Abstract
Most amyloid assemblies are seen as irreversible and exhibit polymorphism because their assembly is kinetically controlled and different structures are trapped during the aggregation process. However, in the specific case of peptide hormones, formation of amyloid assemblies for storage purposes has been reported. This suggests a strict control of assembly and the ability to disassemble upon hormone secretion. In the present work, we have sought to test these assertions with a short peptide, the cholecystokinin (or gastrin) tetrapeptide (CCK-4), that has been found in both gastrointestinal tract and central nervous system, and whose sequence is shared by a large number of hormones. We have thus studied in vitro this peptide's self-assembling properties in dense phases at different pH levels, thus mimicking in vivo storage conditions. The solubility and morphology of the supramolecular assemblies have been shown to vary with the pH. At low pH, the tetrapeptide exhibits a low solubility and forms microcrystals. At higher pH levels, peptide solubility increases and above a high enough concentration, peptide monomers self-assemble into typical amyloid fibrils of 10-20 nm diameter. The physical network formed by these fibrils results in a birefringent hydrogel phase. Despite the different morphological features exhibited at different pH, structural analysis shows strong similarities. Both supramolecular assemblies-microcrystals and fibrils-are structured by β-sheets. We also show that all these morphologies are reversible and can be either dissolved or changed into one another by switching the pH. In addition, we demonstrate that a modification in the charge sequence of the peptide by amino acid mutation modifies its self-assembly properties. In conclusion, just as the CCK-4 sequence is the minimal sequence required for a complete biological activity at CCKB receptors in the brain, it is also sufficient to form amyloid fibers whose properties can be related to hormone storage and release purposes in vivo.
Highlights
Until recently, the study of amyloid fibril assembly and structure has mainly been motivated by the will to understand the formation of pathological aggregations as observed in neurodegenerative diseases such as Alzheimer’s, Parkinson’s or Huntington’s
With only four amino-acids in its sequence the Cholecystokinin/Gastrin tetrapeptide can attain different conformations that lead to polymorphic supramolecular assemblies
By using rather extreme pH conditions, we show that it is possible to choose one or the other morphology
Summary
The study of amyloid fibril assembly and structure has mainly been motivated by the will to understand the formation of pathological aggregations as observed in neurodegenerative diseases such as Alzheimer’s, Parkinson’s or Huntington’s. Values reported for buried tryptophan residues are closer to 330 nm.[50] this value is significantly shifted with respect to the ones measured for the monomers in solutions (366 nm) which are similar to those reported for the amino acid alone in solution.[51] As for the intensity of the emission peak, it decreases as the proportion of assembled peptides increases.
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