Further insight into the role of metals in amyloid formation by IAPP1-37 and ProIAPP1-48

Abstract

Background: IAPP 1-37 and ProIAPP 1-48 are amyloidogenic peptides implicated in β-cell death in diabetes. Interactions with metals may be involved in both the cytotoxicity of these peptides and their deposition as amyloids associated with diabetes-related pathologies. Methods: We have used the complementary methods of thioflavin T (ThT) fluorescence and transmission electron microscopy (TEM) to investigate the role of seeds and specifically metal-peptide seeds in accelerating amyloid formation by ProIAPP. In addition we have used these complementary methods alongside dynamic light scattering (DLS) to observe the dynamics of IAPP amyloid formation during the earliest phase of peptide aggregation. Results: Seeding universally resulted in an acceleration of amyloid formation, as indicated by increased ThT fluorescence, over the shorter term (minutes) while having no influence upon total amyloid deposits (no differences in ThT fluorescence) over many days. Only copper-peptide seeds were ineffective in accelerating amyloid formation above that observed for sham seeds (no peptide). Different seeding environments resulted in amyloid deposits of different fibrillar and non-fibrillar morphologies following longer term incubations regardless of the uniform nature of the respective measurements of ThT fluorescence. The aggregation dynamics of IAPP, mimicking its secretion into extracellular milieus, were complex and suggested that while metals at equimolar, generally increased rates of aggregation, with the possible exception of Cu(II), the range of sub-micron and micron-sized particles observed were not easily explained by either measurement of ThT fluorescence or imaging by TEM. Conclusions: Seeding may be significant in accelerating the formation of amyloid and in influencing the final morphologies of deposited amyloids but not in determining the total deposits of amyloid. It was of interest that copper-peptide seeds did not accelerate amyloid formation in the shorter term and this could indicate an incompatible seeding morphology due to copper? This first attempt to monitor aggregation dynamics of IAPP over only minutes has shown direct impact of metals on peptide particle size which could have implications for the cytotoxicity of IAPP in diabetes.

X-Ray Absorption Spectroscopy Measurements of Cu-ProIAPP Complexes at Physiological Concentrations

“…It remains equivocal as to whether Al(III), Fe(III) and Zn(II) promote amyloid [7], zinc [7,8] and copper [7]. It remains equivocal as to whether Al(III), Fe(III) and Zn(II) promote amyloid (β sheet) formation while it is clear that Cu(II) prevents IAPP from assembling into-sheet structures [7] as recently confirmed [9][10][11][12][13]. ProIAPP1-48 forms amyloid less readily than IAPP and while there are few data on its interactions with metals it is also the case that Cu(II) prevents ProIAPP1-48 from forming -sheets structures more prone to amyloidogenesis [12,14,15].…”

“…It remains equivocal as to whether Al(III), Fe(III) and Zn(II) promote amyloid (β sheet) formation while it is clear that Cu(II) prevents IAPP from assembling into-sheet structures [7] as recently confirmed [9][10][11][12][13]. ProIAPP1-48 forms amyloid less readily than IAPP and while there are few data on its interactions with metals it is also the case that Cu(II) prevents ProIAPP1-48 from forming -sheets structures more prone to amyloidogenesis [12,14,15]. A sketch of the process leading to the formation of ProIAPP1-48 and IAPP and of their effect on islet β cells is depicted in Figure 1.…”

“…A sketch of the process leading to the formation of ProIAPP1-48 and IAPP and of their effect on islet β cells is depicted in Figure 1. It is widely believed that the cytotoxicity of IAPP and ProIAPP1-48 is related to their propensity to form toxic oligomers during the early stages of amyloid formation [4] and it has been suggested that metals and specifically Cu(II) potentiate toxicity through stabilisation of these oligomeric forms [12,16].…”

Aggregation of biologically important peptides and proteins: inhibition or acceleration depending on protein and metal ion concentrations

“…1). 42,43 In addition, their ability to modulate the proteolytic activity of hIAPP-degrading enzymes has been extensively studied. 14,[44][45][46] It was reported that, Zn 2+ plays an important role in glycemic regulation, which is re-ected in their high concentrations in the interior of dense granule cores ranging from 10 to 20 mM, conrming their physiological importance.…”

“…Further tests in the same study using sub-stoichiometric concentrations of the metal ions conrmed the inhibitive properties of Cu 2+ , to a lesser extent for Zn 2+ , and no inuence of Al 3+ on hIAPP aggregation. 43 A recent study applied several experimental techniques such as ThT uorescence and Atomic Force Microscopy (AFM) to examine different characteristic changes of hIAPP, and Dynamic Light Scattering (DLS) analysis was used to determine the particular effects of Au 3+ complexes on the aggregation of hIAPP. 56 56 However, at low concentrations ($5 mM), it inhibited amylin oligomer formation, verifying the concentration dependence of the inhibition process.…”