Structural elements regulating amyloidogenesis: a cholinesterase model system

Polymerization into amyloid fibrils is a crucial step in the pathogenesis of neurodegenerative syndromes. Amyloid assembly is governed by properties of the sequence backbone and specific side-chain interactions, since fibrils from unrelated sequences possess similar structures and morphologies. Therefore, characterization of the structural determinants driving amyloid aggregation is of fundamental importance. We investigated the forces involved in the amyloid assembly of a model peptide derived from the oligomerization domain of acetylcholinesterase (AChE), AChE(586-599), through the effect of single point mutations on beta-sheet propensity, conformation, fibrilization, surfactant activity, oligomerization and fibril morphology. AChE(586-599) was chosen due to its fibrilization tractability and AChE involvement in Alzheimer's disease. The results revealed how specific regions and residues can control AChE(586-599) assembly. Hydrophobic and/or aromatic residues were crucial for maintaining a high beta-strand propensity, for the conformational transition to beta-sheet, and for the first stage of aggregation. We also demonstrated that positively charged side-chains might be involved in electrostatic interactions, which could control the transition to beta-sheet, the oligomerization and assembly stability. Further interactions were also found to participate in the assembly. We showed that some residues were important for AChE(586-599) surfactant activity and that amyloid assembly might preferentially occur at an air-water interface. Consistently with the experimental observations and assembly models for other amyloid systems, we propose a model for AChE(586-599) assembly in which a steric-zipper formed through specific interactions (hydrophobic, electrostatic, cation-pi, SH-aromatic, metal chelation and polar-polar) would maintain the beta-sheets together. We also propose that the stacking between the strands in the beta-sheets along the fiber axis could be stabilized through pi-pi interactions and metal chelation. The dissection of the specific molecular recognition driving AChE(586-599) amyloid assembly has provided further knowledge on such poorly understood and complicated process, which could be applied to protein folding and the targeting of amyloid diseases.     

Molecular dynamics simulations of a fibrillogenic peptide derived from apolipoprotein C-II

The pathway to amyloid fibril formation in proteins involves specific structural changes leading to the combination of misfolded intermediates into oligomeric assemblies. Recent NMR studies showed the presence of “turns” in amyloid peptides, indicating that turn formation may play an important role in the nucleation of the intramolecular folding and possible assembly of amyloid. Fully solvated all-atom molecular dynamics simulations were used to study the structure and dynamics of the apolipoprotein C-II peptide 56 to 76, associated with the formation of amyloid fibrils. The peptide populated an ensemble of turn structures, stabilized by hydrogen bonds and hydrophobic interactions enabling the formation of a strong hydrophobic core which may provide the conditions required to initiate aggregation. Two competing mechanisms discussed in the literature were observed. This has implications in understanding the mechanism of amyloid formation in not only apoC-II and its fragments, but also in other amyloidogenic peptides.     

Analysis of the minimal amyloid-forming fragment of the islet amyloid polypeptide. An experimental support for the key role of the phenylalanine residue in amyloid formation.

The development of type II diabetes was shown to be associated with the formation of amyloid fibrils consisted of the islet amyloid polypeptide (IAPP or amylin). Recently, a short functional hexapeptide fragment of IAPP (NH(2)-NFGAIL-COOH) was found to form fibrils that are very similar to those formed by the full-length polypeptide. To better understand the specific role of the residues that compose the fragment, we performed a systematic alanine scan of the IAPP "basic amyloidogenic units." Turbidity assay experiments demonstrated that the wild-type peptide and the Asn(1) --> Ala and Gly(3) --> Ala peptides had the highest rate of aggregate formation, whereas the Phe(2) --> Ala peptide did not form any detectable aggregates. Dynamic light-scattering experiments demonstrated that all peptides except the Phe(2) --> Ala form large multimeric structures. Electron microscopy and Congo red staining confirmed that the structures formed by the various peptides are indeed amyloid fibrils. Taken together, the results of our study provide clear experimental evidence for the key role of phenylalanine residue in amyloid formation by IAPP. In contrast, glycine, a residue that was suggested to facilitate amyloid formation in other systems, has only a minor role, if any, in this case. Our results are discussed in the context of the remarkable occurrence of aromatic residues in short functional fragments and potent inhibitors of amyloid-related polypeptides. We hypothesize that pi-pi interactions may play a significant role in the molecular recognition and self-assembly processes that lead to amyloid formation.     

