Amyloid Inhibitors

There is great interest in developing inhibitors of amyloid fibril formation so as to help combat diseases such as Alzheimer’s diseases and type 2 diabetes. We combine 2D IR spectroscopy and residue-specific isotope labeling to investigate the interactions between amyloid peptides and amyloid inhibitors, using many of the same techniques developed for studying amyloid folding.

The frequency shifts of 13C=18O isotope labels can be used to detect which residues in an amyloid peptide are incorporated in to amyloid β-sheets. The loss of these shifts for certain residues when the inhibitor is present indicates that the inhibitor prevents the formation of β-sheets at that residue.

Schematic of Rat Inhibition

Figure 1. (Top) Human amylin forms fibrils with β-sheets that induces a shift in the frequency of the isotope label peak. (Bottom) When mixed with the rat amylin inhibitor, the N-terminal β-sheet is prevented from forming at 8 hours, as indicated by the lack of a frequency shift. However, by 24 hours the inhibitor no longer prevents the β-sheet formation and instead forms β-sheets of its own likely on the exterior of the human fibrils.

We used this approach to investigate the inhibition behavior of an inhibitor of human amylin, the peptide implicated in type 2 diabetes. We found that the behavior of the inhibitor is quite complex and surprising. While expected (based on sequence) to disrupt the C-terminal β-sheet, we found that the inhibitor disrupts the N-terminal β-sheet while leaving the C-terminal largely intact. Furthermore, we found that eventually human amylin overcomes the inhibitor and forms the N-terminal β-sheet anyway. In the process, the inhibitor forms its own β-sheets, likely on the outside of the human fibrils. These studies demonstrate how 2D IR spectroscopy can help better understand the behavior of amyloid inhibitors even in very complex cases.