N‐terminal truncated pyroglutamyl β amyloid peptide Aβpy3‐42 shows a faster aggregation kinetics than the full‐length Aβ1‐42

We tested directly the differences in the aggregation kinetics of three important β amyloid peptides, the full‐length Aβ1‐42, and the two N‐terminal truncated and pyroglutamil modified Aβpy3‐42 and Aβpy11‐42 found in different relative concentrations in the brains in normal aging and in Alzheimer disease. By following the circular dichroism signal and the ThT fluorescence of the solution in phosphate buffer, we found substantially faster aggregation kinetics for Aβpy3‐42. This behavior is due to the particular sequence of this peptide, which is also responsible for the specific oligomeric aggregation states, found by TEM, during the fibrillization process, which are very different from those of Aβ1‐42, more prone to fibril formation. In addition, Aβpy3‐42 is found here to have an inhibitory effect on Aβ1‐42 fibrillogenesis, coherently with its known greater infective power. This is an indication of the important role of this peptide in the aggregation process of β‐peptides in Alzheimer disease. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 861–873, 2009. This article was originally published online as an accepted preprint. The “Published Online“ date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

A β‐amino acid modified heptapeptide containing a designed recognition element disrupts fibrillization of the amyloid β‐peptide

We study the complex formation of a peptide βAβAKLVFF, previously developed by our group, with Aβ(1–42) in aqueous solution. Circular dichroism spectroscopy is used to probe the interactions between βAβAKLVFF and Aβ(1–42), and to study the secondary structure of the species in solution. Thioflavin T fluorescence spectroscopy shows that the population of fibers is higher in βAβAKLVFF/Aβ(1–42) mixtures compared to pure Aβ(1–42) solutions. TEM and cryo‐TEM demonstrate that co‐incubation of βAβAKLVFF with Aβ(1–42) causes the formation of extended dense networks of branched fibrils, very different from the straight fibrils observed for Aβ(1–42) alone. Neurotoxicity assays show that although βAβAKLVFF alters the fibrillization of Aβ(1–42), it does not decrease the neurotoxicity, which suggests that toxic oligomeric Aβ(1–42) species are still present in the βAβAKLVFF/Aβ(1–42) mixtures. Our results show that our designed peptide binds to Aβ(1–42) and changes the amyloid fibril morphology. This is shown to not necessarily translate into reduced toxicity. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.     

FTIR spectroscopic studies on aggregation process of the β‐amyloid 11–28 fragment and its variants

Aggregation of Aβ peptides is a seminal event in Alzheimer's disease. Detailed understanding of the Aβ assembly process would facilitate the targeting and design of fibrillogenesis inhibitors. Here, conformational studies using FTIR spectroscopy are presented. As a model peptide, the 11–28 fragment of Aβ was used. This model peptide is known to contain the core region responsible for Aβ aggregation. The structural behavior of the peptide during aggregation provoked by the addition of water to Aβ(11–28) solution in hexafluoroisopropanol was compared with the properties of its variants corresponding to natural, clinically relevant mutants at positions 21–23 (A21G, E22K, E22G, E22Q and D23N). The results showed that the aggregation of the peptides proceeds via a helical intermediate, and it is possible that the formation of α‐helical structures is preceded by creation of 3₁₀‐helix/3₁₀‐turn structures. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd.     

