Thermodynamics of Amyloid Fibril Formation from Chemical Depolymerization

2019 ◽  
Author(s):  
Nicola Vettore ◽  
Alexander Buell
Keyword(s):  
Ionic Strength ◽  
Thermodynamic Stability ◽  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Polymerization Process ◽  
Published Data ◽  
Amyloid Formation ◽  
Ionic Strength Dependence ◽  
Mechanisms Of Formation

Amyloid fibrils are homo-molecular protein polymers that play an important role in disease and biological function. While much is known about their kinetics and mechanisms of formation, the origin and magnitude of their thermodynamic stability has received significantly less attention. This is despite the fact that the thermodynamic stability of amyloid fibrils is an important determinant of their lifetimes and processing in vivo. Here we use depolymerization by chemical denaturants of amyloid fibrils of two different proteins (PI3K-SH3 and glucagon) at different concentrations and show that the previously applied linear polymerization model is an oversimplification that does not capture the concentration dependence of chemical depolymerization of amyloid fibrils. We show that cooperative polymerization, which is compatible with the picture of amyloid formation as a nucleated polymerization process, is able to quantitatively describe the thermodynamic data. We use this combined experimental and conceptual framework in order to probe the ionic strength<br>dependence of amyloid fibril stability. In combination with previously published data on the ionic strength dependence of amyloid fibril growth kinetics, our results provide strong evidence for the product-like nature of the transition state of amyloid fibril growth.

Thermodynamics of Amyloid Fibril Formation from Chemical Depolymerization

2019 ◽  
Author(s):  
Nicola Vettore ◽  
Alexander Buell
Keyword(s):  
Ionic Strength ◽  
Thermodynamic Stability ◽  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Polymerization Process ◽  
Published Data ◽  
Amyloid Formation ◽  
Ionic Strength Dependence ◽  
Mechanisms Of Formation

Amyloid fibrils are homo-molecular protein polymers that play an important role in disease and biological function. While much is known about their kinetics and mechanisms of formation, the origin and magnitude of their thermodynamic stability has received significantly less attention. This is despite the fact that the thermodynamic stability of amyloid fibrils is an important determinant of their lifetimes and processing in vivo. Here we use depolymerization by chemical denaturants of amyloid fibrils of two different proteins (PI3K-SH3 and glucagon) at different concentrations and show that the previously applied linear polymerization model is an oversimplification that does not capture the concentration dependence of chemical depolymerization of amyloid fibrils. We show that cooperative polymerization, which is compatible with the picture of amyloid formation as a nucleated polymerization process, is able to quantitatively describe the thermodynamic data. We use this combined experimental and conceptual framework in order to probe the ionic strength<br>dependence of amyloid fibril stability. In combination with previously published data on the ionic strength dependence of amyloid fibril growth kinetics, our results provide strong evidence for the product-like nature of the transition state of amyloid fibril growth.

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.

Mechanism and ion-dependence of in vitro autoactivation of yeast proteinase A: possible implications for compartmentalized activation in vivo

1997 ◽  
Vol 326 (2) ◽  
pp. 339-344 ◽  
Author(s):  
H. Bart VAN DEN HAZEL ◽  
Anne Mette WOLFF ◽  
Morten C. KIELLAND-BRANDT ◽  
Jakob R. WINTHER
Keyword(s):  
Ionic Strength ◽  
Aspartic Proteinase ◽  
General Mechanism ◽  
Activation Product ◽  
Ionic Strength Dependence ◽  
Strength Dependence ◽  
Proteinase A ◽  
Rapid Activation

Yeast proteinase A is synthesized as a zymogen which transits through the endoplasmic reticulum, the Golgi complex and the endosome to the vacuole. On arrival in the vacuole, activation takes place. It has previously been found that proteinase A can activate autocatalytically; however, the propeptide of proteinase A shows essentially no similarity to other known aspartic proteinase propeptides. To understand why proteinase A activation occurs rapidly in the vacuole but not at all in earlier compartments, we have purified the zymogen and investigated the conditions that trigger autoactivation and the mechanism of autoactivation. Autoactivation was triggered by acidic pH and its rate increased with increasing ionic strength. Kinetic evidence indicates that autoactivation mainly occurs via a bimolecular product-catalysed mechanism in which an active proteinase A molecule activates a zymogen molecule. Both the pH- and ionic-strength-dependence and the predominance of a product-catalysed mechanism are well adapted to the situation in vivo, since slow activation in the absence of active proteinase A helps to prevent activation in prevacuolar compartments, whereas, on delivery to the vacuole, lower pH, higher ionic strength and the presence of already active proteinases ensure rapid activation. Product-catalysed autoactivation may be a general mechanism by which cells ensure autoactivation of intracellular enzymes to be both rapid and compartmentalized.

