Structure−Activity Relationships in Peptide Modulators of β-Amyloid Protein Aggregation: Variation in α,α-Disubstitution Results in Altered Aggregate Size and Morphology (original) (raw)
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Journal of Structural Biology, 2000
The progressive deposition of the amyloid β peptide (Aβ) in fibrillar form is a key feature in the development of the pathology in Alzheimer's disease (AD). We have characterized the time course of Aβ fibril formation using a variety of assays and under different experimental conditions. We describe in detail the morphological development of the Aβ polymerization process from pseudo-spherical
ACS Chemical Neuroscience, 2010
Considerable research effort has focused on the discovery of mitigators that block the toxicity of the βamyloid peptide (Aβ) by targeting a specific step involved in Aβ fibrillogenesis and subsequent aggregation. Given that aggregation intermediates are hypothesized to be responsible for Aβ toxicity, such compounds could likely prevent or mitigate aggregation, or alternatively cause further association of toxic oligomers into larger nontoxic aggregates. Herein we investigate the effect of modifications of the KLVFF hydrophobic core of Aβ by replacing N-and C-terminal groups with various polar moieties. Several of these terminal modifications were found to disrupt the formation of amyloid fibrils and in some cases induced the disassembly of preformed fibrils. Significantly, mitigators that incorporate MiniPEG polar groups were found to be effective against Aβ 1-40 fibrilligonesis. Previously, we have shown that mitigators incorporating alpha,alpha-disubstituted amino acids (RRAAs) were effective in disrupting fibril formation as well as inducing fibril disassembly. In this work, we further disclose that the number of polar residues (six) and RRAAs (three) in the original mitigator can be reduced without dramatically changing the ability to disrupt Aβ 1-40 fibrillization in vitro.
Journal of Structural Biology, 2012
Morphology of aggregation intermediates, polymorphism of amyloid fibrils and aggregation kinetics of the ''Arctic'' mutant of the Alzheimer's amyloid b-peptide, Ab (1-40) (E22G), in a physiologically relevant Tris buffer (pH 7.4) were thoroughly explored in comparison with the human wild type Alzheimer's amyloid peptide, wt-Ab (1-40) , using both in situ atomic force and electron microscopy, circular dichroism and thioflavin T fluorescence assays. For arc-Ab at the end of the 'lag'-period of fibrillization an abrupt appearance of $3 nm size 'spherical aggregates' with a homogeneous morphology, was identified. Then, the aggregation proceeds with a rapid growth of amyloid fibrils with a variety of morphologies, while the spherical aggregates eventually disappeared during in situ measurements. Arc-Ab was also shown to form fibrils at much lower concentrations than wt-Ab (1-40) : 62.5 lM and 12.5 lM, respectively. Moreover, at the same concentration, 50 lM, the aggregation process proceeds more rapidly for arc-Ab : the first amyloid fibrils were observed after c.a. 72 h from the onset of incubation as compared to approximately 7 days for wt-Ab . Amyloid fibrils of arc-Ab (1-40) exhibit a large variety of polymorphs, at least five, both coiled and non-coiled distinct fibril structures were recognized by AFM, while at least four types of arc-Ab (1-40) fibrils were identified by TEM and STEM and their mass-perlength statistics were collected suggesting supramolecular structures with two, four and six b-sheet laminae. Our results suggest a pathway of fibrillogenesis for full-length Alzheimer's peptides with small and structurally ordered transient spherical aggregates as on-pathway immediate precursors of amyloid fibrils.