Dissociation of Abeta(16-22) amyloid fibrils probed by molecular dynamics

The mechanisms of deposition and dissociation are implicated in the assembly of amyloid fibrils. To investigate the kinetics of unbinding of Abeta(16-22) monomers from preformed fibrils, we use molecular dynamics (MD) simulations and the structures for Abeta(16-22) amyloid fibrils. Consistent with experimental studies, the dissociation of Abeta(16-22) peptides involves two main stages, locked and docked, after which peptides unbind. The lifetime of the locked state, in which a peptide retains fibril-like structure and interactions, extends up to 0.5 micros under normal physiological conditions. Upon cooperative rupture of all fibril-like hydrogen bonds (HBs) with the fibril, a peptide enters a docked state. This state is populated by disordered random coil conformations and its lifetime ranges from approximately 10 to 200 ns. The docked state is stabilized by hydrophobic side chain interactions, while the contribution from HBs is small. Our simulations also suggest that the peptides located on fibril edges may form stable beta-strand conformations distinct from the fibril "bulk". We propose that such edge peptides can act as fibril caps, which impede fibril elongation. Our results indicate that the interactions between unbinding peptides constitute the molecular basis for cooperativity of peptide dissociation. The kinetics of fibril growth is reconstructed from unbinding assuming the reversibility of deposition/dissociation pathways. The relation of in silica dissociation kinetics to experimental observations is discussed.     

Characterizing and optimizing protease/peptide inhibitor interactions, a new application for spot synthesis

A new method is presented that uses parallel peptide array synthesis on cellulose membranes to characterize protease/peptide inhibitor interactions. A peptide comprising P5-P4' of the third domain of turkey ovomucoid inhibitor was investigated for both binding to and inhibition of porcine pancreatic elastase. Binding was studied directly on the cellulose membrane, while inhibition was measured by an assay in microtiter plates with punched out peptide spots. The importance of each residue for binding or inhibition was determined by substitutional analyses, exchanging every original amino acid with all other 19 coded amino acids. Seven hundred eighty individual peptides were investigated for binding behavior to porcine pancreatic elastase, and 320 individual peptides were measured in inhibition experiments. The results provide new insights into the interaction between the ovomucoid derived peptide and subsites in the active site of elastase. Combining these data with length analysis we designed new peptides in a step-wise fashion which in the end not only inhibited elastase 400 times more strongly than the original peptide, but are highly specific for the enzyme. In addition, the optimized inhibitor peptide was protected against exopeptidase attack by substituting D-amino acids at both termini.     

Unwinding fibril formation of medin, the peptide of the most common form of human amyloid

Medin amyloid affects the medial layer of the thoracic aorta of most people above 50 years of age. The consequences of this amyloid are not completely known but the deposits may contribute to diseases such as thoracic aortic aneurysm and dissection or to the general diminished elasticity of blood vessels seen in elderly people. We show that the 50-amino acid residue peptide medin forms amyloid-like fibrils in vitro. With the use of Congo red staining, Thioflavin T fluorescence, electron microscopy, and a solid-phase binding assay on different synthetic peptides, we identified the last 18-19 amino acid residues to constitute the amyloid-promoting region of medin. We also demonstrate that the two C-terminal phenylalanines, previously suggested to be of importance for amyloid formation, are not required for medin amyloid formation.     

