scholarly journals Exploring the occurrence of thioflavin-T-positive insulin amyloid aggregation intermediates

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e10918
Author(s):  
Mantas Ziaunys ◽  
Andrius Sakalauskas ◽  
Kamile Mikalauskaite ◽  
Vytautas Smirnovas
Keyword(s):  
Kinetic Data ◽  
Thioflavin T ◽  
Aggregation Kinetics ◽  
Aggregation Process ◽  
Binding Properties ◽  
Aggregation Kinetic ◽  
Fibril Structure ◽  
Random Occurrence ◽  
Amyloidogenic Protein ◽  
Sigmoidal Kinetics

The aggregation of proteins is considered to be the main cause of several neurodegenerative diseases. Despite much progress in amyloid research, the process of fibrillization is still not fully understood, which is one of the main reasons why there are still very few effective treatments available. When the aggregation of insulin, a model amyloidogenic protein, is tracked using thioflavin-T (ThT), an amyloid specific dye, there is an anomalous occurrence of double-sigmoidal aggregation kinetics. Such an event is likely related to the formation of ThT-positive intermediates, which may affect the outcome of both aggregation kinetic data, as well as final fibril structure. In this work we explore insulin fibrillization under conditions, where both normal and double-sigmoidal kinetics are observed and show that, despite their dye-binding properties and random occurrence, the ThT-positive intermediates do not significantly alter the overall aggregation process.

Binding modes of thioflavin T and Congo red to the fibril structure of amyloid-β(1–42)

2020 ◽  
Vol 56 (55) ◽  
pp. 7589-7592 ◽  
Author(s):  
Benedikt Frieg ◽  
Lothar Gremer ◽  
Henrike Heise ◽  
Dieter Willbold ◽  
Holger Gohlke
Keyword(s):  
Congo Red ◽  
Fluorescent Dyes ◽  
Binding Free Energy ◽  
Amyloid Β ◽  
Free Energy Calculations ◽  
Thioflavin T ◽  
Binding Modes ◽  
Fibril Structure ◽  
Binding Free Energy Calculations ◽  
Dynamics Simulations

Binding modes for the amyloid-β(1–42) fibril fluorescent dyes thioflavin T and Congo red were predicted by molecular dynamics simulations and binding free energy calculations.

Simulation on the aggregation process of spherical particle confined in a spherical shell

2016 ◽  
Vol 30 (11) ◽  
pp. 1650065
Author(s):  
J. Wang ◽  
J. J. Xu ◽  
L. Zhang
Keyword(s):  
Spherical Shell ◽  
Density Distribution ◽  
Volume Fraction ◽  
Particle Volume Fraction ◽  
Porous Polymer ◽  
Spherical Particles ◽  
Aggregation Kinetics ◽  
Aggregation Process ◽  
Particle Volume ◽  
Aggregate Structure

The aggregation process of spherical particles confined in a spherical shell was studied by using a diffusion-limited cluster–cluster aggregation (DLCA) model. The influence of geometrical confinement and wetting-like properties of the spherical shell walls on the particle concentration profile, aggregate structure and aggregation kinetics had been explored. The results show that there will be either depletion or absorption particles near the shell walls depending on the wall properties. It is observed that there are four different types of density distribution which can be realized by modifying the property of the inner or outer spherical shell wall. In addition, the aggregate structure will become more compact in the confined spherical shell comparing to bulk system with the same particle volume fraction. The analysis on the aggregation kinetics indicates that geometrical confinement will promote the aggregation process by reducing the invalid movement of the small aggregates and by constraining the movement of those large aggregates. Due to the concave geometrical characteristic of the outer wall of the spherical shell, its effects on the aggregating kinetics and the structure of the formed aggregates are more evident than those of the inner wall. This study will provide some instructive information of controlling the density distribution of low-density porous polymer hollow spherical shells and helps to predict gel structures developed in confined geometries.

scholarly journals The N terminus of α-synuclein dictates fibril formation

2021 ◽  
Vol 118 (35) ◽  
pp. e2023487118
Author(s):  
Ryan P. McGlinchey ◽  
Xiaodan Ni ◽  
Jared A. Shadish ◽  
Jiansen Jiang ◽  
Jennifer C. Lee
Keyword(s):  
Conformational Changes ◽  
Salt Bridge ◽  
Vital Role ◽  
Fibril Formation ◽  
Full Length ◽  
The Other ◽  
Aggregation Kinetics ◽  
Fibril Structure ◽  
N Terminus ◽  
Hairpin Conformation

