Insulin amyloid fibrillation at above 100 degrees C: New insights into protein folding under extreme temperatures

Abstract

To investigate the folding behavior of amyloidogenic proteins under extreme temperatures, the kinetics of fibrillation and accompanying secondary structure transitions of bovine insulin were studied for temperatures ranging tip to 140degreesC. The presence of extreme heat stress had. traditionally been associated with irreversible denaturation of protein while the initial steps of such a denaturation process may be common with a fibril formation pathway of amyloidogenic proteins. The present work demonstrates the ability of insulin to form amyloid fibrils at above 100degreesC. Amyloid formation was gradually replaced by random coil generation after similar to80degreesC until no amyloid was detected at 140degreesC. The morphology of insulin amyloid fibrils underwent sharp changes with increasing the temperature. The dependence of amyloid formation rate on incubation temperature followed non-Arrhenius kinetics, which is explained by temperature-dependent enthalpy change for amyloid formation. The intermediate stage of amyloid formation and random coil generation consisted of a partially folded intermediate common to both pathways. The fully unfolded monomers in random coil conformation showed partial reversibility through this intermediate by reverting back to the amyloid pathway when formed at 140degreesC and incubated at 100degreesC. This Study highlights the non-Arrhenius kinetics of amyloid fibrillation under extreme temperatures, and elucidates its intermediate stage common with random coil formation.