-
Kyle L Morris,
Lin Chen,
Jaclyn Raeburn,
Owen R Sellick,
Pepa Cotanda,
Alison Paul,
Peter C Griffiths,
Stephen M King,
Rachel K O'Reilly, Louise C Serpell,
Dave J Adams
[show abstract]
[hide abstract]
ABSTRACT: Controlling the order and spatial distribution of self-assembly in multicomponent supramolecular systems could underpin exciting new functional materials, but it is extremely challenging. When a solution of different components self-assembles, the molecules can either coassemble, or self-sort, where a preference for like-like intermolecular interactions results in coexisting, homomolecular assemblies. A challenge is to produce generic and controlled 'one-pot' fabrication methods to form separate ordered assemblies from 'cocktails' of two or more self-assembling species, which might have relatively similar molecular structures and chemistry. Self-sorting in supramolecular gel phases is hence rare. Here we report the first example of the pH-controlled self-sorting of gelators to form self-assembled networks in water. Uniquely, the order of assembly can be predefined. The assembly of each component is preprogrammed by the pK(a) of the gelator. This pH-programming method will enable higher level, complex structures to be formed that cannot be accessed by simple thermal gelation.
Nature Communications 02/2013; 4:1480. · 7.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Elaborate morphology: The αSβ1 peptide, a fragment of α-synuclein, assembles into flat tapes consisting of a peptide bilayer, which can be modeled based on the cross-β structure found in amyloid proteins. The tapes are stabilized by hydrogen bonding, whilst the amphiphilic nature of the peptide results in the thin bilayer structure. To further stabilize the structure, these tapes may twist to form helical tapes, which subsequently close into nanotubes.
Angewandte Chemie International Edition 01/2013; · 13.45 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Amyloid fibril formation is associated with misfolding diseases, as well as fulfilling a functional role. The cross-ß molecular architecture has been reported in increasing numbers of amyloid-like fibrillar systems. The Waltz algorithm is able to predict ordered self-assembly of amyloidogenic peptides by taking into account residue type and position. This algorithm has expanded the amyloid sequence space and here we characterise the structures of amyloid-like fibrils formed by three peptides identified by Waltz that form fibrils but not crystals. The structural challenge is met by combining electron microscopy, linear and circular dichroism and X-ray fibre diffraction. We propose structures that reveal a cross-ß conformation with 'steric-zipper' features, giving insights into the role for side chains in peptide packing and stability within fibrils. The amenity of these peptides to structural characterisation makes them compelling model systems to use for understanding the relationship between sequence, self-assembly, stability and structure for amyloid fibrils.
Biochemical Journal 12/2012; · 4.90 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Naphthalene dipeptides have been shown to be useful low-molecular-weight gelators. Here we have used a library to explore the relationship between the dipeptide sequence and the hydrogelation efficiency. A number of the naphthalene dipeptides are crystallizable from water, enabling us to investigate the comparison between the gel/fiber phase and the crystal phase. We succeeded in crystallizing one example directly from the gel phase. Using X-ray crystallography, molecular modeling, and X-ray fiber diffraction, we show that the molecular packing of this crystal structure differs from the structure of the gel/fiber phase. Although the crystal structures may provide important insights into stabilizing interactions, our analysis indicates a rearrangement of structural packing within the fibers. These observations are consistent with the fibrillar interactions and interatomic separations promoting 1D assembly whereas in the crystals the peptides are aligned along multiple axes, allowing 3D growth. This observation has an impact on the use of crystal structures to determine supramolecular synthons for gelators.
Langmuir 05/2012; 28(25):9797-806. · 4.19 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Amyloid fibrils are polymeric assemblies of normally soluble proteins or peptides. To investigate their structure, it is generally not possible to use conventional methods of crystallography and solution nuclear magnetic resonance. To examine the repeating crystalline structure along the fibre axis, X-ray fibre diffraction has been a useful tool. Here we discuss the methods by which amyloid-like fibrils may be prepared to form a sample suitable for structural analysis and describe how data may be collected and then analysed to arrive at a potential model structure.
Methods in molecular biology (Clifton, N.J.) 01/2012; 849:121-35.