Evidence of π‐stacking interactions in the self‐assembly of hIAPP22‐29

The role aromatic amino acids play in the formation of amyloid is a subject of controversy. In an effort to clarify the contribution of aromaticity to the self‐assembly of human islet amyloid polypeptide (hIAPP)_22‐29, peptide analogs containing electron donating groups (EDGs) or electron withdrawing groups (EWGs) as substituents on the aromatic ring of Phe‐23 at the para position have been synthesized and characterized using turbidity measurements in conjunction with Raman and fluorescence spectroscopy. Results indicate the incorporation of EDGs on the aromatic ring of Phe‐23 virtually abolish the ability of hIAPP_22‐29 to form amyloid. Peptides containing EWGs were still capable of forming aggregates. These aggregates were found to be rich in β‐sheet secondary structure. Transmission electron microscopy images of the aggregates confirm the presence of amyloid fibrils. The observed difference in amyloidogenic propensity between peptides containing EDGs and those with EWGs appears not to be based on differences in peptide hydrophobicity. Fluorescence and Raman spectroscopic investigations reveal that the environment surrounding the aromatic ring becomes more hydrophobic and ordered upon aggregation. Furthermore, Raman measurements of peptide analogs containing EWGs, conclusively demonstrate a distinct downshift in the ? C?C? ring mode (ca. 1600 cm^?1) upon aggregation that has previously been shown to be indicative of π‐stacking. While previous work has demonstrated that π‐stacking is not an absolute requirement for fibrillization, our findings indicate that Phe‐23 also contributes to fibril formation through π‐stacking interactions and that it is not only the hydrophobic nature of this residue that is relevant in the self‐assembly of hIAPP_22‐29. © Proteins 2013. © 2012 Wiley Periodicals, Inc.     

Effect of agitation on the peptide fibrillization: Alzheimer's amyloid‐β peptide 1‐42 but not amylin and insulin fibrils can grow under quiescent conditions

Many peptides and proteins can form fibrillar aggregates in vitro, but only a limited number of them are forming pathological amyloid structures in vivo. We studied the fibrillization of four peptides – Alzheimer's amyloid‐β (Aβ) 1‐40 and 1‐42, amylin and insulin. In all cases, intensive mechanical agitation of the solution initiated fast fibrillization. However, when the mixing was stopped during the fibril growth phase, the fibrillization of amylin and insulin was practically stopped, and the rate for Aβ₄₀ substantially decreased, whereas the fibrillization of Aβ₄₂ peptide continued to proceed with almost the same rate as in the agitated conditions. The reason for the different sensitivity of the in vitro fibrillization of these peptides towards agitation in the fibril growth phase remains elusive. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Docosahexaenoic acid disrupts in vitro amyloid β1‐40 fibrillation and concomitantly inhibits amyloid levels in cerebral cortex of Alzheimer’s disease model rats

We have previously reported that dietary docosahexaenoic acid (DHA) improves and/or protects against impairment of cognition ability in amyloid beta_1‐40 (Aβ_1‐40)‐infused Alzheimer’s disease (AD)‐model rats. Here, after the administration of DHA to AD model rats for 12 weeks, the levels of Aβ_1‐40, cholesterol and the composition of fatty acids were investigated in the Triton X100‐insoluble membrane fractions of their cerebral cortex. The effects of DHA on the in vitro formation and kinetics of fibrillation of Aβ_1‐40 were also investigated by thioflavin T fluorescence spectroscopy, transmission electron microscopy and fluorescence microscopy. Dietary DHA significantly decreased the levels of Aβ_1‐40, cholesterol and saturated fatty acids in the detergent insoluble membrane fractions of AD rats. The formation of Aβ fibrils was also attenuated by their incubation with DHA, as demonstrated by the decreased intensity of thioflavin T‐derived fluorescence and by electron micrography. DHA treatment also decreased the intensity of thioflavin fluorescence in preformed‐fibril Aβ peptides, demonstrating the anti‐amyloidogenic effects of DHA. We then investigated the effects of DHA on the levels of oligomeric amyloid that is generated during its in vitro transformation from monomers to fibrils, by an anti‐oligomer‐specific antibody and non‐reducing Tris‐Glycine gradient (4–20%) gel electrophoresis. DHA concentration‐dependently reduced the levels of oligomeric amyloid species, suggesting that dietary DHA‐induced suppression of in vivo Aβ_1‐40 aggregation occurs through the inhibitory effect of DHA on oligomeric amyloid species.     

