Structural mechanism of diabetic amyloid formation

University Of Wisconsin-Madison, Madison WI

Investigators

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Abstract

Structural mechanism of diabetic amyloid formation Abstract 11.6% of the U.S. population is afflicted by type 2 diabetes. It starts as insulin resistance, but ultimately the pancreatic -cells that make insulin fail, resulting in overt diabetes. -cell failure correlates with aggregation of a hormone called the human islet amyloid polypeptide (hIAPP or amylin) into amyloid plaques in the islets of the -cells. Surprisingly, the amyloid fibers themselves are not cytotoxic. Many researchers believe that the toxic species are oligomers of hIAPP that interfere with receptor mediated processes, cause inflammation, or permeabilize the membrane. The oligomer hypothesis is strengthened by recently reported antibodies that bind specifically to oligomers and stall type 2 diabetes in animal models. As a result, there is much interest in elucidating the structures of oligomers and understanding the mechanism by which they form. However, characterizing kinetically evolving proteins is challenging and so little structural information exists about these transient oligomers. We invented a technology, rapid-scan 2D IR spectroscopy, that enables us to monitor the structure of hIAPP as it aggregates. We discovered an oligomeric intermediate with a parallel -sheet in the FGAIL region and an -helix at the N-terminus. We observed this “FGAIL oligomer” in 4 different mammalian species known to contract type 2 diabetes, strengthening our hypothesis that this intermediate is a key player in the disease. Most importantly, we realized that we could trap the oligomer with a few benignly placed mutations. Our trapped oligomers are nearly as toxic as wild-type hIAPP, but persist in vitro for days rather than hours. Because they are stable for so long, we could perform 2D/3D NMR and generate the first structural model of an hIAPP oligomer. Simultaneously, we created a transgenic mouse model that expresses our oligomer, thereby linking our in vitro oligomer to pathophysiology. In this proposal, we capitalize on our new strategy for studying the structures and mechanisms of hIAPP oligomers. Aim 1 refines our structural model, Aim 2 extends our approach to a hereditary missense mutation that causes early onset-diabetes, and Aim 3 tests our in vitro hypotheses with matching transgenic mouse models. We seek to understand hIAPP aggregation from a fundamental perspective and link our in vitro structural mechanisms to in vivo physiology. We believe that our approach is unique to the structural biology community, providing much desired structural and mechanistic information about hIAPP aggregation and its relationship to type 2 diabetes.