Inhibition of peptide amyloid formation by cationic peptides with homologous sequences

We have studied the model peptides that undergo self-initiated structural transition from alpha-helix to beta-sheet and self-assembling into amyloid fibrils. We here constructed an inhibition system of amyloid formation utilizing homologous recognition and assembly of peptides with increased solubility. Among 20 peptides with homologous sequences examined here, cationic peptides showed the stronger inhibition ability against the amyloid formation of a model peptide.     

A single mutation on the human amyloid polypeptide modulates fibril growth and affects the mechanism of amyloid-induced membrane damage

Amyloid fibril formation has been implicated in a wide range of human diseases and the interactions of amyloidogenic proteins with cell membranes are considered to be important in the aetiology of these pathologies. In type 2 diabetes mellitus (T2DM), the human islet amyloid polypeptide (hIAPP) forms amyloid fibrils which impair the functionality and viability of pancreatic β cells. The mechanisms of hIAPP cytotoxicity are linked to the ability of the peptide to self-aggregate and to interact with membranes. Previous studies have shown that the N-terminal part of hIAPP from residues 1 to 19 is the membrane binding domain. The non-amyloidogenic and nontoxic mouse IAPP differs from hIAPP by six residues out of 37, among which a single one, residue 18, lies in the membrane binding region. To gain more insight into hIAPP-membrane interactions we herein performed comprehensive biophysical studies on four analogues (H18R-IAPP, H18K-IAPP, H18E-IAPP and H18A-IAPP). Our data reveal that all peptides are able to insert efficiently in the membrane, indicating that residue 18 is not essential for hIAPP membrane binding and insertion. However, only wild-type hIAPP and H18K-IAPP are able to form fibrils at the membrane. Importantly, all peptides induce membrane damage; wild-type hIAPP and H18K-IAPP presumably cause membrane disruption mainly by fibril growth at the membrane, while for H18R-IAPP, H18E-IAPP and H18A-IAPP, membrane leakage is most likely due to high molecular weight oligomeric species. These results highlight the importance of the residue at position 18 in IAPP for modulating fibril formation at the membrane and the mechanisms of membrane leakage.     

Role of aromatic interactions in amyloid formation by peptides derived from human Amylin

Numerous polypeptides and proteins form amyloid deposits in vivo or in vitro. The mechanism of amyloid formation is not well-understood particularly in the case where unstructured polypeptides assemble to form amyloid. Aromatic-aromatic interactions are known to be important in globular proteins, and the possibility that they might play a key role in amyloid formation has been raised. The results of Ala-scanning experiments on short polypeptides derived from Amylin have suggested that aromatic interactions could be particularly important for this system. Here, we examine a set of Amylin-derived polypeptides in which the single aromatic residue has been substituted with a Leu and Ala. A peptide corresponding to residues 21-29 with a Phe-23 to Leu substitution, a free N terminus, and amidated C terminus readily forms amyloid. Shorter peptides derived from the putative minimal amyloid-forming segment of Amylin, residues 22-27, also form amyloid when Phe-23 is replaced by Leu. Amyloid formation is more facile when the N terminus is deprotonated and the peptide is uncharged. Substitution of the Phe with Ala results in a peptide that is noticeably less prone to form amyloid. A peptide corresponding to residues 10-19 of human Amylin with blocked termini and the sole aromatic residue, Phe-15, substituted by Leu readily forms amyloid. A Phe-15 to Ala substitution reduces significantly the ability to form amyloid. These results indicate that an aromatic residue is not required for amyloid formation in these systems and indicates that other factors such as size, beta-sheet propensity, and hydrophobicity of the side chain in question are also important.     