Retracted: S14G‐humanin inhibits Aβ1–42 fibril formation,disaggregates preformed fibrils,and protects against Aβ‐induced cytotoxicity in vitro

The aggregation of soluble amyloid‐beta (Aβ) peptide into oligomers/fibrils is one of the key pathological features in Alzheimer's disease (AD). The Aβ aggregates are considered to play a pivotal role in the pathogenesis of AD. Therefore, inhibiting Aβ aggregation and destabilizing preformed Aβ fibrils would be an attractive therapeutic target for prevention and treatment of AD. S14G‐humanin (HNG), a synthetic derivative of Humanin (HN), has been shown to be a strong neuroprotective agent against various AD‐related insults. Recent studies have shown that HNG can significantly improve cognitive deficits and reduce insoluble Aβ levels as well as amyloid plaque burden without affecting amyloid precursor protein processing and Aβ production in transgenic AD models. However, the potential mechanisms by which HNG reduces Aβ‐related pathology in vivo remain obscure. In the present study, we found that HNG could significantly inhibit monomeric Aβ1–42 aggregation into fibrils and destabilize preformed Aβ1–42 fibrils in a concentration‐dependent manner by Thioflavin T fluorescence assay. In transmission electron microscope study, we observed that HNG was effective in inhibiting Aβ1–42 fibril formation and disrupting preformed Aβ1–42 fibrils, exhibiting various types of amorphous aggregates without identifiable Aβ fibrils. Furthermore, HNG‐treated monomeric or fibrillar Aβ1–42 was found to significantly reduce Aβ1–42‐mediated cytotoxic effects on PC12 cells in a dose‐dependent manner by MTT assay. Collectively, our results demonstrate for the first time that HNG not only inhibits Aβ1–42 fibril formation but also disaggregates preformed Aβ1–42 fibrils, which provides the novel evidence that HNG may have anti‐Aβ aggregation and fibrillogenesis, and fibril‐destabilizing properties. Together with previous studies, we concluded that HNG may have promising therapeutic potential as a multitarget agent for the prevention and/or treatment of AD. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Side‐chain hydrophobicity and the stability of Aβ16–22 aggregates

Recent mutagenesis studies using the hydrophobic segment of Aβ suggest that aromatic π‐stacking interactions may not be critical for fibril formation. We have tested this conjecture by probing the effect of Leu, Ile, and Ala mutation of the aromatic Phe residues at positions 19 and 20, on the double‐layer hexametric chains of Aβ fragment Aβ_16–22 using explicit solvent all‐atom molecular dynamics. As these simulations rely on the accuracy of the utilized force fields, we first evaluated the dynamic and stability dependence on various force fields of small amyloid aggregates. These initial investigations led us to choose AMBER99SB‐ILDN as force field in multiple long molecular dynamics simulations of 100 ns that probe the stability of the wild‐type and mutants oligomers. Single‐point and double‐point mutants confirm that size and hydrophobicity are key for the aggregation and stability of the hydrophobic core region (Aβ_16–22). This suggests as a venue for designing Aβ aggregation inhibitors the substitution of residues (especially, Phe 19 and 20) in the hydrophobic region (Aβ_16–22) with natural and non‐natural amino acids of similar size and hydrophobicity.     

Aromatic‐interaction‐mediated inhibition of β‐amyloid assembly structures and cytotoxicity

Abnormal aggregation of β‐amyloid (Aβ) peptide plays an important role in the onset and progress of Alzheimer's disease (AD); hence, targeting Aβ aggregation is considered as an effective therapeutic strategy. Here, we studied the aromatic‐interaction‐mediated inhibitory effect of oligomeric polypeptides (K8Y8, K4Y8, K8W8) on Aβ42 fibrillization process. The polypeptides containing lysine as well as representative aromatic amino acids of tryptophan or tyrosine were found to greatly suppress the aggregation as evaluated by thioflavin T assay. Circular dichroism spectra showed that the β‐sheet formation of Aβ42 peptides decreased with the polypeptide additives. Molecular docking studies revealed that the oligomeric polypeptides could preferentially bind to Aβ42 through π–π stacking between aromatic amino acids and Phe19, together with hydrogen bonding. The cell viability assay confirmed that the toxicity of Aβ42 to SH‐SY5Y cells was markedly reduced in the presence of polypeptides. This study could be beneficial for developing peptide‐based inhibitory agents for amyloidoses. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.     

