In silico folding of hydrophobic peptides that form ß-hairpin structures in solution.

Peptides designed with residues that have a high propensity to occur in ß-turns form ß-hairpin structures in apolar as well as in polar organic solvents such as dimethyl sulfoxide (DMSO). Due to limited solubility, their conformations have not been investigated experimentally in water. We have examined the conformations of four of such designed peptides that fold into well-defined ß-hairpin structures facilitated by ß-turns, in the crystalline state and in solution, by molecular dynamics simulations (MDS). The peptides folded into ß-hairpin structures in water, starting from the fully extended conformation. However, in DMSO, neither folding nor unfolding was observed during MDS, when the starting structures were unfolded and folded, respectively. The lack of folding in DMSO was investigated by constructing folding free energy landscapes by umbrella sampling. The folding free energy landscape is smooth in water, whereas in DMSO, folded and unfolded structures are separated by high-energy barriers. The folding free energy is less in DMSO compared with water due to a more stable unfolded structure in DMSO compared with water, which in turn is due to stabilisation of the unfolded state by hydrophobic interactions in DMSO. This finding will be helpful to researchers to accurately model and/or design small peptide structures in water and organic solvents.

Down regulation of defensin genes during SARS-CoV-2 infection.

Defensins, crucial components of the innate immune system, play a vital role against infection as part of frontline immunity. Association of SARS-CoV-2 infection with defensins has not been investigated. In this study, we have investigated the expression of defensin genes in the buccal cavity from patients with COVID-19 infection along with negative control samples. Nasopharyngeal/oropharyngeal swab samples collected for screening SARS-CoV-2 infection in early 2020 from Hyderabad, India, were analyzed for the expression of major defensin genes by the quantitative real-time reverse transcription polymerase chain reaction, qRT-PCR. Forty SARS-CoV-2 infected positive and 40 negative swab samples were selected for this study. Based on the qRT-PCR analysis involving gene specific primers for defensin genes, 9 defensin genes were found to be expressed in the nasopharyngeal/oropharyngeal cavity. Four defensin genes were found to be significantly down regulated in SARS-CoV-2 infected patients in comparison with the control samples based on differential expression analysis. The significantly down regulated genes were defensin beta 4A/B, 106B, 107B, and 103A. Down regulation of human beta defensin 2, 3, 6 and 7 suggests that antiviral innate immune response provided by defensins may be compromised in SARS-CoV-2 infection resulting in progression of the disease. Correction of the down regulation process through appropriate defensin peptide-based therapy could be an attractive method of treatment. Keywords: host defense; defensins; COVID-19; gene regulation; SARS-CoV-2.

Limpid hydrogels from ß-turn motif-connected tandem repeats of Aß16-22.

Self-assembling peptides constitute an important class of functional biomaterials. A number of short amyloidogenic stretches have been identified from amyloid proteins. Such peptides, as such or through subtle modifications, can turn out to be promising candidates for functional biomaterials. End-capped Aß16-22, the well-studied amyloidogenic stretch from ß-amyloid, is reported to be non-hydrogelating up to 20 mM concentration. Here we investigated the hydrogelation propensity of Aß16-22 repeats connected through ß-turn-supporting motifs. The peptide repeats connected through Asn-Gly, Aib-DPro, and DPro-Gly formed transparent hydrogels at concentrations ≥2 mM. The repeats of the aromatic analog Aß16-22(F20Y) also resulted in similar hydrogels. Like other peptide-based gels reported earlier, these gels could trap the anticancer drug doxorubicin and displayed steady release in water. In addition, the gels supported the growth of mammalian cell lines, HEK-293 and RIN-5F. These data show that turn-inducing motifs can have marked effects on the hydrogelating propensity of self-assembling peptides.

Self-assembly of t-butyloxycarbonyl protected dipeptide methyl esters composed of leucine, isoleucine, and valine into highly organized structures from alcohol and aqueous alcohol mixtures.