In vitro creation of amyloid fibrils from native and Arg124Cys mutated betaIGH3((110-131)) peptides, and its relevance for lattice corneal amyloid dystrophy type I

BetaIGH3 protein has been recently involved in the pathogenesis of blinding corneal diseases, some of which have characteristic amyloid corneal deposits. The 124 codon of the betaig-h3 gene seems to be crucial for the amyloidogenicity of the protein product. We presently report an in vitro system that reproducibly forms amyloid fibrils from betaIGH3((110-131)) derived peptides. We also assessed the differences in fibril formation of two 22-amino acid peptides centered on the 124 residue: the native form and the Arg124Cys peptide (mutation linked to lattice corneal amyloid dystrophy type 1). After dialysis of Arg124Cys peptide against PBS 1/15 M pH 7.4 for 72 hours, Congo red staining and electron microscopy demonstrated the presence of abundant material fulfilling the criteria of amyloid. Quantitative analysis with thioflavine T fluorescence studies confirmed the high capacity of Arg124Cys peptide to form amyloid fibrils when compared to the native form.     

The minimal amyloid-forming fragment of the islet amyloid polypeptide is a glycolipid-binding domain

Several proteins that interact with cell surface glycolipids share a common fold with a solvent-exposed aromatic residue that stacks onto a sugar ring of the glycolipid (CH-pi stacking interaction). Stacking interactions between aromatic residues (pi-pi stacking) also play a pivotal role in the assembly process, including many cases of amyloid fibril formation. We found a structural similarity between a typical glycolipid-binding domain (the V3 loop of HIV-1 gp120) and the minimal amyloid-forming fragment of the human islet amyloid polypeptide, i.e. the octapeptide core module NFGAILSS. In a monolayer assay at the air-water interface, the NFGAILSS peptide specifically interacted with the glycolipid lactosylceramide. The interaction appears to require an aromatic residue, as NLGAILSS was poorly recognized by lactosylceramide, whereas NYGAILSS behaved like NFGAILSS. In addition, we observed that the full-length human islet amyloid polypeptide (1-37) did interact with a monolayer of lactosylceramide, and that the glycolipid film significantly affected the aggregation process of the peptide. As glycolipid-V3 interactions are efficiently inhibited by suramin, a polyaromatic compound, we investigated the effects of suramin on amyloid formation by human islet amyloid polypeptide. We found that suramin inhibited amyloid fibril formation at low concentrations, but dramatically stimulated the process at high concentrations. Taken together, our results indicate that the minimal amyloid-forming fragment of human islet amyloid polypeptide is a glycolipid-binding domain, and provide further experimental support for the role of aromatic pi-pi and CH-pi stacking interactions in the molecular control of the amyloidogenesis process.     

Probing the importance of lateral hydrophobic association in self-assembling peptide hydrogelators

A class of peptides has been designed whose ability to self-assemble into hydrogel is dependent on their conformationally folded state. Under unfolding conditions aqueous peptide solutions are freely flowing having the viscosity of water. When folding is triggered by external stimuli, peptides adopt a β-hairpin conformation that self-assembles into a highly crosslinked network of fibrils affording mechanically rigid hydrogels. MAX 1, a 20 residue, amphiphilic hairpin self-assembles via a mechanism which entails both lateral and facial self-assembly events to form a network of fibrils whose local structure consists of a bilayer of hairpins hydrogen bonded in the direction of fibril growth. Lateral self-assembly along the long axis of the fibril is mainly facilitated by intermolecular hydrogen bonding between the strands of distinct hairpins and the formation of hydrophobic contacts between residue side chains of laterally associating hairpins. Facial assembly is driven by the hydrophobic collapse of the valine-rich faces of the amphiphilic hairpins affording a bilayer laminate. The importance of forming lateral hydrophobic contacts during hairpin self-assembly and the relative contribution these interactions have towards nano-scale morphology and material rigidity is probed via the study of: MAX1, a hairpin designed to exploit lateral hydrophobic interactions; MAX 4, a peptide with reduced ability to form these interactions; and MAX5, a control peptide. CD spectroscopy and rheological experiments suggest that the formation of lateral hydrophobic interactions aids the kinetics of assembly and contributes to the mechanical rigidity of the hydrogel. Transmission electron microscopy (TEM) shows that these interactions play an essential role in the self-assembly process leading to distinct nano-scale morphologies. Electronic Supplementary Material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Polar residues in transmembrane helices can decrease electrophoretic mobility in polyacrylamide gels without causing helix dimerization