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.

Amyloidosis

2020 ◽  
pp. 2218-2234
Author(s):  
Mark B. Pepys ◽  
Philip N. Hawkins
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Polarized Light ◽  
Serum Amyloid ◽  
Amyloid P Component ◽  
Serum Amyloid P ◽  
Serum Amyloid P Component ◽  
Amyloid Deposits ◽  
Amyloid P

Amyloidosis is the clinical condition caused by extracellular deposition of amyloid in the tissues. Amyloid deposits are composed of amyloid fibrils, abnormal insoluble protein fibres formed by misfolding of their normally soluble precursors. About 30 different proteins can form clinically or pathologically significant amyloid fibrils in vivo as a result of either acquired or hereditary abnormalities. Small, focal, clinically silent amyloid deposits in the brain, heart, seminal vesicles, and joints are a universal accompaniment of ageing. Clinically important amyloid deposits usually accumulate progressively, disrupting the structure and function of affected tissues and lead inexorably to organ failure and death. There is no licensed treatment which can specifically clear amyloid deposits, but intervention which reduces the availability of the amyloid fibril precursor proteins can arrest amyloid accumulation and may lead to amyloid regression with clinical benefit. Pathology—amyloid fibrils bind Congo red dye producing pathognomonic green birefringence when viewed in high-intensity cross-polarized light, and the protein type can be identified by immunostaining or proteomic analysis. Amyloid deposits always contain a nonfibrillar plasma glycoprotein, serum amyloid P component, the universal presence of which is the basis for use of radioisotope-labelled serum amyloid P component as a diagnostic tracer. Clinicopathological correlation—amyloid may be deposited in any tissue of the body, including blood vessels walls and connective tissue matrix; clinical manifestations are correspondingly diverse. Identification of the amyloid fibril protein is always essential for appropriate clinical management. The specific types of amyloidosis covered in this chapter are reactive systemic (AA) amyloidosis, monoclonal immunoglobulin light chain (AL) amyloidosis, and hereditary systemic amyloidoses (including familial amyloid polyneuropathy).

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 Seminal Plasma Accelerates Semen-derived Enhancer of Viral Infection (SEVI) Fibril Formation by the Prostatic Acid Phosphatase (PAP248–286) Peptide

2012 ◽  
Vol 287 (15) ◽  
pp. 11842-11849 ◽  
Author(s):  
Joanna S. Olsen ◽  
John T. M. DiMaio ◽  
Todd M. Doran ◽  
Caitlin Brown ◽  
Bradley L. Nilsson ◽  
...  
Keyword(s):  
Acid Phosphatase ◽  
Viral Infection ◽  
Seminal Plasma ◽  
Amyloid Fibrils ◽  
Divalent Cations ◽  
Prostatic Acid Phosphatase ◽  
Fibril Formation ◽  
Amyloid Formation ◽  
Precursor Peptide

Amyloid fibrils contained in semen, known as SEVI, or semen-derived enhancer of viral infection, have been shown to increase the infectivity of HIV dramatically. However, previous work with these fibrils has suggested that extensive time and nonphysiologic levels of agitation are necessary to induce amyloid formation from the precursor peptide (a proteolytic cleavage product of prostatic acid phosphatase, PAP248–286). Here, we show that fibril formation by PAP248–286is accelerated dramatically in the presence of seminal plasma (SP) and that agitation is not required for fibrillization in this setting. Analysis of the effects of specific SP components on fibril formation by PAP248–286revealed that this effect is primarily due to the anionic buffer components of SP (notably inorganic phosphate and sodium bicarbonate). Divalent cations present in SP had little effect on the kinetics of fibril formation, but physiologic levels of Zn2+strongly protected SEVI fibrils from degradation by seminal proteases. Taken together, these data suggest that in thein vivoenvironment, PAP248–286is likely to form fibrils efficiently, thus providing an explanation for the presence of SEVI in human semen.

scholarly journals Pathogenesis, diagnosis and treatment of systemic amyloidosis

2001 ◽  
Vol 356 (1406) ◽  
pp. 203-211 ◽  
Author(s):  
M. B. Pepys
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Current Treatment ◽  
High Concentration ◽  
Calcium Dependent ◽  
Amyloid Deposits ◽  
Precursor Proteins ◽  
End Stage ◽  
Native Fold

