stage ◽  
Fibril Formation ◽  
Amyloid Formation ◽  
Amyloid Fibril Formation

Our study demonstrates that organic dots can be used for the imaging and early stage detection of amyloid fibril formation and the modulation of amyloid formation pathways.

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 Inhibition of peptide aggregation by means of enzymatic phosphorylation

2016 ◽  
Vol 12 ◽  
pp. 2462-2470 ◽  
Author(s):  
Kristin Folmert ◽  
Malgorzata Broncel ◽  
Hans v. Berlepsch ◽  
Christopher Hans Ullrich ◽  
Mary-Ann Siegert ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Conformational Transition ◽  
Structural Characteristics ◽  
Random Coil ◽  
Amyloid Formation ◽  
Initial Growth ◽  
Amyloid Fibril Formation ◽  
Dependent Protein ◽  
Enzymatic Phosphorylation ◽  
Β Sheet

As is the case in numerous natural processes, enzymatic phosphorylation can be used in the laboratory to influence the conformational populations of proteins. In nature, this information is used for signal transduction or energy transfer, but has also been shown to play an important role in many diseases like tauopathies or diabetes. With the goal of determining the effect of phosphorylation on amyloid fibril formation, we designed a model peptide which combines structural characteristics of α-helical coiled-coils and β-sheets in one sequence. This peptide undergoes a conformational transition from soluble structures into insoluble amyloid fibrils over time and under physiological conditions and contains a recognition motif for PKA (cAMP-dependent protein kinase) that enables enzymatic phosphorylation. We have analyzed the pathway of amyloid formation and the influence of enzymatic phosphorylation on the different states along the conformational transition from random-coil to β-sheet-rich oligomers to protofilaments and on to insoluble amyloid fibrils, and we found a remarkable directing effect from β-sheet-rich structures to unfolded structures in the initial growth phase, in which small oligomers and protofilaments prevail if the peptide is phosphorylated.

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 Exploring the sequence–structure relationship for amyloid peptides

2013 ◽  
Vol 450 (2) ◽  
pp. 275-283 ◽  
Author(s):  
Kyle L. Morris ◽  
Alison Rodger ◽  
Matthew R. Hicks ◽  
Maya Debulpaep ◽  
Joost Schymkowitz ◽  
...  
Keyword(s):  
Self Assembly ◽  
Amyloid Fibrils ◽  
Linear Dichroism ◽  
Model Systems ◽  
Molecular Architecture ◽  
Amyloid Peptides ◽  
Amyloid Fibril Formation ◽  
Structure Relationship ◽  
Fibre Diffraction ◽  
Misfolding Diseases

Amyloid fibril formation is associated with misfolding diseases, as well as fulfilling a functional role. The cross-β molecular architecture has been reported in increasing numbers of amyloid-like fibrillar systems. The Waltz algorithm is able to predict ordered self-assembly of amyloidogenic peptides by taking into account the residue type and position. This algorithm has expanded the amyloid sequence space, and in the present study we characterize the structures of amyloid-like fibrils formed by three peptides identified by Waltz that form fibrils but not crystals. The structural challenge is met by combining electron microscopy, linear dichroism, CD and X-ray fibre diffraction. We propose structures that reveal a cross-β conformation with ‘steric-zipper’ features, giving insights into the role for side chains in peptide packing and stability within fibrils. The amenity of these peptides to structural characterization makes them compelling model systems to use for understanding the relationship between sequence, self-assembly, stability and structure of 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 The Kinetics of Amyloid Fibril Formation by de Novo Protein Albebetin and Its Mutant Variants

Biomolecules ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 241
Author(s):  
Vitalii Balobanov ◽  
Rita Chertkova ◽  
Anna Egorova ◽  
Dmitry Dolgikh ◽  
Valentina Bychkova ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
De Novo ◽  
Point Mutations ◽  
Inhibitory Effect ◽  
Structural Basis ◽  
Amyloid Formation ◽  
Amyloid Fibril Formation ◽  
External Conditions ◽  
New Perspective ◽  
Kinetics Of