-
Kris Pauwels,
Thomas L Williams,
Kyle L Morris,
Wim Jonckheere,
Annelies Vandersteen,
Geoff Kelly,
Joost Schymkowitz,
Frederic Rousseau,
Annalisa Pastore, Louise C Serpell,
Kerensa Broersen
[show abstract]
[hide abstract]
ABSTRACT: The β-amyloid peptide (Aβ) is directly related to neurotoxicity in Alzheimer disease (AD). The two most abundant alloforms of the peptide co-exist under normal physiological conditions in the brain in an Aβ(42):Aβ(40) ratio of ∼1:9. This ratio is often shifted to a higher percentage of Aβ(42) in brains of patients with familial AD and this has recently been shown to lead to increased synaptotoxicity. The molecular basis for this phenomenon is unclear. Although the aggregation characteristics of Aβ(40) and Aβ(42) individually are well established, little is known about the properties of mixtures. We have explored the biophysical and structural properties of physiologically relevant Aβ(42):Aβ(40) ratios by several techniques. We show that Aβ(40) and Aβ(42) directly interact as well as modify the behavior of the other. The structures of monomeric and fibrillar assemblies formed from Aβ(40) and Aβ(42) mixtures do not differ from those formed from either of these peptides alone. Instead, the co-assembly of Aβ(40) and Aβ(42) influences the aggregation kinetics by altering the pattern of oligomer formation as evidenced by a unique combination of solution nuclear magnetic resonance spectroscopy, high molecular weight mass spectrometry, and cross-seeding experiments. We relate these observations to the observed enhanced toxicity of relevant ratios of Aβ(42):Aβ(40) in synaptotoxicity assays and in AD patients.
Journal of Biological Chemistry 12/2011; 287(8):5650-60. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Addition of divalent cations to a solution of a naphthalene-diphenylalanine that forms worm-like micelles at high pH results in the formation of a rigid, self-supporting hydrogel.
Chemical Communications 11/2011; 47(44):12071-3. · 6.17 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Alzheimer's disease is the most common form of dementia and its pathological hallmarks include the loss of neurones through cell death, as well as the accumulation of amyloid fibres in the form of extracellular neuritic plaques. Amyloid fibrils are composed of the amyloid-β peptide (Aβ), which is known to assemble to form 'toxic' oligomers that may be central to disease pathology. Aβ is produced by cleavage from the amyloid precursor protein within the transmembrane region, and the cleaved peptide may retain some membrane affinity. It has been shown that Aβ is capable of specifically binding to phospholipid membranes with a relatively high affinity, and that modulation of the composition of the membrane can alter both membrane-amyloid interactions and toxicity. Various biomimetic membrane models have been used (e.g. lipid vesicles in solution and tethered lipid bilayers) to examine the binding and interactions between Aβ and the membrane surfaces, as well as the resulting permeation. Oligomeric Aβ has been observed to bind more avidly to membranes and cause greater permeation than fibrillar Aβ. We review some of the recent advances in studying Aβ-membrane interactions and discuss their implications with respect to understanding the causes of Alzheimer's disease.
FEBS Journal 07/2011; 278(20):3905-17. · 3.79 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Amyloid-like fibrous crystals formed by the peptide KFFEAAAKKFFE have been previously characterized and provide an ideal model system to examine the importance of specific interactions by introducing specific substitutions. We find that the removal of any phenylalanine residue completely abrogates assembly ability, while charged residues modulate interactions within the structure resulting in alternative fibrillar morphologies. X-ray fiber diffraction analysis reveals that the essential backbone packing of the peptide molecules is maintained, while small changes accommodate differences in side chain size in the variants. We conclude that even very short peptides are adaptable and add to the growing knowledge regarding amyloid polymorphisms. Additionally, this work impacts on our understanding of the importance of residue composition for amyloidogenic peptides, in particular the roles of electrostatic, aromatic, and hydrophobic interactions in amyloid assembly.
Biochemistry 02/2011; 50(12):2061-71. · 3.42 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: We have previously shown that overexpressing subunits of the iron-binding protein ferritin can rescue the toxicity of the amyloid β (Aβ) peptide in our Drosophila model system. These data point to an important pathogenic role for iron in Alzheimer disease. In this study, we have used an iron-selective chelating compound and RNAi-mediated knockdown of endogenous ferritin to further manipulate iron in the brain. We confirm that chelation of iron protects the fly from the harmful effects of Aβ. To understand the pathogenic mechanisms, we have used biophysical techniques to see how iron affects Aβ aggregation. We find that iron slows the progression of the Aβ peptide from an unstructured conformation to the ordered cross-β fibrils that are characteristic of amyloid. Finally, using mammalian cell culture systems, we have shown that iron specifically enhances Aβ toxicity but only if the metal is present throughout the aggregation process. These data support the hypothesis that iron delays the formation of well ordered aggregates of Aβ and so promotes its toxicity in Alzheimer disease.
