(756d) Kinetics of Bovine and Human Insulin Amyloid Fibril Formation In Bulk and In the Presence of Solid/Liquid Interfaces
Amyloid fibrils are ordered aggregates of peptides or proteins that are associated with many diseases (e.g. type 2 diabetes mellitus, Alzheimer’s disease, Parkinson’s disease, etc). Amyloid diseases are apparently unconnected; however, intermolecular secondary structure (mostly β-sheets) is present in all amyloid aggregates. One protein that forms amyloid fibrils is insulin. Despite the fact that insulin aggregation has been extensively studied, the differences in the aggregation behavior between proteins from different species have not been explored.
The objective of this work is to explore the effect of solid/liquid interfaces on the aggregation behavior of bovine and human insulins. The rate and extent of aggregation of both proteins in bulk and in the presence of either polystyrene microspheres with different surface chemistry or liposomes of different compositions were studied. The aggregation was followed by turbidity and dynamic light scattering. The Congo red assay and FT-IR were used corroborate the formation of fibrils after they reached a constant length.
The surfaces were either amino, carboxy, and hydroxyl polystyrene microbeads or the following liposomes: 1) 80%/20% PC/PS; 2) 20%/80% PC /Cholesterol; 3) 80%/20% PC /Cholesterol; 4) 2:2:1:1 Cholesterol/PC/PG/PE; and 5) 10:5:7.5:16 PC/PE/PS/Cholesterol.
It was found that bovine insulin forms amyloid fibrils faster than human insulin. Amino, carboxy, and hydroxy polystyrene microbeads shorted the lag time of human insulin by 20 minutes. However, amino, carboxy, and hydroxy polystyrene extended the lag time of insulin by 20 minutes. The lag time of bulk bovine insulin was not affected by liposome 5. The lag time of bulk human insulin was only affected by liposome 2 which has the highest cholesterol content.
Some of the results may be explained by considering the (minute) differences in the polypeptide chains of both proteins. Bovine insulin differs from human insulin in three of its amino acids; alanine substitutes threonine twice, and valine substitutes isoleucine once. It is known that Thr, Val and Ile are “beta sheet” forming amino acids but Ala is an alpha helix forming one. Moreover, aromatic amino-acids, like Thr, are found to have the highest amyloidogenic propensity.