Short peptides composed of phenylalanine and sequences derived from amyloidogenic peptides have the ability to self-assemble to form nanostructures including hydrogels. The self-assembly of peptides composed of only hydrophobic amino acids and aliphatic protecting groups have not been investigated in detail. We have examined various aspects of nanostructures formed by N-terminal t-butyloxycarbonyl-protected aliphatic dipeptide methyl esters dissolved in various solvents. Scanning electron microscopic images indicate that depending on the sequence, position of the amino acid and solvent of dissolution, the peptides self-assemble into superstructures such as nanotubes and needles particularly from aqueous mixtures of organic solvents. Crystallization was not required for self-assembly into nanostructures. Circular dichroism and attenuated total internal reflection fourier transform infrared spectroscopy studies indicate that the peptides adopt ß-conformation in the superstructures both in solution and solid state. The nanostructures composed of entirely aliphatic moieties have the ability to bind to aromatic dyes such as Rhodamine 6G, Nile red and Congo red. They also bind to Thioflavin T although the structures do not resemble amyloid fibrils. The powder X-ray diffraction patterns suggest distinctive packing of the monomers. These structures are stabilized by intermolecular hydrogen bonds and hydrophobic interactions resulting in superstructures containing long distance order and were devoid of hemolytic activity.

A hybrid cationic peptide composed of human ß-defensin-1 and humanized θ-defensin sequences exhibits salt-resistant antimicrobial activity.

We have designed a hybrid peptide by combining sequences of human ß-defensin-1 (HBD-1) and θ-defensin, in an attempt to generate a molecule that combines the diversity in structure and biological activity of two different peptides to yield a promising therapeutic candidate. HBD-1 was chosen as it is a natural defensin of humans that is constitutively expressed, but its antibacterial activity is considerably impaired by elevated ionic strength. θ-Defensins are expressed in human bone marrow as a pseudogene and are homologous to rhesus monkey circular minidefensins. Retrocyclins are synthetic human θ-defensins. The cyclic nature of the θ-defensin peptides makes them salt resistant, nonhemolytic, and virtually noncytotoxic in vitro. However, a nonhuman circular molecule developed for clinical use would be less viable than a linear molecule. In this study, we have fused the C-terminal region of HBD-1 to the nonapeptide sequence of a synthetic retrocyclin. Cyclization was achieved by joining the terminal ends of the hybrid peptide by a disulfide bridge. The hybrid peptide with or without the disulfide bridge exhibited enhanced antimicrobial activity against both Gram-negative and Gram-positive bacteria as well as against fungi, including clinical bacterial isolates from eye infections. The peptide retained activity in the presence of NaCl and serum and was nonhemolytic in vitro. Thus, the hybrid peptide generated holds potential as a new class of antibiotics.

Self-assembly of a peptide with a tandem repeat of the Aß16-22 sequence linked by a ß turn-promoting dipeptide sequence.

Amyloid deposits have been found to be abundant in patients with Alzheimer's disease due to fibril formation by the Aß peptides. Peptide Aß16-22, comprising of the seven-residue segment KLVFFAE, spanning residues 16-22 of the full length Aß42 peptide, aggregates to form fibrils or other nanostructures in isolation, depending on the conditions of dissolution and incubation. In this study, we have examined the self-assembly of PAß, a tandem repeat peptide of the Aß16-22 sequence, joined by a ß-turn-inducing sequence Asn-Gly. To study the effect of various solvents on the self-association, hexafluoroisopropanol (HFIP), trifluoroethanol (TFE) and methanol were used. The peptide was also incubated in fibril-promoting conditions of 20% fluorinated alcohol-water mixtures which form dynamical solvent clusters, as well as in 20% MeOH-water mixture which does not form solvent clusters. Secondary structural studies suggest the presence of ß-structures. Electron microscopic images indicate that fibril formation occurs in a time-dependent manner, under different conditions of solvent composition. Thioflavin-T fluorescence studies confirm the presence of amyloid fibrils in the aggregates. Although the insertion of the Asn-Gly sequence has not facilitated the formation of an ideal Type I' rigid turn, the intramolecular interactions aid the formation of a flexible ß-turn conformation, with twisted ß-sheets. Interactions between the intermolecular ß-sheets result in the formation of amyloid fibrils. Organic solvents appear to play an important role in modulating self-assembly of peptide PAß during fibril formation. Studies on ß-hairpin engineered amyloidogenic peptides could lead to knowledge about suitable conditions for generating a diverse range of polymorphic structures.

