Molecular aspects of copper and zinc binding to the prion protein

 描述(申请人提供)：这项研究计划集中在蛋白(PrP)对铜和锌的吸收，以及金属结合在PrP结构、调节、体内平衡和神经退行性疾病中的作用。PrP折叠错误的脑沉积导致了一类显着的神经退行性疾病，称为传染性海绵状脑病(TSE)。TSE与流行的、与年龄相关的神经退行性疾病，包括阿尔茨海默氏症和帕金森氏病，具有共同的病理基础。虽然PrP最初被认为在神经退行性疾病中起作用，但现在很清楚PrP对维持神经元功能是必不可少的。PrP在对PrP活性至关重要的位置同时吸收铜和锌。通过之前的资助阶段，该计划充分阐明了铜和锌结合部位的结构和热力学。最近，发现了一种全新的锌离子促进的相互作用，在这种相互作用中，金属离子驱动PrP的N端和C端结构域之间的接触。这一发现表明，许多遗传性Pron疾病可能是由于这一新发现的结构的减弱而引起的。也是在上一个供资期间，确定了新的a-卵裂机制和地点，从根本上改变了PrP条例中的概念。拟议的工作将以这些调查结果为基础，有以下三个目标。1)新的结构研究将探讨铜如何影响PrP三级接触，家族突变如何调节这种相互作用，以及PrP N末端是否有助于金属离子促进的稳定。2)神经退行性变研究的新发现表明，PrP是Abeta寡聚体的主要受体，与阿尔茨海默病有关。有趣的是，Abeta结合位点位于易受a-切割的PrP片段的近端。这个项目将进一步研究-裂解是如何调节的，以及Abeta等有毒物种是否会阻止酶进入裂解位点。3)PrP引起的非调节性跨膜电流可引起早期细胞损伤。几个实验室的研究表明，这些电流来自一个调控不佳的PrP N-末端结构域。这将通过使用全细胞膜片钳电生理学来确定PrP结构、a-裂解和Abeta结合的影响而被共同检测。总之，这些实验将测试一个模型，在该模型中，PrP必须在金属离子动态平衡中受到严格调控，以及调控缺失可能会如何导致Prion和阿尔茨海默病。

 DESCRIPTION (provided by applicant): This research program focuses on copper and zinc uptake by the prion protein (PrP), and the role of metal binding in PrP structure, regulation, homeostasis, and neurodegenerative diseases. Brain deposits of misfolded PrP are responsible for a remarkable class of neurodegenerative diseases termed the Transmissible Spongiform Encephalopathies (TSEs). TSEs share pathologies with prevalent, age-related, neurodegenerative illnesses including Alzheimer's and Parkinson's disease. Although originally recognized for its role in neurodegenerative disease, it is now clear that PrP is essential for sustaining neuron function. PrP takes up both copper and zinc at sites that are essential for PrP activity. Through previous funding periods, this program fully elucidated the structure and thermodynamics of the copper and zinc binding sites. Most recently, a fundamentally new Zn2+ promoted interaction was discovered in which the metal ion drives contact between PrP's N-terminal and C-terminal domains. This finding suggests that many inherited prion diseases may arise from a weakening of this newly identified structure. Also in the last funding period, new mechanisms and sites for a- cleavage were identified, fundamentally modifying concepts in PrP regulation. The proposed work will build on these findings with the following three aims. 1) New structural studies will examine how copper influences PrP tertiary contacts, how familial mutations modulate this interaction, and whether the PrP N-terminus contributes to metal ion promoted stabilization. 2) New findings in the study of neurodegeneration suggest that PrP is a primary receptor for Abeta oligomers, implicated in Alzheimer's disease. Interestingly, the Abeta binding site is proximal to the PrP segment susceptible to a-cleavage. This project will further examine how -cleavage is regulated and whether toxic species, such as Abeta, occlude enzymatic access to the cleavage sites. 3) Early stage cellular damage arises from unregulated transmembrane currents caused by PrP. Work from several labs suggests that these currents arise from a poorly regulated PrP N-terminal domain. This will be examined collaboratively using whole cell patch clamp electrophysiology to determine the influence of PrP structure, a-cleavage and Abeta binding. Together, these experiments will test a model in which PrP must be tightly regulated in metal ion homeostasis, and how loss of regulation might contribute to prion and Alzheimer's disease.