(631f) Formation of Amyloid Fibrils By Protein Fragments

Many diseases such as Alzheimer’s and Parkinson’s are linked to amyloid deposits, which are insoluble protein aggregates with a characteristic intermolecular beta-sheet structure. The buildup of these deposits is caused by the aggregation of the amyloid peptide that is found in the human body. It is known that aggregate formation damages tissue, but the kinetics of their formation is not well understood. One protein that also forms amyloid fibrils is insulin.  It is known that bovine and human insulin have different fibrillation kinetics in spite of the fact of being almost identical molecules (they differ in four amino acids).  In a previous study, we have found that bovine insulin  has a shorter lag time and a faster fibrillation rate than human insulin. 

In this work the fibrillation of bovine and human insulin fragments, which include the region lacking homology, was studied.  The sequences used were Asn-Glu-Leu-Gln-Tyr-Leu-Ser-Cys-Ile-Ser-Thr for the human insulin fragment and Asn-Glu-Leu-Gln-Tyr-Leu-Ser-Cys-Val-Ser-Ala for the bovine insulin fragment.  In both fragment synthesis, the peptide was had a lysine tail in an attempt to increase its solubility. 

Fourier Transform Infrared (FTIR) Spectroscopy, Transmission Electron Microscopy (TEM), and Thioflavin-T (ThT) Fluorescent Spectroscopy were used to explore the differences in kinetics of these two fragments. We have found that both fragments form aggregates rich in beta sheet structure but some TEM micrographs suggest that they are forming spherulites rather than fibrils.  The aggregation process is very slow taking several months to form large aggregates.  Our results also show that the bovine fragment has a faster kinetics than the human one.