There are only a few available methods to study lateral interactions and self assembly of transmembrane helices. One of the most frequently used methods is sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) which can report on strong interactions between peptides in SDS solution. Here we offer a cautionary tale about studying the folding and assembly of membrane proteins using peptides and SDS-PAGE experiments as a membrane mimetic system. At least for the specific peptide and detergent systems studied here, we show that a polar asparagine residue in the 12th position of an otherwise hydrophobic helical segment of 20 amino acids causes a peptide to migrate on SDS-PAGE gels with an apparent molecular weight that is twice its true molecular weight, suggesting dimerization. However when examined carefully in SDS solutions and in situ in the polyacrylamide gel itself using Forster resonance energy transfer no interaction can be detected. Instead we show evidence suggesting that differential interactions between peptide and detergent drive the differences in electrophoretic mobility without any interaction between peptides. These results emphasize the need to apply multiple independent techniques to the study of membrane protein folding, and they highlight the usefulness of studying folding and structure of membrane proteins in lipid membranes rather than in detergents.     

Characterizing the Assembly of the Sup35 Yeast Prion Fragment, GNNQQNY: Structural Changes Accompany a Fiber-to-Crystal Switch

Amyloid-like fibrils can be formed by many different proteins and peptides. The structural characteristics of these fibers are very similar to those of amyloid fibrils that are deposited in a number of protein misfolding diseases, including Alzheimer's disease and the transmissible spongiform encephalopathies. The elucidation of two crystal structures from an amyloid-like fibril-forming fragment of the yeast prion, Sup35, with sequence GNNQQNY, has contributed to knowledge regarding side-chain packing of amyloid-forming peptides. Both structures share a cross-β steric zipper arrangement but vary in the packing of the peptide, particularly in terms of the tyrosine residue. We investigated the fibrillar and crystalline structure and assembly of the GNNQQNY peptide using x-ray fiber diffraction, electron microscopy, intrinsic and quenched tyrosine fluorescence, and linear dichroism. Electron micrographs reveal that at concentrations between 0.5 and 10 mg/mL, fibers form initially, followed by crystals. Fluorescence studies suggest that the environment of the tyrosine residue changes as crystals form. This is corroborated by linear dichroism experiments that indicate a change in the orientation of the tyrosine residue over time, which suggests that a structural rearrangement occurs as the crystals form. Experimental x-ray diffraction patterns from fibers and crystals also suggest that these species are structurally distinct. A comparison of experimental and calculated diffraction patterns contributes to an understanding of the different arrangements accessed by the peptide.     

Secondary structure propensity and chirality of the amyloidophilic peptide p5 and its analogues impacts ligand binding - In vitro characterization

BackgroundPolybasic helical peptides, such as peptide p5, bind human amyloid extracts and synthetic amyloid fibrils. When radiolabeled, peptide p5 has been shown to specifically bind amyloid in vivo thereby allowing imaging of the disease. Structural requirements for heparin and amyloid binding have been studied using analogues of p5 that modify helicity and chirality.MethodsPeptide-ligand interactions were studied using CD spectroscopy and solution-phase binding assays with radiolabeled p5 analogues. The interaction of a subset of peptides was further studied by using molecular dynamics simulations.ResultsDisruption of the peptide helical structure reduced peptide binding to heparin and human amyloid extracts. The all-D enantiomer and the β-sheet-structured peptide bound all substrates as well as, or better than, p5. The interaction of helical and β-sheet structured peptides with Aβ fibrils was modeled and shown to involve both ionic and non-ionic interactions.ConclusionsThe α-helical secondary structure of peptide p5 is important for heparin and amyloid binding; however, helicity is not an absolute requirement as evidenced by the superior reactivity of a β-sheet peptide. The differential binding of the peptides with heparin and amyloid fibrils suggests that these molecular interactions are different. The all-D enantiomer of p5 and the β-sheet peptide are candidates for amyloid targeting reagents in vivo.General SignificanceEfficient binding of polybasic peptides with amyloid is dependent on the linearity of charge spacing in the context of an α-helical secondary structure. Peptides with an α-helix or β-sheet propensity and with similar alignment of basic residues is optimal.     

