Fibril formation and toxicity of the non-amyloidogenic rat amylin peptide

Abstract

Full-length native rat amylin 1–37 has previously been widely shown to be unable to form fibrils and to lack the toxicity of the human amylin form leading to its use as a non-amyloidogenic control peptide. A recent study has suggested that rat amylin 1–37 forms amyloidogenic β-sheet structures in the presence of the human amylin form and suggested that this property could promote toxicity. Using TEM analysis we show here fibril formation by synthetic rat amylin 1–37 and 8–37 peptides when the lyophilized HPLC purified peptides are initially dissolved in 20mM Tris–HCl. Dissolution of synthetic rat amylin 1–37 and 8–37 peptides in H2O or phosphate buffered saline failed to produce fibrils. Addition of 20mM Tris–HCl to synthetic rat amylin 1–37 and 8–37 peptides initially dissolved in H2O also failed to induce fibril formation. The rat amylin fibrils have a uniform structure and bind Congo red suggesting that they are amyloid fibrils. The rat amylin fibrils also bind catalase, which could be inhibited by Amyloid-β 31–35 and a catalase amyloid-β binding domain-like peptide (R9). The rat amylin 1–37 and 8–37 fibrils are toxic in both human pancreatic islet and neuronal cell culture systems. The toxicity of rat amylin fibrils can be inhibited by an amylin receptor antagonist (AC187) and a caspase inhibitor (zVAD-fmk) in a similar manner to previous observations for human amylin toxicity. Chemically induced rat amylin fibril formation of uniform structured fibrils provides a potentially novel anti-amyloid drug discovery tool.