The Properties of Amyloid-β Fibrils Are Determined by their Path of Formation
Journal of molecular biology, 2018
Fibril formation of the amyloid-β peptide (Aβ) follows a nucleation-dependent polymerization process and is associated with Alzheimer's disease. Several different lengths of Aβ are observed in vivo, but Aβ1-40 and Aβ1-42 are the dominant forms. The fibril architectures of Aβ1-40 and Aβ1-42 differ and Aβ1-42 assemblies are generally considered more pathogenic. We show here that monomeric Aβ1-42 can be cross-templated and incorporated into the ends of Aβ1-40 fibrils, while incorporation of Aβ1-40 monomers into Aβ1-42 fibrils is very poor. We also show that via cross-templating incorporated Aβ monomers acquire the properties of the parental fibrils. The suppressed ability of Aβ1-40 to incorporate into the ends of Aβ1-42 fibrils and the capacity of Aβ1-42 monomers to adopt the properties of Aβ1-40 fibrils may thus represent two mechanisms reducing the total load of fibrils having the intrinsic, and possibly pathogenic, features of Aβ1-42 fibrils in vivo. We also show that the transf...
Effects of Aβ-derived peptide fragments on fibrillogenesis of Aβ
Scientific Reports
Amyloid β (Aβ) peptide aggregation plays a central role in Alzheimer’s disease (AD) etiology. AD drug candidates have included small molecules or peptides directed towards inhibition of Aβ fibrillogenesis. Although some Aβ-derived peptide fragments suppress Aβ fibril growth, comprehensive analysis of inhibitory potencies of peptide fragments along the whole Aβ sequence has not been reported. The aim of this work is (a) to identify the region(s) of Aβ with highest propensities for aggregation and (b) to use those fragments to inhibit Aβ fibrillogenesis. Structural and aggregation properties of the parent Aβ1–42 peptide and seven overlapping peptide fragments have been studied, i.e. Aβ1–10 (P1), Aβ6–15 (P2), Aβ11–20 (P3), Aβ16–25 (P4), Aβ21–30 (P5), Aβ26–36 (P6), and Aβ31–42 (P7). Structural transitions of the peptides in aqueous buffer have been monitored by circular dichroism and Fourier transform infrared spectroscopy. Aggregation and fibrillogenesis were analyzed by light scatteri...
Journal of the American Chemical Society, 2016
Evidence suggests that oligomers of the 42residue form of the amyloid β-protein (Aβ), Aβ42, play a critical role in the etiology of Alzheimer's disease (AD). Here we use high resolution atomic force microscopy to directly image populations of small oligomers of Aβ42 that occur at the earliest stages of aggregation. We observe features that can be attributed to a monomer and to relatively small oligomers, including dimers, hexamers, and dodecamers. We discovered that Aβ42 hexamers and dodecamers quickly become the dominant oligomers after peptide solubilization, even at low (1 μM) concentrations and short (5 min) incubation times. Soon after (≥10 min), dodecamers are observed to seed the formation of extended, linear preprotofibrillar β-sheet structures. The preprotofibrils are a single Aβ42 layer in height and can extend several hundred nanometers in length. To our knowledge this is the first report of structures of this type. In each instance the preprotofibril is associated off center with a single layer of a dodecamer. Protofibril formation continues at longer times, but is accompanied by the formation of large, globular aggregates. Aβ40, by contrast, does not significantly form the hexamer or dodecamer but instead produces a mixture of smaller oligomers. These species lead to the formation of a branched chain-like network rather than discrete structures.
Stability of Aβ (1-42) peptide fibrils as consequence of environmental modifications
European Biophysics Journal, 2010
Amyloid peptide (A ) plays a key role in the pathogenesis of Alzheimer disease (AD). Monomeric A undergoes aggregation, forming oligomers and Wbrils, resulting in the deposition of plaques in the brain of AD patients. A widely used protocol for Wbril formation in vitro is based on incubation of the peptide at low pH and ionic strength, which generates A Wbrils several microns long. What happens to such Wbrils once they are brought to physiological pH and ionic strength for biological studies is not fully understood. In this investigation, we show that these changes strongly aVect the morphology of Wbrils, causing their fragmentation into smaller ones followed by their aggregation into disordered structures. We show that an increase in pH is responsible for Wbril fragmentation, while increased ionic strength is responsible for the aggregation of Wbril fragments. This behavior was conWrmed on diVerent batches of peptide either produced by the same company or of diVerent origin. Similar aggregates of short Wbrils are obtained when monomeric peptide is incubated under physiological conditions of pH and ionic strength, suggesting that Wbril morphology is independent of the Wbrillation protocol but depends on the Wnal chemical environment. This was also conWrmed by experiments with cell cultures showing that the toxicity of Wbrils with diVerent initial morphology is the same after addition to the medium. This information is of fundamental importance when A Wbrils are prepared in vitro at acidic pH and then diluted into physiological buVer for biological investigations.