A cyclic peptide inhibitor of apoC-II peptide fibril formation: mechanistic insight from NMR and molecular dynamics analysis

The misfolding and aggregation of proteins to form amyloid fibrils is a characteristic feature of several common age-related diseases. Agents that directly inhibit formation of amyloid fibrils represent one approach to combating these diseases. We have investigated the potential of a cyclic peptide to inhibit fibril formation by fibrillogenic peptides from human apolipoprotein C-II (apoC-II). Cyc[60-70] was formed by disulfide cross-linking of cysteine residues added to the termini of the fibrillogenic peptide comprising apoC-II residues 60-70. This cyclic peptide did not self-associate into fibrils. However, substoichiometric concentrations of cyc[60-70] significantly delayed fibril formation by the fibrillogenic, linear peptides apoC-II[60-70] and apoC-II[56-76]. Reduction of the disulfide bond or scrambling the amino acid sequence within cyc[60-70] significantly impaired its inhibitory activity. The solution structure of cyc[60-70] was solved using NMR spectroscopy, revealing a well-defined structure comprising a hydrophilic face and a more hydrophobic face containing the Met60, Tyr63, Ile66 and Phe67 side chains. Molecular dynamics (MD) studies identified a flexible central region within cyc[60-70], while MD simulations of "scrambled" cyc[60-70] indicated an increased formation of intramolecular hydrogen bonds and a reduction in the overall flexibility of the peptide. Our structural studies suggest that the inhibitory activity of cyc[60-70] is mediated by an elongated structure with inherent flexibility and distinct hydrophobic and hydrophilic faces, enabling cyc[60-70] to interact transiently with fibrillogenic peptides and inhibit fibril assembly. These results suggest that cyclic peptides based on amyloidogenic core peptides could be useful as specific inhibitors of amyloid fibril formation.     

Promotion of formation of amyloid fibrils by aluminium adenosine triphosphate (AlATP)

The formation of amyloid fibrils is considered to be an important step in the aetiology of Alzheimer's disease and other amyloidoses. Fibril formation in vitro has been shown to depend on many different factors including modifications to the amino acid profile of fibrillogenic peptides and interactions with both large and small molecules of physiological significance. How these factors might contribute to amyloid fibril formation in vivo is not clear as very little is known about the promotion of fibril formation in undersaturated solutions of amyloidogenic peptides. We have used thioflavin T fluorescence and reverse phase high performance liquid chromatography to show that ATP, and in particular AlATP, promoted the formation of thioflavin T-reactive fibrils of beta amyloid and, an unrelated amyloidogenic peptide, amylin. Evidence is presented that induction of fibril formation followed the complexation of AIATP by one or more monomers of the respective peptide. However, the complex formed could not be identified directly and it is suggested that AlATP might be acting as a chaperone in the assembly of amyloid fibrils. The effect of AlATP was not mimicked by either AlADP or AlAMP. However, it was blocked by suramin, a P2 ATP receptor antagonist, and this has prompted us to speculate that the precursor proteins to beta amyloid and amylin may be substrates or receptors for ATP in vivo.     

