Lamination and spherulite-like compaction of a hormone's native amyloid-like nanofibrils: spectroscopic insights into key interactions.

The native hierarchical self-assembly process of natural somatostatin-14, a highly aromatic and charged peptide hormone involved in various inhibitory functions, was investigated mainly using vibrational spectroscopy (ATR-FTIR and Raman scattering) combined with electron microscopy. Generic kinetic features of amyloid fibrillogenesis were confirmed for the somatostatin-14 case, together with new insights into key interactions involved in the further hierarchical assembly of the somatostatin-14 nanofibrils into i) laterally associated nanofibers and ii) spherulite-like amyloid droplets resulting from the compaction of the nanofibers. In particular, the key role of aromatic side-chains in both fibrillogenesis and the association of the nanofibrils into higher order structures could be followed. It is proposed that the compaction propensity of the somatostatin-14 nanofibrils is relevant to the current hypothesis of the biological function of hormone self-assembly processes: hormone storage inside secretory granules.

Self-assembly of the octapeptide lanreotide and lanreotide-based derivatives: the role of the aromatic residues.

We investigated the spectroscopic properties of the aromatic residues in a set of octapeptides with various self-assembly properties. These octapeptides are based on lanreotide, a cyclic peptide analogue of somatostatin-14 that spontaneously self-assembles into very long and monodisperse hollow nanotubes. A previous study on these lanreotide-based derivatives has shown that the disulfide bridge, the peptide hairpin conformation and the aromatic residues are involved in the self-assembly process and that modification of these properties either decreases the self-assembly propensity or modifies the molecular packing resulting in different self-assembled architectures. In this study we probed the local environment of the aromatic residues, naphthyl-alanine, tryptophan and tyrosine, by Raman and fluorescence spectroscopy, comparing nonassembled peptides at low concentrations with the self-assembled ones at high concentrations. As expected, the spectroscopic characteristics of the aromatic residues were found to be sensitive to the peptide-peptide interactions. Among the most remarkable features we could record a very unusual Raman spectrum for the tyrosine of lanreotide in relation to its propensity to form H-bonds within the assemblies. In Lanreotide nanotubes, and also in the supramolecular architectures formed by its derivatives, the tryptophan side chain is water-exposed. Finally, the low fluorescence polarization of the peptide aggregates suggests that fluorescence energy transfer occurs within the nanotubes.

Molecular origin of the self-assembly of lanreotide into nanotubes: a mutational approach.

Lanreotide, a synthetic, therapeutic octapeptide analog of somatostatin, self-assembles in water into perfectly hollow and monodisperse (24-nm wide) nanotubes. Lanreotide is a cyclic octapeptide that contains three aromatic residues. The molecular packing of the peptide in the walls of a nanotube has recently been characterized, indicating four hierarchical levels of organization. This is a fascinating example of spontaneous self-organization, very similar to the formation of the gas vesicle walls of Halobacterium halobium. However, this unique peptide self-assembly raises important questions about its molecular origin. We adopted a directed mutation approach to determine the molecular parameters driving the formation of such a remarkable peptide architecture. We have modified the conformation by opening the cycle and by changing the conformation of a Lys residue, and we have also mutated the aromatic side chains of the peptide. We show that three parameters are essential for the formation of lanreotide nanotubes: i), the specificity of two of the three aromatic side chains, ii), the spatial arrangement of the hydrophilic and hydrophobic residues, and iii), the aromatic side chain in the beta-turn of the molecule. When these molecular characteristics are modified, either the peptides lose their self-assembling capability or they form less-ordered architectures, such as amyloid fibers and curved lamellae. Thus we have determined key elements of the molecular origins of lanreotide nanotube formation.

Spontaneous fibrillation of the native neuropeptide hormone Somatostatin-14.

Natural Somatostatin-14 is a small cyclic neuropeptide hormone with broad inhibitory effects on endocrine secretions. Here we show that natural Somatostatin-14 spontaneously self-assembles in water and in 150 mM NaCl into liquid crystalline nanofibrils, which follow characteristic structural features of amyloid fibrils. These non-covalent highly stable structures are based on the Somatostatin native backbone conformation and are formed under non-denaturing conditions. Our results support the hypothesis that self-assembly into amyloid fibrils is a generic property of the polypeptide chain under appropriate conditions. Given recent advances on the mechanisms of biological storage and sorting modes of peptide/protein hormones into secretory granules, we propose that Somatostatin-14 fibrillation could be relevant to the regulated secretion pathway of this neuropeptide hormone. Such a hypothesis is consistent with the emerging concept of the existence of non-disease related but functional amyloids.

Biomimetic organization: Octapeptide self-assembly into nanotubes of viral capsid-like dimension.

The controlled self-assembly of complex molecules into well defined hierarchical structures is a promising route for fabricating nanostructures. These nanoscale structures can be realized by naturally occurring proteins such as tobacco mosaic virus, capsid proteins, tubulin, actin, etc. Here, we report a simple alternative method based on self-assembling nanotubes formed by a synthetic therapeutic octapeptide, Lanreotide in water. We used a multidisciplinary approach involving optical and electron microscopies, vibrational spectroscopies, and small and wide angle x-ray scattering to elucidate the hierarchy of structures exhibited by this system. The results revealed the hexagonal packing of nanotubes, and high degree of monodispersity in the tube diameter (244 A) and wall thickness (approximately equal to 18 A). Moreover, the diameter is tunable by suitable modifications in the molecular structure. The self-assembly of the nanotubes occurs through the association of beta-sheets driven by amphiphilicity and a systematic aromatic/aliphatic side chain segregation. This original and simple system is a unique example for the study of complex self-assembling processes generated by de novo molecules or amyloid peptides.