The generation of α-synuclein (α-syn) truncations from incomplete proteolysis plays a significant role in the pathogenesis of Parkinson’s disease. It is well established that C-terminal truncations exhibit accelerated aggregation and serve as potent seeds in fibril propagation. In contrast, mechanistic understanding of N-terminal truncations remains ill defined. Previously, we found that disease-related C-terminal truncations resulted in increased fibrillar twist, accompanied by modest conformational changes in a more compact core, suggesting that the N-terminal region could be dictating fibril structure. Here, we examined three N-terminal truncations, in which deletions of 13-, 35-, and 40-residues in the N terminus modulated both aggregation kinetics and fibril morphologies. Cross-seeding experiments showed that out of the three variants, only ΔN13-α-syn (14‒140) fibrils were capable of accelerating full-length fibril formation, albeit slower than self-seeding. Interestingly, the reversed cross-seeding reactions with full-length seeds efficiently promoted all but ΔN40-α-syn (41–140). This behavior can be explained by the unique fibril structure that is adopted by 41–140 with two asymmetric protofilaments, which was determined by cryogenic electron microscopy. One protofilament resembles the previously characterized bent β-arch kernel, comprised of residues E46‒K96, whereas in the other protofilament, fewer residues (E61‒D98) are found, adopting an extended β-hairpin conformation that does not resemble other reported structures. An interfilament interface exists between residues K60‒F94 and Q62‒I88 with an intermolecular salt bridge between K80 and E83. Together, these results demonstrate a vital role for the N-terminal residues in α-syn fibril formation and structure, offering insights into the interplay of α-syn and its truncations.

In vitro Aggregation Ability of Five Commercially Available Aβ42 Peptides

2021 ◽  
Vol 18 ◽  
Author(s):  
Zhaoji Lv ◽  
Xi Du ◽  
Zhongsheng Chen ◽  
Fengjuan Liu ◽  
Rong Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Amyloid Β ◽  
Cell Counting ◽  
Basic Material ◽  
Aggregation Kinetics ◽  
Basic Study ◽  
Sds Page ◽  
Sigmoidal Kinetics ◽  
High Cytotoxicity

Background: As the most basic material, synthetic human Amyloid-β (1-42) (Aβ42) pep- tides from different manufacturers have been widely used. Their aggregation ability is vital to the reliability, repeatability and comparability of studies on Aβ42 physiology and pathology. However, it has not been evaluated and compared. Objectives: To analyze the consistency of the aggregation ability of 5 commercially available Aβ42 peptides. Methods: 5 Aβ42 peptides represented as A, B, C, D and E were pretreated by HFIP. The pretreated Aβ42 peptides were dissolved in Thioflavin T (ThT) solution, and their aggregation kinetics was monitored for 30 h with the aggregation kinetics test. Meanwhile, the pretreated peptides were ag- gregated in phosphate buffered saline. After aggregated for 12 h, they were detected by methods of ThT fluorescence, far-UV circular dichroism (CD), SDS-PAGE, western blot, and transmission electron microscopy (TEM), respectively. After aggregation for 8 h and 12 h, their cytotoxicity to SH-SY5Y cells was further evaluated using Cell Counting Kit-8. Results: For aggregation kinetics, peptides A, C and E remained low level curves, while peptides B and D presented typical sigmoidal kinetics curves. In CD measurement, the aggregates of pep- tides B and D showed relatively high negative CD peaks with the height of -8.09 mdeg and -14.37 mdeg, while the height of peptides A, C and E was -1.04, -3.55, and -3.88. In ThT assay, relative fluorescence intensity of the aggregates of peptides B and D were 7.79 and 8.82, higher than 1.19, 1.71, and 2.70 of peptide A, C and E, respectively. In SDS-PAGE, all aggregates contained monomers and eleven polymers. Moreover, peptide B-E presented a trapezoidal distribution from dimers to trimers, and peptide A aggregated to dimers. By western blot, the bands of monomers re- mained in all aggregates. Furthermore, peptides B and D aggregated to dimers and trimers, pep- tides A and C only aggregated to dimers, and peptide E showed a strong band of trimers. By TEM, protofibrils were observed only in peptide B, while substantial spherical aggregates were formed in other peptides. Additionally, peptides B, D and E exhibited higher cytotoxicity after being aggregat- ed for 8 h, whereas peptides A, B and D presented relatively high cytotoxicity after 12-hour aggre- gation. Conclusion: Commercially available Aβ42 peptides showed obvious differences in aggregation abil- ity, which should arouse enough attention in the field of basic study related to Aβ42. The aggrega- tion ability evaluation with the various assay methods has some discrepancies, and it is highly ur- gent to establish a reasonable and uniform measurement strategy.

scholarly journals Familial Alzheimer's Disease Mutations within the Amyloid Precursor Protein Alter the Aggregation and Conformation of the Amyloid-β Peptide

2017 ◽  
Vol 292 (8) ◽  
pp. 3172-3185 ◽  
Author(s):  
Asa Hatami ◽  
Sanaz Monjazeb ◽  
Saskia Milton ◽  
Charles G. Glabe
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Monoclonal Antibodies ◽  
Amyloid Precursor Protein ◽  
Amyloid Β ◽  
Thioflavin T ◽  
Precursor Protein ◽  
Aggregation Kinetics ◽  
Wide Range ◽  
Amyloid Structures