Journal of Biological Chemistry 02/2011; 286(6):4248-56. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Filamentous inclusions made of α-synuclein are found in nerve cells and glial cells in a number of human neurodegenerative diseases, including Parkinson disease, dementia with Lewy bodies, and multiple system atrophy. The assembly and spreading of these inclusions are likely to play an important role in the etiology of common dementias and movement disorders. Both α-synuclein and the homologous β-synuclein are abundantly expressed in the central nervous system; however, β-synuclein is not present in the pathological inclusions. Previously, we observed a poor correlation between filament formation and the presence of residues 73-83 of α-synuclein, which are absent in β-synuclein. Instead, filament formation correlated with the mean β-sheet propensity, charge, and hydrophilicity of the protein (global physicochemical properties) and β-strand contiguity calculated by a simple algorithm of sliding averages (local physicochemical property). In the present study, we rendered β-synuclein fibrillogenic via one set of point mutations engineered to enhance global properties and a second set engineered to enhance predominantly β-strand contiguity. Our findings show that the intrinsic physicochemical properties of synucleins influence their fibrillogenic propensity via two distinct but overlapping modalities. The implications for filament formation and the pathogenesis of neurodegenerative diseases are discussed.
Journal of Biological Chemistry 12/2010; 285(49):38555-67. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Alzheimer's disease is characterized by the aggregation and deposition of the Aβ peptide. This 40 or 42 residue peptide is the product of the proteolysis of the amyloid precursor protein membrane protein and is able to assemble to form ordered, stable amyloid fibrils as well as small, soluble, and potentially cytotoxic oligomers. The toxicity of the oligomers may be associated with the ability to bind to and affect the integrity of lipid membranes. In this novel work, we have monitored and compared the ability of the potent Aβ42 peptide, the less amyloidogenic Aβ40 peptide, and a variant with reduced amyloidogenicity to bind to and affect the integrity of membranes using dye-filled synthetic vesicles. We reveal that the potency of the assemblies reduces with incubation time of the peptide and that maximal effect occurs when a particular species is apparent by electron microscopy. We have investigated the effect of lipid vesicle composition, and our results suggest that charge on the vesicles is important and that binding may partly be mediated by the GM1 ganglioside receptors expressed in the outer leaflet of vertebrate membranes.
Langmuir 10/2010; 26(22):17260-8. · 4.19 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Incorporation of dextran into a hydrogel formed using a low molecular weight dipeptide-conjugate gelator results in controlled modification of the material properties.
Chemical Communications 09/2010; 46(36):6738-40. · 6.17 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Dipeptide-conjugates can be efficient low molecular weight hydrogelators. However, the effective design of a gelator for a specific application is compromised by the lack of a clear understanding of the design rules that govern assembly and hence gelation. Here, we report a library of naphthalene-dipeptides, their physical chemical properties, and gelation ability. We have varied both the amino acids in the dipeptides and the substitution on the naphthalene ring to allow variation of the structure throughout the molecule. We have examined the effects of these permutations on the critical micelle concentration and air-water partition coefficient at high pH and the apparent pK(a). We show that there is a clear link between these properties and the predicted hydrophobicity of the overall conjugates, rather than the properties varying with, for example, the dipeptide sequence. The majority of these dipeptide-conjugates are effective hydrogelators, although there is no apparent link between the solution properties and whether or not a conjugate is a hydrogelator. Nevertheless, where gelation occurs, the link between hydrophobicity and apparent pK(a) allows the prediction of the pH at which a gel will be formed and hence informed choice of gelator for specific applications.
Langmuir 08/2010; 26(16):13466-71. · 4.19 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: 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-beta 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.
Biophysical Journal 01/2010; 98(2):330-8. · 3.65 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The folding of a protein from a sequence of amino acids to a well-defined tertiary structure is one of the most studied and enigmatic events to take place in biological systems. Relatively recently, it has been established that some proteins and peptides are able to take on conformations other than their native fold to form long fibres known as amyloid. In vivo, these are associated with misfolding diseases, such as Alzheimer's disease, Type 2 diabetes and the amyloidoses. In vitro, peptide assembly leads to amyloid-like fibres that have high stability, resistance to degradation and high tensile strength. Remarkably, despite the lack of any obvious sequence similarity between these fibrillogenic proteins and peptides, all amyloid fibrils share common structural characteristics and their underlying structure is known as 'cross-beta'. Nature is rich in beta-sheet protein assemblies such as spider silk and other 'useful' amyloids such as curli from Escherichia coli, where the strength of fibrils is fundamental to their function.