Ethanol induced the formation of β‐sheet and amyloid‐like fibrils by surfactant‐like peptide A6K

Self‐assembly of natural or designed peptides into fibrillar structures based on β‐sheet conformation is a ubiquitous and important phenomenon. Recently, organic solvents have been reported to play inductive roles in the process of conformational change and fibrillization of some proteins and peptides. In this study, we report the change of secondary structure and self‐assembling behavior of the surfactant‐like peptide A6K at different ethanol concentrations in water. Circular dichroism indicated that ethanol could induce a gradual conformational change of A6K from unordered secondary structure to β‐sheet depending upon the ethanol concentration. Dynamic light scattering and atomic force microscopy revealed that with an increase of ethanol concentration the nanostructure formed by A6K was transformed from nanosphere/string‐of‐beads to long and smooth fibrils. Furthermore, Congo red staining/binding and thioflavin‐T binding experiments showed that with increased ethanol concentration, the fibrils formed by A6K exhibited stronger amyloid fibril features. These results reveal the ability of ethanol to promote β‐sheet conformation and fibrillization of the surfactant‐like peptide, a fact that may be useful for both designing self‐assembling peptide nanomaterials and clarifying the molecular mechanism behind the formation of amyloid fibrils. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Fe2+ binding on amyloid β‐peptide promotes aggregation

The metal ions Zn²⁺, Cu²⁺, and Fe²⁺ play a significant role in the aggregation mechanism of Aβ peptides. However, the nature of binding between metal and peptide has remained elusive; the detailed information on this from the experimental study is very difficult. Density functional theory (dft) (M06‐2X/6‐311++G (2df,2pd) +LANL2DZ) has employed to determine the force field resulting due to metal and histidine interaction. We performed 200 ns molecular dynamics (MD) simulation on Aβ_1‐42‐Zn²⁺, Aβ_1‐42‐Cu²⁺, and Aβ_1‐42‐Fe²⁺ systems in explicit water with different combination of coordinating residues including the three Histidine residues in the N‐terminal. The present investigation, the Aβ_1‐42‐Zn²⁺ system possess three turn conformations separated by coil structure. Zn²⁺ binding caused the loss of the helical structure of N‐terminal residues which transformed into the S‐shaped conformation. Zn²⁺ has reduced the coil and increases the turn content of the peptide compared with experimental study. On the other hand, the Cu²⁺ binds with peptide, β sheet formation is observed at the N‐terminal residues of the peptide. Fe²⁺ binding is to promote the formation of Glu22‐Lys28 salt‐bridge which stabilized the turn conformation in the Phe19‐Gly25 residues, subsequently β sheets were observed at His13‐Lys18 and Gly29‐Gly37 residues. The turn conformation facilitates the β sheets are arranged in parallel by enhancing the hydrophobic contact between Gly25 and Met35, Lys16 and Met35, Leu17 and Leu34, Val18 and Leu34 residues. The Fe²⁺ binding reduced the helix structure and increases the β sheet content in the peptide, which suggested, Fe²⁺ promotes the oligomerization by enhancing the peptide‐peptide interaction. Proteins 2016; 84:1257–1274. © 2016 Wiley Periodicals, Inc.     

Design, synthesis, and biophysical properties of a helical Abeta1-42 analog: Inhibition of fibrillogenesis and cytotoxicity

The aggregation of amyloid β-peptide (Aβ) into β-sheet-rich aggregates is a crucial step in the etiology of Alzheimer’s disease. Helical forms of Aβ have been suggested to be intermediates in the aggregation process of the peptide in aqueous phase, micelles and membranes. A stable helical Aβ analog would be useful to investigate the role of helical intermediates in fibrillization by Aβ. Here we designed a helical analog by simply cross-linking the Cys residues of A30C, G37C-Aβ1-42 with 1,6-bismaleimidohexane. The analog assumed a weak α-helical conformation in model membranes mimicking lipid raft microdomains of neuronal membranes under conditions in which the wild-type Aβ1-42 formed a β-sheet, indicating the cross-linking locally induced a helical conformation. Furthermore, addition of equimolar helical Aβ analog significantly reduced the amyloid formation and cytotoxicity by Aβ1-42. Thus, our helical Aβ1-42 is not only a model peptide to investigate the role of helical intermediates in fibrillization by Aβ, but also an inhibitor of Aβ-induced cytotoxicity.     

