How mutations in proinsulin cause diabetes: a protein-misfolding disease Insulin plays a central role in the regulation of vertebrate metabolism. The hormone, the post-translational product of a single-chain precursor (proinsulin), is a globular protein containing two chains, A (21 residues) and B (30 residues). Recent advances in human genetics have identified dominant negative mutations in the insulin gene causing permanent neonatal-onset diabetes mellitus (DM). The objective of this interdisciplinary application is to investigate the biochemical, structural, and cell-biological mechanisms of this syndrome as a model disease of protein misfolding. We hypothesize that the clinical mutations block folding of the precursor in the endoplasmic reticulum (ER) of pancreatic ?-cells. Structural analysis of the mutant proinsulins will provide insight into native determinants of foldability. Although expression of the wild-type allele would in other circumstances be sufficient to maintain homeostasis, studies of a corresponding mouse model motivate the hypothesis that the misfolded variant perturbs wild-type biosynthesis through formation of non-native aggregates containing both wild-type and mutant polypeptides. Impaired ?-cell secretion is associated with ER stress, distorted organelle architecture, and eventual cell death. To test this central hypothesis and to define the structural bases of pathological misfolding, a team has been assembled at CWRU, University of Michigan, and University of Chicago to bring to bear the combined power of biochemistry, biophysics, structural biology, cell biology, and transgenic mouse models. This collaborative proposal thus offers the exciting possibility of deciphering the molecular basis of a human disease of protein misfolding. Although neonatal diabetes is uncommon, the proposed contribution of proinsulin misfolding and ER stress to the mechanism of ?-cell dysfunction in the metabolic syndrome and type 2 diabetes mellitus extends the significance of this application to diverse human populations.
Our goal is to determine the molecular bases of toxic proinsulin misfolding in the pathogenesis of neonatal-onset diabetes mellitus due to mutations in the insulin gene. An interdisciplinary strategy is proposed that integrates structural biology with biochemistry, synthetic chemistry, and cell biology. The results promise to eludidate the structural principles of native disulfide pairing in the biosynthesis of insulin.
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