Investigation of the role of the N17 headpiece in huntingtin aggregation

描述(申请人提供)：亨廷顿S病(HD)是一种毁灭性的神经退行性疾病，由亨廷顿蛋白(Htt)中扩张的聚谷氨酰胺束引起，导致其聚集成富含β-折叠的纤维。有证据表明，Htt的17个氨基酸头饰(N17)强烈调节其聚集。本研究的目标之一是通过分子动力学模拟预测N17；S的结构和在齐聚中的作用，并进行实验验证。另一个目标是利用这些结构信息和实验数据来设计Htt聚集的抑制剂。更好地了解Htt蛋白聚集的早期步骤的结构对于设计亨廷顿和S病的治疗方法将是非常宝贵的。具体目的是：1.研究N17头饰在亨廷顿蛋白聚集中的结构作用。好了！以前对孤立的N17头盔进行的MD模拟表明，它是高度螺旋的，并采用了由一个和两个两亲性螺旋组成的两个高度聚集的构象。我们将：a.使用马尔可夫状态模型对Folding@Home上的N17-PolyQ聚集进行分子动力学模拟。我们将调查我们的假设，即多个N17疏水表面的埋葬通过正确地定位它们彼此之间的位置来启动多聚Q区段的聚集。基于模拟，N17的突变将被提出，预计将抑制Htt聚集。B.通过计算预测会破坏聚集的N17突变将通过与斯坦福大学朱迪思·弗莱德曼和S实验室的合作进行实验验证。2.模仿N17头戴式耳机的结构，对聚集肽抑制剂进行了计算机设计和模拟，并通过实验验证了有希望的抑制肽。好了！我将通过调节N17&Lt；S假设的在齐聚过程中的作用来设计针对Htt聚集的“装订多肽”抑制剂。哈佛大学的格雷格·韦尔丁和S实验室最近首创了一项名为“装订的多肽”的技术，它将多肽“装订”成一种α-螺旋构象。我们将：a.使用“装订的多肽”来测试关于亨廷顿蛋白寡聚体结构的假设。在上面的MD模拟中，装订的多肽将被设计成模仿N17在聚集核中的螺旋结构(S)，并将用装订的多肽重新运行模拟。B.设计亨廷顿蛋白聚集的装订多肽抑制剂，通过掺入实验证明的N17突变来阻断聚集，同时维持N17区域之间的结合界面。C.弗莱德曼实验室将在体外和体内获得有希望的多肽，并进行实验测试。“装订多肽”作为“纳米药物疗法”在抑制Htt聚集方面具有潜在的应用前景。 公共卫生相关性：外行描述：！！亨廷顿S病是一种由亨廷顿蛋白错误折叠和聚集引起的破坏性神经退行性疾病。有证据表明，亨廷顿蛋白的前17个氨基酸(N17)强烈刺激其聚集。本研究的目标是通过计算和实验验证来预测N17和S在这一聚集过程中的结构和作用，然后利用这些结构信息和实验数据来设计和测试亨廷顿蛋白聚集的抑制剂。

DESCRIPTION (provided by applicant): 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.