Rational design of amyloid β peptide–binding proteins: Pseudo‐Aβ β‐sheet surface presented in green fluorescent protein binds tightly and preferentially to structured Aβ

Some neurodegenerative diseases such as Alzheimer disease (AD) and Parkinson disease are caused by protein misfolding. In AD, amyloid β‐peptide (Aβ) is thought to be a toxic agent by self‐assembling into a variety of aggregates involving soluble oligomeric intermediates and amyloid fibrils. Here, we have designed several green fluorescent protein (GFP) variants that contain pseudo‐Aβ β‐sheet surfaces and evaluated their abilities to bind to Aβ and inhibit Aβ oligomerization. Two GFP variants P13H and AP93Q bound tightly to Aβ, K_d = 260 nM and K_d = 420 nM, respectively. Moreover, P13H and AP93Q were capable of efficiently suppressing the generation of toxic Aβ oligomers as shown by a cell viability assay. By combining the P13H and AP93Q mutations, a super variant SFAB4 comprising four strands of Aβ‐derived sequences was designed and bound more tightly to Aβ (K_d = 100 nM) than those having only two pseudo‐Aβ strands. The SFAB4 protein preferentially recognized the soluble oligomeric intermediates of Aβ more than both unstructured monomer and mature amyloid fibrils. Thus, the design strategy for embedding pseudo‐Aβ β‐sheet structures onto a protein surface arranged in the β‐barrel structure is useful to construct molecules capable of binding tightly to Aβ and inhibiting its aggregation. This strategy may provide implication for the diagnostic and therapeutic development in the treatment of AD. Proteins 2010. © 2009 Wiley‐Liss, Inc.     

Nitric oxide‐mediated regulation of ‐amyloid clearance via alterations of MMP‐9/TIMP‐1

Fibrillar amyloid plaques are largely composed of amyloid‐beta (Aβ) peptides that are metabolized into products, including Aβ1‐16, by proteases including matrix metalloproteinase 9 (MMP‐9). The balance between production and degradation of Aβ proteins is critical to amyloid accumulation and resulting disease. Regulation of MMP‐9 and its endogenous inhibitor tissue inhibitor of metalloproteinase (TIMP)‐1 by nitric oxide (NO) has been shown. We hypothesize that nitric oxide synthase (NOS2) protects against Alzheimer's disease pathology by increasing amyloid clearance through NO regulation of MMP‐9/TIMP‐1 balance. We show NO‐mediated increased MMP‐9/TIMP‐1 ratios enhanced the degradation of fibrillar Aβ in vitro, which was abolished when silenced for MMP‐9 protein translation. The in vivo relationship between MMP‐9, NO and Aβ degradation was examined by comparing an Alzheimer's disease mouse model that expresses NOS2 with a model lacking NOS2. To quantitate MMP‐9 mediated changes, we generated an antibody recognizing the Aβ1‐16 fragment, and used mass spectrometry multi‐reaction monitoring assay for detection of immunoprecipitated Aβ1‐16 peptides. Aβ1‐16 levels decreased in brain lysates lacking NOS2 when compared with strains that express human amyloid precursor protein on the NOS2 background. TIMP‐1 increased in the APPSwDI/NOS2^?/? mice with decreased MMP activity and increased amyloid burden, thereby supporting roles for NO in the regulation of MMP/TIMP balance and plaque clearance.     

