A dimer model of human calcitonin13-32 forms an α-helical structure and robustly aggregates in 50% aqueous 2,2,2-trifluoroethanol solution

Abstract

Determining the cause of human calcitonin (hCT) aggregation could be of help in the effort to utilize hCT for treatment of hypercalcemia. Here we report that a dimer model of hCT13-32 aggregated to a greater degree than native hCT under aqueous 2,2,2-trifluoroethanol conditions. Analyses using circular dichroism spectroscopy, thioflavine-T binding assays and atomic force microscopy suggest that the α-helical portion of hCT is important for initiation of the aggregation process, which yields long fibrils. Dimerization, which stabilizes the β-sheet structure of hCT, enhances aggregation potency. Dimerization of hCT stabilizes the α-helix under aqueous TFE conditions, leading to the long fibril formation. Up to now, there have been no reports of using a dimer model to investigate the properties of hCT aggregation. Our findings could potentially serve as the basis for development of novel hCT derivatives that could be utilized for treatment of hypercalcemia, as well as for development of novel therapeutics for other ailments caused by amyloid peptides. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

Flavonoids with Vicinal Hydroxyl Groups Inhibit Human Calcitonin Amyloid Formation

“…Notably, after reaching the maximum, the fluorescence intensity slowly declines over time. This is consistent with other reports where the decrease of the signal is likely attributed to the precipitation of hCT aggregates over the incubation period . When 5 μ m EGCG is added, there is a ≈50 % decrease in the maximum ThT fluorescence intensity compared to that of hCT only (Figure B).…”

“…This is consistent with other reports where the decrease of the signali s likely attributed to the precipitation of hCT aggregates over the incubation period. [36][37] When 5 mm EGCG is added, there is a % 50 %d ecreasei nt he maximum ThT fluorescencei ntensity compared to that of hCT only ( Figure 1B). In the presence of 50 mm EGCG,t he maximum fluorescencei ntensity is reduced by % 80 %.…”

Structural Biology of Calcitonin: From Aqueous Therapeutic Properties to Amyloid Aggregation

“…Despite this variability in fibrillation, α‐helical intermediates have been observed under a range of conditions, and play a significant role in the biological function of CT ,,,,. Regarding the α‐helical intermediate, it is reported that dimerization of hCT stabilize the α‐helix under aqueous TFE solutions, leading to the long fibril formation . Within individual CT monomers, there are often multiple secondary structural motifs present that depend on the aggregation environment and could influence peptide behavior, but given the ubiquity of α‐helical intermediates, it seems generally accepted that they are a key species in the in vivo aggregation of CT ,,…”

Effects of disulfide bond and cholesterol derivatives on human calcitonin amyloid formation

“…Our result is consistent with the study of Kawashima et al . who also reported the loss of CD signal of hCT after aggregation, likely attributed to the precipitation of hCT aggregates over the course of incubation . This may also account for the decrease of ThT fluorescence intensity observed in the aggregation kinetics result (Figure A).…”