Pre-Amyloid States of Islet Amyloid Polypeptide Examined by Single-Particle Fluorescence

Abstract

Islet amyloid polypeptide (IAPP or amylin) is a peptide hormone cosecreted with insulin by the pancreas that displays potent amyloidogenic activity. In vitro studies demonstrate that IAPP is capable of disrupting lipid bilayers, suggesting a possible mechanism for IAPP-induced beta-cell death in Type II Diabetes Mellitus. Of particular interest are oligomeric IAPP species, which are believed to mediate membrane leakage, as well as to be intermediates in amyloid formation. IAPP oligomers are likely to be transient and heterogeneous, and so a detailed dynamic and functional characterization of these critical structures has been challenging. We have used single-molecule Forster resonance energy transfer (FRET) to study IAPP conformations in solution; bound to model membranes; and in the presence of insulin, which exerts a cytoprotective effect. Intermolecular FRET was used to characterize the topology, packing and dimensions of IAPP oligomers. FRET measurements provided experimental constraints for atomistic modeling and MD simulations both of IAPP monomers and oligomers. Simultaneously, we have used fluorescence correlation spectroscopy (FCS) to explore the interactions of IAPP with lipid bilayers, probing the thermodynamic landscape of membrane-catalyzed IAPP oligomer assembly. Single-molecule techniques provide unique experimental insight into the processes governing the membrane binding and aggregation of IAPP and similar amyloidogenic peptides.