Huntington<s Disease (HD) is a devastating neurodegenerative disease caused by an expanded polyglutamine tract in the huntingtin (Htt) protein, leading to its aggregation into beta-sheet-rich fibrils. Evidence suggests the 17 amino acid headpiece (N17) of Htt strongly modulates its aggregation. One goal of this research is to predict N17;s structure and role in oligomerization via molecular dynamics simulations, with experimental verification. Another goal is to leverage this structural information and experimental data to design inhibitors of Htt aggregation. Better structural understanding of the early steps of aggregation of the Htt protein would be invaluable for designing therapeutics for Huntington<s Disease. The specific aims are: 1. To investigate the structural role of the N17 headpiece in huntingtin aggregation. ! Previous MD simulations of the isolated N17 headpiece suggest that it is highly helical, and adopts two highly populated conformations consisting of one and two amphipathic helices. We will: a. Perform molecular dynamics simulations of N17-polyQ aggregation on Folding@home using Markov state models. We will investigate our hypothesis that burial of multiple N17 hydrophobic surfaces initiates aggregation of the polyQ tracts by positioning them correctly relative to each other. Based on the simulations, mutations in N17 will be proposed that would be expected to inhibit Htt aggregation. b. N17 mutations computationally predicted to disrupt aggregation will be experimentally verified, via a collaboration with Judith Frydman<s lab at Stanford. 2. To computationally design and simulate peptide inhibitors of aggregation by mimicking the structure adopted by the N17 headpiece, and verify promising inhibitory peptides experimentally. ! I will design "stapled peptide" inhibitors against Htt aggregation, specifically by modulating N17<s hypothesized role in the oligomerization process. "Stapled peptides", a recent technique pioneered by Greg Verdine<s lab at Harvard, "staples" peptides into an alpha-helical conformation. We will: a. Use "stapled peptides" to test hypotheses about the huntingtin oligomeric structure. Stapled peptides will be designed to mimic the proposed helical structure(s) of N17 in the aggregation nucleus in the MD simulations above, and simulations will be re-run with the stapled peptides. b. Design stapled peptide inhibitors of huntingtin aggregation, by incorporating N17 mutations shown experimentally to block aggregation, while maintaining the binding interface between N17 regions. c. Promising peptides will be procured and experimentally tested both in vitro and in vivo by the Frydman Lab. "Stapled peptides" have potential applications as "nanomedicine therapeutics" in inhibiting Htt aggregation. PUBLIC HEALTH RELEVANCE: Lay description:! ! Huntington<s Disease is a devastating neurodegenerative disease caused by misfolding and aggregation of the huntingtin protein. Evidence suggests the first 17 amino acids (N17) of the huntingtin protein strongly stimulate its aggregation. The goals of this research are to predict N17<s structure and role in this aggregation process computationally, with experimental verification, and then to leverage this structural information and experimental data to design and test inhibitors of huntingtin aggregation.