Antimicrobial activity of human α-defensin 6 analogs: insights into the physico-chemical reasons behind weak bactericidal activity of HD6 in vitro.

Human α-defensin 6 (HD6), unlike other mammalian defensins, does not exhibit bactericidal activity, particularly against aerobic bacteria. Monomeric HD6 has a tertiary structure similar to other α-defensins in the crystalline state. However, the physico-chemical reasons behind the lack of antibacterial activity of HD6 are yet to be established unequivocally. In this study, we have investigated the antimicrobial activity of HD6 analogs. A linear analog of HD6, in which the distribution of arginine residues was similar to active α-defensins, shows broad-spectrum antimicrobial activity, indicating that atypical distribution of arginine residues contributes to the inactivity of HD6. Peptides spanning the N-terminal cationic segment were active against a wide range of organisms. Antimicrobial potency of these shorter analogs was further enhanced when myristic acid was conjugated at the N-terminus. Cytoplasmic localization of the analogs without fatty acylation was observed to be necessary for bacterial killing, while they exhibited fungicidal activity by permeabilizing Candida albicans membranes. Myristoylated analogs and the linear full-length arginine analog exhibited activity by permeabilizing bacterial and fungal membranes. Our study provides insights into the lack of bactericidal activity of HD6 against aerobic bacteria.

Engineering of a linear inactive analog of human ß-defensin 4 to generate peptides with potent antimicrobial activity.

Human ß-defensins (HBDs) are cationic antimicrobial peptides constrained by three disulfide bridges. They have diverse range of functions in the innate immune response. It is of interest to investigate whether linear analogs of defensins can be generated, which possess antimicrobial activity. In this study, we have designed linear peptides with potent antimicrobial activity from an inactive peptide spanning the N-terminus of HBD4. Our results show that l-arginine to d-arginine substitution imparts considerable antimicrobial activity against both bacteria and Candida albicans. Increase in hydrophobicity by fatty acylation of the peptides with myristic acid further enhances their potency. In the presence of high concentrations of salt, antimicrobial activity of the myristoylated peptide with l-arginine is attenuated relatively to a lesser extent as compared with the linear active peptide with d-arginine. Substitution of cysteine with the hydrophobic helix-promoting amino acid α-aminoisobutyric acid favors candidacidal activity but not antibacterial activity. The mechanism of killing by d-arginine substituted unacylated analog involves transient interaction with the bacterial membrane followed by translocation into the cytoplasm without membrane permeabilization. Accumulation of peptides in the cytoplasm can affect various cellular processes that lead to cell death. However, the peptide causes membrane permeabilization in case of C. albicans. Myristoylation results in greater interaction of the peptide chain with the microbial cell surface and causes membrane permeabilization. Results described in the study demonstrate that it is possible to generate highly active linear analogs of defensins by selective introduction of d-amino acids and fatty acids, which could be attractive candidates for development as therapeutic agents.

Covalently attached fatty acyl chains alter the aggregation behavior of an amyloidogenic peptide derived from human ß(2)-microglobulin.

Aggregation of a polypeptide chain into highly ordered amyloid aggregates is a complex process. Various factors, both extrinsic and intrinsic to the polypeptide chain, have been shown to perturb this process, leading to a drastic change in the amyloidogenic behavior, which is reflected in the polymorphism of amyloid aggregates at various levels of self-assembly. In this paper, we have investigated the ability of covalently linked long-chain fatty acids in modulating the self-assembly of an aromatic amino acid-rich highly amyloidogenic sequence derived from the amino acid region 59-71 of human ß2-microglobulin by thioflavin T (ThT) fluorescence microscopy, circular dichroism, and fluorescence spectroscopy. Our results indicate that under identical conditions of dissolution and concentration, each peptide enhances the fluorescence of ThT. However, the aggregates are morphologically distinct. For the same peptide, the aggregate morphologies are dependent on peptide concentration. Further, an optimum concentration, which varies with solution ionic strength, is required for the formation of fibrillar aggregates. We show that covalent modification of this amyloidogenic sequence, with long-chain fatty acids, affects the way the higher order amyloid structures assemble from the cross-ß units, in fatty acyl chain-dependent and position-dependent manner. Our data indicate that noncovalent interactions leading to amyloid fibril formation can be modulated by the hydrophobicity of covalently attached long-chain fatty acids resulting in self-assembly of the peptide chain to form nonfibrillar aggregates.

