title: 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 20 mM Tris-HCl. Dissolution of synthetic rat amylin 1-37 and 8-37 peptides in H(2)O or phosphate buffered saline failed to produce fibrils. Addition of 20 mM Tris-HCl to synthetic rat amylin 1-37 and 8-37 peptides initially dissolved in H(2)O 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.
authors.affiliation: Department of Human & Health Sciences, School of Life Sciences, University of Westminster, London W1W 6UW, UK. n.milton@westminster.ac.uk
authors.forename: Nathaniel G N
authors.initials: NG
authors.lastname: Milton
authors.forename: J Robin
authors.initials: JR
authors.lastname: Harris
mesh_terms: D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000682:Amyloid / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism / Q000633:toxicity; D058227:Amyloidogenic Proteins; D000818:Animals; D002374:Catalase / Q000378:metabolism; D002478:Cells, Cultured; D006801:Humans; D058228:Islet Amyloid Polypeptide / Q000378:metabolism*; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D018696:Neuroprotective Agents / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D051381:Rats
chemical_list: D000590:Amino Acid Chloromethyl Ketones; D000682:Amyloid; D058227:Amyloidogenic Proteins; D058228:Islet Amyloid Polypeptide; D018696:Neuroprotective Agents; D010446:Peptide Fragments; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C087166:AC 187; D002374:Catalase