The Effects of Naturally Occurring Phenols on Amyloid Fibrils from Insulin in the Liquid Phase Using Atomic Force Microscopy

Objectives Amyloid fibrils are misfolded proteins that self-assemble to form insoluble, elongated, β-sheet structures. These fibrils are linked with the progression of neurodegenerative diseases like Alzheimer's Disease and metabolic disorders such as Type II diabetes. A range of molecules have shown the ability to impede the formation of these fibrils, often by acting on the monomeric and oligomeric forms of the protein. Our work focuses on the inhibitory effect of curcumin, gallic acid and mangiferin on the formation of amyloid fibrils. Materials and methods Bovine insulin was used as a model system for amyloid formation by lowering the pH at 60 °C, and several different small molecules were used to test inhibition. Samples were analyzed using Atomic Force Microscopy (AFM) in a liquid environment, as well as Congo Red and Thioflavin T assays. Results We have been able to observe significant inhibition of amyloid fibril formed from insulin by these compounds as well as observe beginning stages and the morphology of fibrils. Mangiferin, gallic acid, and curcumin were all shown to slow down fibril formation, though by 5 days fibrils have started to form in the gallic acid solution. After 7 days, solutions containing curcumin or mangiferin still show no strong evidence of structured fibrils. Conclusion Gallic acid, curcumin and mangiferin are all molecules that have inhibitory effects on amyloid formation using insulin as a model system, with gallic acid being somewhat less effective than the other two. The AFM technique used to image fibrils was performed in an environment that more closely resembles in vivo conditions than air dried samples.