Wild‐type,Flemish, and Dutch amyloid‐β exhibit different cytotoxicities depending on Aβ40 to Aβ42 interaction time and concentration ratio

Addition of amyloid β (Aβ) peptide Aβ40 to Aβ42 can delay Aβ42 aggregation, but consequent cytotoxicity has been reported to be enhanced or diminished. In the present study, we found that cytotoxicity was enhanced when human neuroblastoma SH‐SY5Y cells were incubated in a mixture of wt Aβ42 and Aβ40wt at a ratio of 1 : 10–20 (0.1 : 1–2 μM) for 24–36 h, whereas the enhancement was detected in cells incubated for longer times (48–60 h) with the less amyloidogenic Flemish Aβ40 variant or in cells incubated for as short as 12 h with the more amyloidogenic Dutch variant. Reductions in cytotoxicity by Aβ40 were most prominently observed in the Flemish and wt Aβ40/Aβ42 mixture at ratio 1 : 20 incubated for a short time (~12 h). The most cytotoxic Aβ40/Aβ42 mixtures were enriched in Aβ protofibril‐like structures, implying a strong correlation between cytotoxicity and this structure, the formation of which was dependent on amyloidogenic properties and incubation time. The consequences of the interactions were probably because of the different amyloidogenic properties of the Aβ40 variants, rather than to those of Aβ42, because aggregation rates of Aβ40 variants were highly dependent on sequence, whereas those of Aβ42 variants were not. These studies highlight a potential role for Aβ40 in cytotoxicity and provide novel mechanistic insights into the pathogenesis of each familial Alzheimer's disease‐associated Aβ40 variant. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Rationally designed dehydroalanine (ΔAla)‐containing peptides inhibit amyloid‐β (Aβ) peptide aggregation

Among the pathological hallmarks of Alzheimer's disease (AD) is the deposition of amyloid‐β (Aβ) peptides, primarily Aβ (1–40) and Aβ (1–42), in the brain as senile plaques. A large body of evidence suggests that cognitive decline and dementia in AD patients arise from the formation of various aggregated forms of Aβ, including oligomers, protofibrils and fibrils. Hence, there is increasing interest in designing molecular agents that can impede the aggregation process and that can lead to the development of therapeutically viable compounds. Here, we demonstrate the ability of the specifically designed α,β‐dehydroalanine (ΔAla)‐containing peptides P1 (K‐L‐V‐F‐ΔA‐I‐ΔA) and P2 (K‐F‐ΔA‐ΔA‐ΔA‐F) to inhibit Aβ (1–42) aggregation. The mechanism of interaction of the two peptides with Aβ (1–42) seemed to be different and distinct. Overall, the data reveal a novel application of ΔAla‐containing peptides as tools to disrupt Aβ aggregation that may lead to the development of anti‐amyloid therapies not only for AD but also for many other protein misfolding diseases. © 2009 Wiley Periodicals, Inc. Biopolymers 91: 456–465, 2009. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

The protonation state of histidine 111 regulates the aggregation of the evolutionary most conserved region of the human prion protein

In a group of neurodegenerative diseases, collectively termed transmissible spongiform encephalopathies, the prion protein aggregates into β‐sheet rich amyloid‐like deposits. Because amyloid structure has been connected to different prion strains and cellular toxicity, it is important to obtain insight into the structural properties of prion fibrils. Using a combination of solution NMR spectroscopy, thioflavin‐T fluorescence and electron microscopy we here show that within amyloid fibrils of a peptide containing residues 108–143 of the human prion protein [humPrP (108–143)]—the evolutionary most conserved part of the prion protein ‐ residue H111 and S135 are in close spatial proximity and their interaction is critical for fibrillization. We further show that residues H111 and H140 share the same microenvironment in the unfolded, monomeric state of the peptide, but not in the fibrillar form. While protonation of H140 has little influence on fibrillization of humPrP (108–143), a positive charge at position 111 blocks the conformational change, which is necessary for amyloid formation of humPrP (108–143). Our study thus highlights the importance of protonation of histidine residues for protein aggregation and suggests point mutations to probe the structure of infectious prion particles.     

Zinc-induced heterodimer formation between metal-binding domains of intact and naturally modified amyloid-beta species: implication to amyloid seeding in Alzheimer’s disease?

