Self-assembly of β-turn forming synthetic tripeptides into supramolecular β-sheets and amyloid-like fibrils in the solid state
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Introduction
Oligopeptides with appropriate conformations can self-assemble to form many regular structures such as sheets, ribbons, rods and tubes which have numerous applications in biological and material sciences.1., 2. For example, Ghadiri and his coworkers have established that self-assembling cyclic peptides form hollow nanotubes, which can act as artificial ion channels and biosensors.3 Zhang and his colleagues have shown that a self-assembling peptide scaffold can serve as a substrate for neurite outgrowth and synapse formation and this type of biologically compatible scaffold is also important for tissue repair and tissue engineering.4 Self-assembling peptides sometimes form gels when they encapsulate solvent molecules under suitable conditions.5 Very recently, Artzner and his co-workers have demonstrated that self-assembly of a synthetic therapeutic octapeptide, Lanreotide, leads to the formation of supramolecular β-sheets which upon further self-assembly ultimately form monodisperse nanotubes in water with diameters that are tunable by suitable modifications in the molecular structure.6 Higher order molecular self-assembly of a peptide into a β-sheet structure is not only important for designing biomaterials, but also useful in studying pathogenesis of certain age-related disease causing fibrils where self-assembly of mis-folded proteins or protein fragments leads to the formation of the aggregated mass that is known as amyloid fibrils.7 The supramolecular β-sheet stabilization and consequent insoluble amyloid plaque formation are associated with several neurodegenerative diseases including Alzheimer's disease8 and Prion-protein diseases.9 Sequences and three dimensional structures of disease-causing amyloid proteins and/or protein fragments are enormously varied. However, they self-assemble into supramolecular β-sheets and consequently form protease resistant amyloid fibrils and exhibit similar physicochemical properties (viz.: congophilicity, binds to thioflavin T).10 The therapeutic challenge in all forms of these fatal neurodegenerative diseases is to prevent amyloid fibril formation, a goal that requires a detailed understanding of the pathway(s) of β-sheet aggregation as well as fibrillation. Recently, many research groups also have used self-assembling, β-sheet forming peptides as amyloid fibril inhibitors.11
Previously, we have demonstrated that short peptides with extended backbone conformations can self-assemble to form supramolecular β-sheet structures in crystals and amyloid-like fibrils in the solid state.12 As Aβ-peptides (Amyloid β-peptide) contain many short loops and turn conformations in their backbones,13 self-assembly of turn-forming peptides is important for model studies. Recently, Kirschner and his co-workers measured the powder diffraction patterns of a solubilized and dried Aβ31-35 sample and demonstrated that this peptide adopts an intramolecular hydrogen bonded reverse-turn conformation which is important for amyloid fibril formation and its cytotoxicity.14 However, the crystal structures of model peptides which form an intermolecularly hydrogen-bonded supramolecular β-sheet from a turn-forming semi-cyclic peptide backbone are still rarely obtained. In our very recent communication, we have demonstrated that a β-turn forming peptide can form supramolecular β-sheet structure through self-aggregation and can exhibit amyloid-like fibrillar morphology in the solid state.15 Continuing our research in this field, we report here the results of our studies on three synthetic terminally blocked tripeptides, Boc-Ala(1)-Aib(2)-Val(3)-OMe 1, Boc-Ala(1)-Aib(2)-Ile(3)-OMe 2 and Boc-Ala(1)-Gly(2)-Val(3)-OMe 3 which all adopt reverse turn conformations and self-assemble to form supramolecular β-sheets in crystals and amyloid-like fibrils in the solid state.
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Results and discussion
Peptides 1 and 2 contain the centrally located conformationally constrained Aib (α-aminoisobutyric acid) residue together with hydrophobic Val (valine) and Ile (isoleucine) residues at the C terminus and both adopt a turn structure. In tripeptide 3, the centrally positioned Aib has been substituted by the structurally very flexible Gly (glycine) residue with the aim of investigating whether or not peptide 3 forms the folded turn conformation. These peptides were studied using X-ray
Conclusion
In spite of having different sequences and compositions, all three reported peptides adopt a distorted β-turn conformation in their crystal structures. Moreover, these turn-forming peptides self-associate to form supramolecular β-sheet structures via non-covalent interactions including intermolecular hydrogen bonds, in which all the hydrogen bonds are formed between the peptide linkages. These turn-forming peptides also form amyloid-like fibrils upon further self-aggregation, as is evident from
Peptide synthesis
The tripeptides were synthesized by conventional solution phase methods using racemization-free fragment condensation strategy.18 The Boc group was used for N-terminal protection and the C terminus was protected as a methyl ester. Couplings were mediated by dicyclohexylcarbodiimide-1-hydroxybenzotriazole (DCC/HOBt). All intermediates have been characterized by 1H NMR (300 MHz) and thin layer chromatography (TLC) on silica gel and used without further purification. The final products were
Acknowledgements
We thank EPSRC and the University of Reading, UK for funds for the Image Plate System. We also acknowledge Department of Science and Technology, New Delhi, India for the grant No. SR/S5/OC-29/2003.
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