Biochemical Society Transactions 09/2009; 37(Pt 4):671-6. · 3.71 Impact Factor
-
Benjamin M Bulheller,
Alison Rodger,
Matthew R Hicks,
Timothy R Dafforn, Louise C Serpell,
Karen E Marshall,
Elizabeth H C Bromley,
Patrick J S King,
Kevin J Channon,
Derek N Woolfson,
Jonathan D Hirst
[show abstract]
[hide abstract]
ABSTRACT: Flow linear dichroism (LD) spectroscopy provides information on the orientation of molecules in solution and hence on the relative orientation of parts of molecules. Long molecules such as fibrous proteins can be aligned in Couette flow cells and characterized using LD. We have measured using Couette flow and calculated from first principles the LD of proteins representing prototypical secondary structure classes: a self-assembling fiber and tropomyosin (all-alpha-helical), FtsZ (an alphabeta protein), an amyloid fibril (beta-sheet), and collagen [poly(proline)II helices]. The combination of calculation and experiment allows elucidation of the protein orientation in the Couette flow and the orientation of chromophores within the protein fibers.
Journal of the American Chemical Society 09/2009; 131(37):13305-14. · 9.91 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Amyloid fibril deposition is central to the pathology of more than 30 unrelated diseases including Alzheimer's disease and Type 2 diabetes. It is generally accepted that amyloid fibrils share common structural features despite each disease being characterised by the deposition of an unrelated protein or peptide. The structure of amyloid fibrils has been studied using X-ray fibre diffraction and crystallography, solid-state NMR and electron paramagnetic resonance, and many different, sometimes opposing, models have been suggested. Many of these models are based on the original interpretation of the cross-beta diffraction pattern for cross-beta silk in which beta-strands run perpendicular to the fibre axis, although alternative models include beta-helices and natively structured proteins. Here, we have analysed opposing model structures and examined the necessary structural elements within the amyloid core structure, as well as producing idealised models to test the limits of the core conformation. Our work supports the view that amyloid fibrils share a number of common structural features, resulting in characteristic diffraction patterns. This pattern may be satisfied by structures in which the strands align close to perpendicular to the fibre axis and are regularly arranged to form beta-sheet ribbons. Furthermore, the fibril structure contains several beta-sheets that associate via side-chain packing to form the final protofilament structure.
Journal of Molecular Biology 09/2009; 395(4):717-27. · 4.00 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The amyloid fibrils of familial anryloidotic polyneuropathy (FAP) are composed of genetic variants of transthyretin (TTR). Over 40 amino-acid variants have been identified associated with amyloidosis. We have performed an analysis of the effect of the different substitutions, such as location and changes in side-chain volume, hydrophilicity and charge on the TTR molecule. We have also analyzed the distribution of variants along the peptide chain. While we could demonstrate no pattern of change associated with the bulk properties of the anryloidogenic variant residues, the distribution analysis suggested a significant correlation with the position of variant residues along the polypeptide chain. We identified a significant peak in the distribution of variants which, in the folded protein chain, was associated with the edge strands (residues 45–58) of the two β-sheets that form the structural framework of the TTR molecule. In contrast, the regions of the molecule associated with the subunit interactions are almost devoid of anryloidogenic variants. To test this finding, we constructed two variant TTRS using recombinant techniques, one with a triple substitution and one with a triple deletion in the edge strand region. On expression, the variants spontaneously finned material that produced a characteristic red-shill upon Congo red staining, was composed of fibrils 80–110Å in diameter and gave β-structure X-ray patterns suggesting the formation of amyloid fibrils. We conclude that the effect of anryloidogenic variants may be to alter the structure or stability of the edge strands of the β-sheets of TTR, thus promoting the growth of inter-molecular β-sheet structures characteristic of amyloid.
07/2009; 3(2):75-85.
-
[show abstract]
[hide abstract]
ABSTRACT: Biocompatible hydrogels have a wide variety of potential applications in biotechnology and medicine, such as the controlled delivery and release of cells, cosmetics and drugs, and as supports for cell growth and tissue engineering. Rational peptide design and engineering are emerging as promising new routes to such functional biomaterials. Here, we present the first examples of rationally designed and fully characterized self-assembling hydrogels based on standard linear peptides with purely alpha-helical structures, which we call hydrogelating self-assembling fibres (hSAFs). These form spanning networks of alpha-helical fibrils that interact to give self-supporting physical hydrogels of >99% water content. The peptide sequences can be engineered to alter the underlying mechanism of gelation and, consequently, the hydrogel properties. Interestingly, for example, those with hydrogen-bonded networks of fibrils melt on heating, whereas those formed through hydrophobic fibril-fibril interactions strengthen when warmed. The hSAFs are dual-peptide systems that gel only on mixing, which gives tight control over assembly. These properties raise possibilities for using the hSAFs as substrates in cell culture. We have tested this in comparison with the widely used Matrigel substrate, and demonstrate that, like Matrigel, hSAFs support both growth and differentiation of rat adrenal pheochromocytoma cells for sustained periods in culture.
Nature Material 07/2009; 8(7):596-600. · 32.84 Impact Factor
