Toxic Intermediates in Islet Amyloid Formation: Analysis of IAPP Mutants Reveals a Correlation between Lag Time and Toxicity

Abstract

Islet amyloid polypeptide (IAPP, Amylin) is responsible for amyloid formation in type 2 diabetes and in islet cell transplants. The mechanism of IAPP toxicity is poorly understood. Here we identify and characterize the toxic species produced during amyloid formation by human IAPP. using a combination of time-dependent biophysical and biological assays, we show that the transiently populated pre-fibrillar intermediates formed in the early phase of amyloid formation are toxic and they are loosely packed with very modest amounts of secondary structure. The only known natural mutation found in mature human IAPP is a Ser20-to-Gly mis-sense mutation, which appears to be associated with an increased risk of early-onset type 2 diabetes. We demonstrate that the mutant accelerates amyloid formation under physiologically relevant conditions. We rationally design another variant at residue 20, S20K-IAPP, which is much slower to aggregate with an 18-fold longer lag phase and inhibits wild type amyloid formation. The pronounced effects of the Ser20 mutants highlight the sensitivity of amyloid formation to the identity of position 20. By comparing the kinetics and toxicity profiles of the two Ser20 mutants to wild type IAPP, we demonstrate that changes in the length of the lag phase directly correlate with changes in the onset and the duration of toxicity. Our findings provide direct evidence that the toxic species are transient intermediates and have implications for the treatment of type 2 diabetes and other amyloid related diseases.