Tau fibrillogenesis.

The protein tau is the most abundant microtubule associated protein in the central and peripheral nervous system. In the brain, tau plays a role in the assembly and stabilization of microtubules. The function of tau, however, appears to overlap with other microtubule binding proteins. The observation that tau is associated with neurodegenerative diseases has renewed interest in this protein. Various aspects of structure and biochemistry of tau, fibril formation and clinical perspectives, including therapeutic strategies are reviewed in this chapter.

A proteomic perspective and involvement of cytokines in SARS-CoV-2 infection.

Infection with the SARS-CoV-2 virus results in manifestation of several clinical observations from asymptomatic to multi-organ failure. Biochemically, the serious effects are due to what is described as cytokine storm. The initial infection region for COVID-19 is the nasopharyngeal/oropharyngeal region which is the site where samples are taken to examine the presence of virus. We have now carried out detailed proteomic analysis of the nasopharyngeal/oropharyngeal swab samples collected from normal individuals and those tested positive for SARS-CoV-2, in India, during the early days of the pandemic in 2020, by RTPCR, involving high throughput quantitative proteomics analysis. Several proteins like annexins, cytokines and histones were found differentially regulated in the host human cells following SARS-CoV-2 infection. Genes for these proteins were also observed to be differentially regulated when their expression was analyzed. Majority of the cytokine proteins were found to be up regulated in the infected individuals. Cell to Cell signaling interaction, Immune cell trafficking and inflammatory response pathways were found associated with the differentially regulated proteins based on network pathway analysis.

Interaction of peptides spanning the transmembrane domain of caveolin-1 with model membranes.

Caveolin-1 has an atypical membrane-spanning domain comprising of 34 residues. Caveolin-1 targets to lipid droplets under certain conditions, where they are involved in signaling and cholesterol balance. In the present study, membrane association of synthetic peptides corresponding to the membrane-spanning domain of caveolin-1 has been investigated to obtain an insight into the topology of transmembrane region in the lipid bilayer and the effect of truncations in this sequence, as observed in the targeting to lipid droplets, by using model membranes. Fluorescence studies revealed strong association of the peptide corresponding to the membrane-spanning domain of caveolin-1 with anionic lipids as compared with zwitterionic lipids, which is consistent with the location of this domain in the cytoplasmic side of the plasma membrane. Association of a short 9 residue peptide corresponding to the C-terminus of caveolin-1 membrane-spanning domain with lipid vesicles revealed the importance of this region for association with model membranes. Our investigations indicate that the peptide corresponding to the membrane-spanning domain of caveolin-1 does not span the lipid bilayer. We propose that both caveolin scaffolding domain and transmembrane segment of caveolin-1 contribute to the strong association with the plasma membrane rendering the protein highly detergent resistant.

Hexafluoroisopropanol induces self-assembly of ß-amyloid peptides into highly ordered nanostructures.

Deposition of insoluble fibrillar aggregates of ß-amyloid (Aß) peptides in the brain is a hallmark of Alzheimer's disease. Apart from forming fibrils, these peptides also exist as soluble aggregates. Fibrillar and a variety of nonfibrillar aggregates of Aß have also been obtained in vitro. Hexafluoroisopropanol (HFIP) has been widely used to dissolve Aß and other amyloidogenic peptides. In this study, we show that the dissolution of Aß40, 42, and 43 in HFIP followed by drying results in highly ordered aggregates. Although α-helical conformation is observed, it is not stable for prolonged periods. Drying after prolonged incubation of Aß40, 42, and 43 peptides in HFIP leads to structural transition from α-helical to ß-conformation. The peptides form short fibrous aggregates that further assemble giving rise to highly ordered ring-like structures. Aß16-22, a highly amyloidogenic peptide stretch from Aß, also formed very similar rings when dissolved in HFIP and dried. HFIP could not induce α-helical conformation in Aß16-22, and rings were obtained from freshly dissolved peptide. The rings formed by Aß40, 42, 43, and Aß16-22 are composed of the peptides in ß-conformation and cause enhancement in thioflavin T fluorescence, suggesting that the molecular architecture of these structures is amyloid-like. Our results clearly indicate that dissolution of Aß40, 42 and 43 and the amyloidogenic fragment Aß16-22 in HFIP results in the formation of annular amyloid-like structures.