Proteins: Structure, Function, and Bioinformatics, 2016
The histopathological hallmark of Alzheimer's disease (AD) is the aggregation and accumulation of the amyloid beta peptide (Ab) into misfolded oligomers and fibrils. Here we examine the biophysical properties of a protective Ab variant against AD, A2T, and a causative mutation, A2V, along with the wild type (WT) peptide. The main finding here is that the A2V native monomer is more stable than both A2T and WT, and this manifests itself in different biophysical behaviors: the kinetics of aggregation, the initial monomer conversion to an aggregation prone state (primary nucleation), the abundances of oligomers, and extended conformations. Aggregation reaction modeling of the conversion kinetics from native monomers to fibrils predicts the enhanced stability of the A2V monomer, while ion mobility spectrometry-mass spectrometry measures this directly confirming earlier predictions. Additionally, unique morphologies of the A2T aggregates are observed using atomic force microscopy, providing a basis for the reduction in long term potentiation inhibition of hippocampal cells for A2T compared with A2V and the wild type (WT) peptide. The stability difference of the A2V monomer and the difference in aggregate morphology for A2T (both compared with WT) are offered as alternate explanations for their pathological effects.
Journal of Molecular Biology, 2010
The ability of a single polypeptide sequence to grow into multiple stable amyloid fibrils sets these aggregates apart from most native globular proteins. The existence of multiple amyloid forms is the basis for strain effects in yeast prion biology, and may also contribute to variations in Alzheimer's disease pathology. However, the structural basis for amyloid polymorphism is poorly understood. We report here five structurally distinct fibrillar aggregates of the Alzheimer's plaque peptide Aβ(1-40), as well as a non-fibrillar aggregate induced by Zn +2. Each of these conformational forms exhibits a unique profile of physical properties, and all the fibrillar forms "breed true" in elongation reactions at a common set of growth conditions. Consistent with their defining cross-β structure, we find that in this series the amyloid fibrils containing more extensive β-sheet exhibit greater stability. At the same time, side chain packing outside of the β-sheet regions also contributes to stability, and to stability differences between polymorphic forms. Stability comparison is facilitated by the unique feature that the free energy of the monomer (equivalent to the unfolded state in a protein folding reaction) does not vary, and hence can be ignored, in the comparison of ΔG° of elongation values for each polymorphic fibril obtained at a single set of conditions. The aggregated, β-sheet rich amyloid structure represents a stable, alternatively folded state of polypeptides. Amyloid fibrils are associated with several important neurodegenerative diseases, such as Alzheimer's and Huntington's diseases 1, as well as a number of peripheral diseases of organ failure 2. Amyloid fibrils can be produced in vitro from many proteins, consistent with the polymeric structure of proteins and the relationship of amyloid fibrils to synthetic polymers 3. The fundamental unit of amyloid fibrils is the cross-β structure, in which β-sheet extended chains and sheet-sheet stacking interactions are perpendicular to the fibril axis and β-sheet H-bonds are parallel to the fibril axis 4. Details of the threedimensional structures of amyloid fibrils are still being elucidated 5 ; 6 ; 7 ; 8 ; 9 ; 10 ; 11 ; 12 One striking feature of amyloid fibrils that sets them apart from most globular proteins is the ability of a single polypeptide chain to grow into more than one stable structure 13. The existence of multiple protein aggregate conformations, each of which can propagate with