Amyloidosis is a disorder of protein folding in which normally soluble proteins are deposited as abnormal, insoluble fibrils that disrupt tissue structure and cause disease. Although about 20 different unrelated proteins can form amyloid fibrils in vivo , all such fibrils share a common cross–β core structure. Some natural wild–type proteins are inherently amyloidogenic, form fibrils and cause amyloidosis in old age or if present for long periods at abnormally high concentration. Other amyloidogenic proteins are acquired or inherited variants, containing amino–acid substitutions that render them unstable so that they populate partly unfolded states under physiological conditions, and these intermediates then aggregate in the stable amyloid fold. In addition to the fibrils, amyloid deposits always contain the non–fibrillar pentraxin plasma protein, serum amyloid P component (SAP), because it undergoes specific calcium–dependent binding to amyloid fibrils. SAP contributes to amyloidogenesis, probably by stabilizing amyloid fibrils and retarding their clearance. Radiolabelled SAP is an extremely useful, safe, specific, non–invasive, quantitative tracer for scintigraphic imaging of systemic amyloid deposits. Its use has demonstrated that elimination of the supply of amyloid fibril precursor proteins leads to regression of amyloid deposits with clinical benefit. Current treatment of amyloidosis comprises careful maintenance of impaired organ function, replacement of end–stage organ failure by dialysis or transplantation, and vigorous efforts to control underlying conditions responsible for production of fibril precursors. New approaches under development include drugs for stabilization of the native fold of precursor proteins, inhibition of fibrillogenesis, reversion of the amyloid to the native fold, and dissociation of SAP to accelerate amyloid fibril clearance in vivo .

scholarly journals Secondary Nucleation and the Conservation of Structural Characteristics of Amyloid Fibril Strains

2021 ◽  
Vol 8 ◽  
Author(s):  
Saeid Hadi Alijanvand ◽  
Alessia Peduzzo ◽  
Alexander K. Buell
Keyword(s):  
Amyloid Fibrils ◽  
Amyloid Fibril ◽  
Structural Characteristics ◽  
Secondary Nucleation ◽  
Primary Nucleation ◽  
Structural Preferences ◽  
Initial Seed ◽  
The Brain

Amyloid fibrils are ordered protein aggregates and a hallmark of many severe neurodegenerative diseases. Amyloid fibrils form through primary nucleation from monomeric protein, grow through monomer addition and proliferate through fragmentation or through the nucleation of new fibrils on the surface of existing fibrils (secondary nucleation). It is currently still unclear how amyloid fibrils initially form in the brain of affected individuals and how they are amplified. A given amyloid protein can sometimes form fibrils of different structure under different solution conditions in vitro, but often fibrils found in patients are highly homogeneous. These findings suggest that the processes that amplify amyloid fibrils in vivo can in some cases preserve the structural characteristics of the initial seed fibrils. It has been known for many years that fibril growth by monomer addition maintains the structure of the seed fibril, as the latter acts as a template that imposes its fold on the newly added monomer. However, for fibrils that are formed through secondary nucleation it was, until recently, not clear whether the structure of the seed fibril is preserved. Here we review the experimental evidence on this question that has emerged over the last years. The overall picture is that the fibril strain that forms through secondary nucleation is mostly defined by the solution conditions and intrinsic structural preferences, and not by the seed fibril strain.

scholarly journals Pathogenic D76N Variant of β2-Microglobulin: Synergy of Diverse Effects in Both the Native and Amyloid States

Biology ◽  
2021 ◽  
Vol 10 (11) ◽  
pp. 1197
Author(s):  
Éva Bulyáki ◽  
Judit Kun ◽  
Tamás Molnár ◽  
Alexandra Papp ◽  
András Micsonai ◽  
...  
Keyword(s):  
Amyloid Fibrils ◽  
Conformational Stability ◽  
Point Mutations ◽  
Amyloid Formation ◽  
Mhc I ◽  
Mutant Proteins ◽  
Β2 Microglobulin ◽  
Partial Unfolding ◽  
The Stability

β2-microglobulin (β2m), the light chain of the MHC-I complex, is associated with dialysis-related amyloidosis (DRA). Recently, a hereditary systemic amyloidosis was discovered, caused by a naturally occurring D76N β2m variant, which showed a structure remarkably similar to the wild-type (WT) protein, albeit with decreased thermodynamic stability and increased amyloidogenicity. Here, we investigated the role of the D76N mutation in the amyloid formation of β2m by point mutations affecting the Asp76-Lys41 ion-pair of WT β2m and the charge cluster on Asp38. Using a variety of biophysical techniques, we investigated the conformational stability and partial unfolding of the native state of the variants, as well as their amyloidogenic propensity and the stability of amyloid fibrils under various conditions. Furthermore, we studied the intermolecular interactions of WT and mutant proteins with various binding partners that might have in vivo relevance. We found that, relative to WT β2m, the exceptional amyloidogenicity of the pathogenic D76N β2m variant is realized by the deleterious synergy of diverse effects of destabilized native structure, higher sensitivity to negatively charged amphiphilic molecules (e.g., lipids) and polyphosphate, more effective fibril nucleation, higher conformational stability of fibrils, and elevated affinity for extracellular components, including extracellular matrix proteins.

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