Isotope-assisted vibrational circular dichroism investigations of amyloid β peptide fragment, Aβ(16-22)

Isotope-assisted vibrational circular dichroism (VCD) investigations have been used to probe the site specific local structure of an amyloid peptide for the first time. A seven residue peptide, NH₂-KLVFFAE-COOH, which represents the Aβ(16–22) fragment of the Alzheimer’s amyloid β peptide, was used for these investigations. ¹³C labels were introduced separately at the carbonyl group of leucine (residue 17), alanine (residue 21) and also at both sites together. Since VCD spectra provide structure dependent signs, band shapes and frequencies, the isotope-assisted VCD spectroscopy revealed information on site specific secondary structure of the polypeptide. Isotope dilution VCD experiments provided a means to distinguish between parallel and anti-parallel nature of the β-sheet structure formed by the Aβ(16–22) fragment. The current results establish the usefulness of isotope-assisted VCD analysis in determining the site specific secondary structure of amyloid peptides.     

The influence of phospholipid membranes on bovine calcitonin peptide's secondary structure and induced neurotoxic effects

The peptide hormone, calcitonin, which is associated with medullary carcinoma of the thyroid, has a marked tendency to form amyloid fibrils and may be a useful model in probing the role of peptide-membrane interactions in beta-sheet and amyloid formation and amyloid neurotoxicity. Using bovine calcitonin, we found that, like other amyloids, the peptide was toxic only when in a beta-sheet-rich, amyloid form, but was non-toxic, when it lacked an amyloid structure. We found that the peptide bound with significant affinity to membranes that contained either cholesterol and gangliosides. In addition, incubation of calcitonin with cholesterol-rich and ganglioside-containing membranes resulted in significant changes in peptide structure yielding a peptide enriched in beta-sheet and amyloid content. Because the cholesterol- and ganglioside-rich phospholipid systems enhanced the calcitonin beta-sheet and amyloid contents, and peptide amyloid content was associated with neurotoxicity, we then investigated whether depleting cellular cholesterol and gangliosides affected calcitonin neurotoxicity. We found that cholesterol and ganglioside removal significantly reduced the calcitonin-induced PC12 cell neurotoxicity. Similar results have been observed with other amyloid-forming peptides such as beta-amyloid (A beta) of Alzheimer's disease and suggest that modulation of membrane composition and peptide-membrane interactions may prove useful in the control of amyloid formation and amyloid neurotoxicity.     

The Role of Pro, Gly Lys, and Arg Containing Peptides on Amyloid-Beta Aggregation

Genetic, biochemical and pathological evidence support that self-assembly of amyloid-beta (Aβ) peptide into toxic aggregates is implicated as the cause of Alzheimer’s disease. An attractive therapeutic strategy for the treatment of AD is to prevent or interfere with Aβ aggregation. A systematic investigation of the effects of proline-, glycine-, arginine- and lysine- containing peptides (PGKLVYA, KKLVFFARRRRA and KKLVFFA) on the beta-amyloid aggregation was made using FTIR, circular dichroism, ANS binding, ThT binding and TEM analysis. These peptides are based on the central hydrophobic region of Aβ (residues 16–20), which is believed to be crucial in Aβ self-association. There is increasing evidence to suggest that protein aggregation, including amyloid fibril formation results from the strong self-association tendency of the partially folded intermediates. Addition of PGKLVYA and KKLVFFARRRRA resulted in increase in ANS fluorescence intensity, suggesting enhanced exposure of hydrophobic surface area. As observed by ThT and TEM analysis PGKLVYA and KKLVFFARRRRA promote non-fibrillar ensembles, while peptide KKLVFFA accelerated the fibrillization of Aβ peptide by stabilizing intermolecular interactions. Circular dichroism and FTIR data showed that PGKLVYA and KKLVFFARRRRA effectively prevented amyloid-beta (Aβ) peptide adopting the beta-sheet secondary structure correlated with fibrillogenesis. This result indicates that PGKLVYA and KKLVFFARRRRA might have triggered another mechanism of Aβ assembly.