Most cases of Alzheimer's disease (AD) are sporadic, but a small percentage of AD cases, called familial AD (FAD), are associated with mutations in presenilin 1, presenilin 2, or the amyloid precursor protein. Amyloid precursor protein mutations falling within the amyloid-β (Aβ) sequence lead to a wide range of disease phenotypes. There is increasing evidence that distinct amyloid structures distinguished by amyloid conformation-dependent monoclonal antibodies have similarly distinct roles in pathology. It is possible that this phenotypic diversity of FAD associated with mutations within the Aβ sequence is due to differences in the conformations adopted by mutant Aβ peptides, but the effects of FAD mutations on aggregation kinetics and conformational and morphological changes of the Aβ peptide are poorly defined. To gain more insight into this possibility, we therefore investigated the effects of 11 FAD mutations on the aggregation kinetics of Aβ, as well as its ability to form distinct conformations recognized by a panel of amyloid conformation-specific monoclonal antibodies. We found that most FAD mutations increased the rate of aggregation of Aβ. The FAD mutations also led to the adoption of alternative amyloid conformations distinguished by monoclonal antibodies and resulted in the formation of distinct aggregate morphologies as determined by transmission electron microscopy. In addition, several of the mutant peptides displayed a large reduction in thioflavin T fluorescence, despite forming abundant fibrils indicating that thioflavin T is a probe of conformational polymorphisms rather than a reliable indicator of fibrillization. Taken together, these results indicate that FAD mutations falling within the Aβ sequence lead to dramatic changes in aggregation kinetics and influence the ability of Aβ to form immunologically and morphologically distinct amyloid structures.

scholarly journals S100A9 Alters the Pathway of Alpha-Synuclein Amyloid Aggregation

2021 ◽  
Vol 22 (15) ◽  
pp. 7972
Author(s):  
Zigmantas Toleikis ◽  
Mantas Ziaunys ◽  
Lina Baranauskiene ◽  
Vytautas Petrauskas ◽  
Kristaps Jaudzems ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Amyloid Fibril ◽  
Alpha Synuclein ◽  
Aggregation Kinetics ◽  
Neuron Death ◽  
Force Microscopy ◽  
Atomic Force ◽  
Fibril Structure ◽  
Parkinson’S Diseases ◽  
Main Protein

The formation of amyloid fibril plaques in the brain creates inflammation and neuron death. This process is observed in neurodegenerative disorders, such as Alzheimer’s and Parkinson’s diseases. Alpha-synuclein is the main protein found in neuronal inclusions of patients who have suffered from Parkinson’s disease. S100A9 is a calcium-binding, pro-inflammation protein, which is also found in such amyloid plaques. To understand the influence of S100A9 on the aggregation of α-synuclein, we analyzed their co-aggregation kinetics and the resulting amyloid fibril structure by Fourier-transform infrared spectroscopy and atomic force microscopy. We found that the presence of S100A9 alters the aggregation kinetics of α-synuclein and stabilizes the formation of a particular amyloid fibril structure. We also show that the solution’s ionic strength influences the interplay between S100A9 and α-synuclein, stabilizing a different structure of α-synuclein fibrils.

Fitting Yeast and Mammalian Prion Aggregation Kinetic Data with the Finke−Watzky Two-Step Model of Nucleation and Autocatalytic Growth†

Biochemistry ◽  
2008 ◽  
Vol 47 (40) ◽  
pp. 10790-10800 ◽  
Author(s):  
Murielle A. Watzky ◽  
Aimee M. Morris ◽  
Eric D. Ross ◽  
Richard G. Finke
Keyword(s):  
Kinetic Data ◽  
Aggregation Kinetic ◽  
Step Model ◽  
Autocatalytic Growth

scholarly journals Modulating Kinetics of the Amyloid-Like Aggregation of S. aureus Phenol-Soluble Modulins by Changes in pH

2021 ◽  
Vol 9 (1) ◽  
pp. 117
Author(s):  
Masihuz Zaman ◽  
Maria Andreasen
Keyword(s):  
Staphylococcus Aureus ◽  
Clear Correlation ◽  
Aggregation Kinetics ◽  
Ph Values ◽  
Fibril Structure ◽  
Lag Times ◽  
Functional Amyloids ◽  
Important Basis ◽  
Kinetics Of ◽  
Microscopic Techniques

The pathogen Staphylococcus aureus is recognized as one of the most frequent causes of biofilm-associated infections. The recently identified phenol-soluble modulin (PSM) peptides act as the key molecular effectors of staphylococcal biofilm maturation and promote the formation of an aggregated fibril structure. The aim of this study was to evaluate the effect of various pH values on the formation of functional amyloids of individual PSM peptides. Here, we combined a range of biophysical, chemical kinetics and microscopic techniques to address the structure and aggregation mechanism of individual PSMs under different conditions. We established that there is a pH-induced switch in PSM aggregation kinetics. Different lag times and growth of fibrils were observed, which indicates that there was no clear correlation between the rates of fibril elongation among different PSMs. This finding confirms that pH can modulate the aggregation properties of these peptides and suggest a deeper understanding of the formation of aggregates, which represents an important basis for strategies to interfere and might help in reducing the risk of biofilm-related infections.

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