Resveratrol prevents amyloid fibrillation of insulin by arresting it in a bioactive oligomeric form

Insulin fibrillation is a limiting factor for its long-term storage because of considerably reduced bioavailable moieties. Deposition of fibrillated insulin can also cause subcutaneous insulin amyloidoma. Toxic phenolic compounds along with Zinc are used during commercial preparation of insulin to stabilize it in a hexameric form. Designed or repurposed natural small molecules with anti-amyloidogenic properties could thus be attractive agents for preventing insulin fibrillation. Natural polyphenolic compounds which have been shown to serve as anti-amyloid agents for proteins associated with neurodegenerative diseases are potential candidates for such function.In this study we have demonstrated that resveratrol, a natural polyphenol, can not only prevent insulin fibrillation but can also preserve insulin in a bioactive oligomeric form even at high temperature. While investigating the influence of some natural polyphenols on human insulin (hINS) in a condition inductive to amyloid fibrillation at physiological pH, we found attenuation, to different extents, of insulin fibril formation. However, visualization of polyphenol-treated hINS revealed that resveratrol in particular has the unique ability to arrest hINS before the onset of fibrillation growth in soluble oligomeric forms with discrete spherical morphology. Importantly, insulin treated with resveratrol retains its full biological activity in vivo and exerts no toxicity towards cell lines. Structural characterization of the major population of resveratrol-induced insulin oligomers by cryo-EM and single particle 3D reconstruction revealed its morphology that resembled crystal structure of insulin hexamer formulated with phenolic compounds. Thus, our study suggests that resveratrol can be an effective nontoxic substituent of phenolic compounds for insulin preservation.