Engineering of amyloid structures is one of the new perspective areas of protein engineering. Studying the process of amyloid formation can help find ways to manage it in the interests of medicine and biotechnology. One of the promising candidates for the structural basis of artificial functional amyloid fibrils is albebetin (ABB), an artificial protein engineered under the leadership of O.B. Ptitsyn. Various aspects of the amyloid formation of this protein and some methods for controlling this process are investigated in this paper. Four stages of amyloid fibrils formation by this protein from the first non-fibrillar aggregates to mature fibrils and large micron-sized complexes have been described in detail. Dependence of albebetin amyloids formation on external conditions and some mutations also have been described. The introduction of similar point mutations in the two structurally identical α-β-β motifs of ABB lead to different amiloidogenesis kinetics. The inhibitory effect of a disulfide bond and high pH on amyloid fibrils formation, that can be used to control this process, was shown. The results of this work are a good basis for the further design and use of ABB-based amyloid constructs.

scholarly journals The Aggregation Conditions Define Whether EGCG is an Inhibitor or Enhancer of α-Synuclein Amyloid Fibril Formation

2020 ◽  
Vol 21 (6) ◽  
pp. 1995 ◽  
Author(s):  
Rebecca Sternke-Hoffmann ◽  
Alessia Peduzzo ◽  
Najoua Bolakhrif ◽  
Rainer Haas ◽  
Alexander K. Buell
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Epigallocatechin Gallate ◽  
Small Region ◽  
Fibril Formation ◽  
Oxidation Products ◽  
Relevant Parameter ◽  
Amyloid Formation ◽  
Experimental Conditions ◽  
Amyloid Fibril Formation

The amyloid fibril formation by α -synuclein is a hallmark of various neurodegenerative disorders, most notably Parkinson’s disease. Epigallocatechin gallate (EGCG) has been reported to be an efficient inhibitor of amyloid formation by numerous proteins, among them α -synuclein. Here, we show that this applies only to a small region of the relevant parameter space, in particular to solution conditions where EGCG readily oxidizes, and we find that the oxidation product is a much more potent inhibitor compared to the unmodified EGCG. In addition to its inhibitory effects, EGCG and its oxidation products can under some conditions even accelerate α -synuclein amyloid fibril formation through facilitating its heterogeneous primary nucleation. Furthermore, we show through quantitative seeding experiments that, contrary to previous reports, EGCG is not able to re-model α -synuclein amyloid fibrils into seeding-incompetent structures. Taken together, our results paint a complex picture of EGCG as a compound that can under some conditions inhibit the amyloid fibril formation of α -synuclein, but the inhibitory action is not robust against various physiologically relevant changes in experimental conditions. Our results are important for the development of strategies to identify and characterize promising amyloid inhibitors.

scholarly journals Amyloid formation of bovine insulin is retarded in moderately acidic pH and by addition of short-chain alcohols

2020 ◽  
Vol 49 (2) ◽  
pp. 145-153 ◽  
Author(s):  
David Bernson ◽  
Almedina Mecinovic ◽  
Md Tuhin Abed ◽  
Fredrik Limé ◽  
Per Jageland ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Current Knowledge ◽  
Solvent Polarity ◽  
Bovine Insulin ◽  
Short Chain ◽  
Amyloid Formation ◽  
Wide Ph Range ◽  
Model Protein ◽  
Ph Range ◽  
Simplistic Model

AbstractProtein aggregation and amyloid formation are associated with multiple human diseases, but are also a problem in protein production. Understanding how aggregation can be modulated is therefore of importance in both medical and industrial contexts. We have used bovine insulin as a model protein to explore how amyloid formation is affected by buffer pH and by the addition of short-chain alcohols. We find that bovine insulin forms amyloid fibrils, albeit with different rates and resulting fibril morphologies, across a wide pH range (2–7). At pH 4.0, bovine insulin displayed relatively low aggregation propensity in combination with high solubility; this condition was therefore chosen as basis for further exploration of how bovine insulin’s native state can be stabilized in the presence of short-chain alcohols that are relevant because of their common use as eluents in industrial-scale chromatography purification. We found that ethanol and isopropanol are efficient modulators of bovine insulin aggregation, providing a three to four times retardation of the aggregation kinetics at 30–35% (vol/vol) concentration; we attribute this to the formation of oligomers, which we detected by AFM. We discuss this effect in terms of reduced solvent polarity and show, by circular dichroism recordings, that a concomitant change in α-helical packing of the insulin monomer occurs in ethanol. Our results extend current knowledge of how insulin aggregates, and may, although bovine insulin serves as a simplistic model, provide insights into how buffers and additives can be fine-tuned in industrial production of proteins in general and pharmaceutical insulin in particular.

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