Self-assembly of short peptides composed of only aliphatic amino acids and a combination of aromatic and aliphatic amino acids.

The morphology of structures formed by the self-assembly of short N-terminal t-butyloxycarbonyl (Boc) and C-terminal methyl ester (OMe) protected and Boc-deprotected hydrophobic peptide esters was investigated. We have observed that Boc-protected peptide esters composed of either only aliphatic hydrophobic amino acids or aliphatic hydrophobic amino acids in combination with aromatic amino acids, formed highly organized structures, when dried from methanol solutions. Transmission and scanning electron microscopic images of the peptides Boc-Ile-Ile-OMe, Boc-Phe-Phe-Phe-Ile-Ile-OMe and Boc-Trp-Ile-Ile-OMe showed nanotubular structures. Removal of the Boc group resulted in disruption of the ability to form tubular structures though spherical aggregates were formed. Both Boc-Leu-Ile-Ile-OMe and H-Leu-Ile-Ile-OMe formed only spherical nanostructures. Dynamic light scattering studies showed that aggregates of varying dimensions were present in solution suggesting that self-assembly into ordered structures is facilitated by aggregation in solution. Fourier transform infrared spectroscopy and circular dichroism spectroscopy data show that although all four of the protected peptides adopt well-defined tertiary structures, upon removal of the Boc group, only H-Phe-Phe-Phe-Ile-Ile-OMe had the ability to adopt ß-structure. Our results indicate that hydrophobic interaction is a very important determinant for self-assembly and presence of charged and aromatic amino acids in a peptide is not necessary for self-assembly.

Effect of introducing a short amyloidogenic sequence from the Aß 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. Aß40 self-associates to form amyloid fibrils. Its central seven-residue segment KLVFFAE (Aß16-22), 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 Aß16-22 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 ß-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 Aß16-22 sequence is attached at the N-terminus.

Gramicidin S and melittin: potential anti-viral therapeutic peptides to treat SARS-CoV-2 infection.

The COVID19 pandemic has led to multipronged approaches for treatment of the disease. Since de novo discovery of drugs is time consuming, repurposing of molecules is now considered as one of the alternative strategies to treat COVID19. Antibacterial peptides are being recognized as attractive candidates for repurposing to treat viral infections. In this study, we describe the anti-SARS-CoV-2 activity of the well-studied antibacterial peptides gramicidin S and melittin obtained from Bacillus brevis and bee venom respectively. The EC50 values for gramicidin S and melittin were 1.571 µg and 0.656 µg respectively based on in vitro antiviral assay. Significant decrease in the viral load as compared to the untreated group with no/very less cytotoxicity was observed. Both the peptides treated to the SARS-CoV-2 infected Vero cells showed viral clearance from 12 h onwards with a maximal viral clearance after 24 h post infection. Proteomics analysis indicated that more than 250 proteins were differentially regulated in the gramicidin S and melittin treated SARS-CoV-2 infected Vero cells against control SARS-CoV-2 infected Vero cells after 24 and 48 h post infection. The identified proteins were found to be associated in the metabolic and mRNA processing of the Vero cells post-treatment and infection. Both these peptides could be attractive candidates for repurposing to treat SARS-CoV-2 infection.

Determinants of membrane association in the SH4 domain of Fyn: roles of N-terminus myristoylation and side-chain thioacylation.