Zinc ions and modified amyloid-beta peptides (Aβ) play a critical role in the pathological aggregation of endogenous Aβ in Alzheimer’s disease (AD). Zinc-induced Aβ oligomerization is mediated by the metal-binding domain (MBD) which includes N-terminal residues 1–16 (Aβ_1–16). Earlier, it has been shown that Aβ_1–16 as well as some of its naturally occurring variants undergoes zinc-induced homodimerization via the interface in which zinc ion is coordinated by Glu11 and His14 of the interacting subunits. In this study using surface plasmon resonance technique, we have found that in the presence of zinc ions Aβ_1–16 forms heterodimers with MBDs of two Aβ species linked to AD: Aβ containing isoAsp7 (isoAβ) and Aβ containing phosphorylated Ser8 (pS8-Aβ). The heterodimers appear to possess the same interface as the homodimers. Simulation of 200 ns molecular dynamic trajectories in two constructed models of dimers ([Aβ_1–16/Zn/Aβ_1–16] and [isoAβ_1–16/Zn/Aβ_1–16]), has shown that conformational flexibility of the N-terminal fragments of the dimer subunits is controlled by the structure of corresponding sites 6–8. The data suggest that isoAβ and pS8-Aβ can be involved in the AD pathogenesis by means of their zinc-dependent interactions with endogenous Aβ resulting in the formation of heterodimeric seeds for amyloid aggregation.     

Effect of introducing a short amyloidogenic sequence from the Aβ peptide at the N‐terminus of 18‐residue amphipathic helical peptides

Fibril formation is the hallmark of pathogenesis in Alzheimer's disease and other amyloid disorders caused by conformational alterations leading to the aggregation of soluble monomers. Aβ40 self‐associates to form amyloid fibrils. Its central seven‐residue segment KLVFFAE (Aβ16–22), which is thought to be crucial for fibril formation of the full‐length peptide, forms fibrils even in isolation. Context‐dependent induction of amyloid formation by such sequences in peptides, which otherwise do not have that propensity, is of considerable interest. We have examined the effect of introducing the Aβ16–22 sequence at the N‐terminus of two amphipathic helical 18‐residue peptides Ac‐WYSEMKRNVQRLERAIEE‐am and Ac‐KQLIRFLKRLDRNLWGLA‐am, which have high average hydrophobic moment <μH> values but have net charges of 0 and +4, respectively, at neutral pH. Upon incubation in aqueous buffer, fibril‐like aggregates were discernible by transmission electron microscopy for the peptide with only 0 net charge, which also displayed ThT binding and β‐structure. Although both the sequences have been derived from amphipathic helical segments in globular proteins and possess high average hydrophobic moments, the +4 charge peptide lacks the ability to form fibrils, while the peptide with 0 charge has the tendency to form fibrillar structures. Variation in the net charge and the presence of several glutamic acids in the sequence of the peptide with net charge 0 appear to favor the formation of fibrils when the Aβ16–22 sequence is attached at the N‐terminus. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd.     

Structural analysis of the pyroglutamate‐modified isoform of the Alzheimer's disease‐related amyloid‐β using NMR spectroscopy

The aggregation of the Aβ plays a fundamental role in the pathology of AD. Recently, N‐terminally modified Aβ species, pE‐Aβ, have been described as major constituents of Aβ deposits in the brains of AD patients. pE‐Aβ has an increased aggregation propensity and shows increased toxicity compared with Aβ1‐40 and Aβ1‐42. In the present work, high‐resolution NMR spectroscopy was performed to study pE‐Aβ3‐40 in aqueous TFE‐containing solution. Two‐dimensional TOCSY and NOESY experiments were performed. On the basis of NOE and chemical shift data, pE‐Aβ3‐40 was shown to contain two helical regions formed by residues 14–22 and 30–36. This is similar as previously described for Aβ1‐40. However, the secondary chemical shift data indicate decreased helical propensity in pE‐Aβ3‐40 when compared with Aβ1‐40 under exactly the same conditions. This is in agreement with the observation that pE‐Aβ3‐40 shows a drastically increased tendency to form β‐sheet‐rich structures under more physiologic conditions. Structural studies of pE‐Aβ are crucial for better understanding the structural basis of amyloid fibril formation in the brain during development of AD, especially because an increasing number of reports indicate a decisive role of pE‐Aβ for the pathogenesis of AD. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.