The SH4 domain of Fyn, a member of the Src family of tyrosine kinases, though rich in polar amino acid residues, anchors to the cytosolic face of membranes upon fatty acylation. In order to probe the requirement of specific fatty acylation at the N-terminus and at the side-chain of this domain for membrane-association, we have studied the interaction of peptides corresponding to the polar segment of the SH4 domain of Fyn and its mono- and dually fatty acylated analogs with model membranes. While the polar segment without covalently linked fatty acids (KDKEATKLTEW-amide) does not interact with lipid vesicles, peptides with one covalently linked fatty acid at the N-terminus or in the side-chain, associate with zwitterionic and anionic lipids to varying degrees. The interaction of dually acylated peptides (Myr-GK(epsilon-myr)KDKEATKLTEW-amide and Myr-GC(S-pal)KDKEATKLTEW-amide) with lipids depends on the linkage between fatty acyl side-chain and peptide backbone. The peptide chain associates with membranes only when the side-chain acylation is via an amide bond and not via a thioester bond. Our investigations indicate that acylation is essential for membrane targeting and unacylated polar stretch of the SH4 domain does not have a role in membrane-anchoring. Side-chain acylation via a thioester bond not only provides membrane anchorage but also directs the peptide chain away from the bilayer which might be important to enable the full length protein to interact with other signaling partners.

Dysfunction of store-operated calcium channel in muscle cells lacking mg29.

The store-operated calcium channel (SOC) located in the plasma membrane (PM) mediates capacitative entry of extracellular calcium after depletion of intracellular calcium stores in the endoplasmic or sarcoplasmic reticulum (ER/SR). An intimate interaction between the PM and the ER/SR is essential for the operation of this calcium signalling pathway. Mitsugumin 29 (MG29) is a synaptophysin-family-related protein located in the junction between the PM and SR of skeletal muscle. Here, we identify SOC in skeletal muscle and characterise its regulation by MG29 and the ryanodine receptor (RyR) located in the SR. Targeted deletion of mg29 alters the junctional membrane structure, causes severe dysfunction of SOC and SR calcium homeostasis and increases the susceptibility of muscle to fatigue stimulation. Severe dysfunction of SOC is also identified in muscle cells lacking both type 1 and type 3 RyRs, indicating that SOC activation requires an intact interaction between the PM and the SR, and is linked to conformational changes of RyRs. Whereas defective SOC seems to be inconsequential to short-term excitation-contraction coupling, the slow cumulative calcium entry through SOC is crucial for long-term calcium homeostasis, such that reduced SOC activity exaggerates muscle fatigue under conditions of intensive exercise.

Impact on the replacement of Phe by Trp in a short fragment of Aß amyloid peptide on the formation of fibrils.

Aß(16-22) (Ac-KLVFFAE-NH(2) ) is one of the shortest amyloid fibril-forming sequences identified in ß-amyloid peptide. At neutral pH, the peptide forms fibrils in the concentration range of 0.2-2.0 mM after ≥ 10 days of incubation. Structures of the fibrils proposed based on solid-state NMR and MD simulations studies suggest antiparallel arrangement of ß-strands and aromatic interactions between the Phe residues. In an effort to examine the role of aromatic interactions between two Phe residues in Aß(16-22) , we have studied the self-assembly of Aß(16-22) (AßFF) and two of its variants, Ac-KLVFWAE-NH(2) (AßFW) and Ac-KLVWFAE-NH(2) (AßWF). The peptides were dissolved in methanol (MeOH) at a concentration of 1 mM and in water (AßFW and AßWF, 1 mM; AßFF, 330 µM). Peptide solutions (100 µM) were prepared in 50 mM sodium phosphate buffer at pH 7 by diluting from MeOH and water stock solutions. AßFW forms amyloid-like fibrils immediately from MeOH, as indicated by atomic force microscopy. Dilution of AßFW into phosphate buffer from stock solution prepared in MeOH results in fibrils, but with different morphology and dimensions. The secondary structure potentiated by MeOH seems to be important for the self-assembly of AßFW, as fibrils are not formed from water where the peptide is unordered. On the other hand, AßFF and AßWF do not form amyloid fibrils rapidly from any of the solvents used for dissolution. However, drying of AßWF from MeOH on mica surface gives rod-like and fibrous structures. Our study indicates that positioning of the aromatic residues F and W has an important role to play in promoting self-assembly of the Aß(16-22) peptides.