Characterization of prion protein conformational changes

描述(由申请人提供)：传染性海绵状脑病(TSE)是由PrP构象变化引起的，PrP是一种膜相关的糖蛋白。TSE或Prion疾病，包括克雅氏病，人类致命的家族性失眠和库鲁，绵羊的瘙痒病，牛的牛海绵状脑病，以及鹿和麋鹿的慢性消瘦疾病。虽然罕见，但这些疾病总是致命的。鉴于它们也是可传播的，它们不一定只影响患病的患者或动物：人类形式对血液和器官接受者构成威胁，动物形式对我们的食物供应构成威胁。 Prion病的中心假设是，它是一种纯蛋白质疾病，因此Prion蛋白是繁殖和传播所必需的唯一病原体。在疾病过程中，Prion蛋白从其主要的螺旋细胞形式(PrPC)转变为具有丰富的β结构并保留大部分螺旋结构的构象(PrPSc)。PrPSc聚集性、致病和传染性。虽然大多数研究只关注蛋白质，但PrP的N-连接糖链影响PrP在大脑中的表达、分布和沉积，这反过来又决定了患者的症状。不幸的是，事实证明，不可能从实验中获得转化过程和相应的疾病相关构象的高分辨率结构信息。因此，我们的假设是，全原子分子动力学模拟将提供可测试的模型(与NIH RML的拜伦·考伊博士合作进行测试)，以了解转化过程、物种障碍、感染性低聚体以及糖基化和脂膜对转化过程的影响。具体来说，我们会： ？对不同物种(人、牛、仓鼠、绵羊、小鼠、鸡、乌龟、猪和麋鹿)在中性和低pH(低pH值在体内和体外都会引发PrPSc构象变化)的水中的PrPC进行多分子动力学(MD)模拟。 ？通过包括多糖、GPI锚和中性和低pH条件下的膜环境，对更多生物相关的蛋白质结构进行MD模拟，以评估它们对转化的影响 ？构建具有感染性的、有毒的PrPSc原纤维低聚物(不是原纤维)的分子模型，并对这些结构以及异源低聚物(例如牛PrPSc人PrPC)进行MD模拟，以研究物种屏障。

DESCRIPTION (provided by applicant): Transmissible spongiform encephalopathies (TSEs) are caused by a conformational change in the prion protein (PrP), which is a membrane-associated, glycoprotein. TSEs, or prion diseases, include Creutzfeldt-Jakob disease, fatal familial insomnia and Kuru in humans, scrapie in sheep, bovine spongiform encephalopathy in cattle, and chronic wasting disease in deer and elk. While rare, these diseases are always fatal. Given that they are also transmissible, they don't necessarily just affect the afflicted patient or animal: the human forms represent a threat to blood and organ recipients and the animal forms are a threat to our food supply. The central hypothesis in prion disease is that it is a protein-only disease, whereby the prion protein is the only agent necessary for propagation and transmission. In the disease process, the prion protein converts from its primarily helical, cellular form (PrPC) to a conformer rich in beta-structure with retention of most of the helical structure (PrPSc). PrPSc aggregates, causes disease, and is infectious. While most studies focus on just the protein, PrP's N-linked glycans influence PrP expression, distribution, and deposition in the brain, which in turn determines patient symptoms. Unfortunately it has proved impossible to obtain high-resolution structural information for the conversion process and corresponding disease-associated conformers from experiment. Consequently, our hypothesis is that all-atom molecular dynamics simulations will provide testable models (to be tested in collaboration with Dr. Byron Caughey, NIH RML) for the conversion process, species barriers, the infectious oligomers, and the effect of glycosylation and the lipid membrane on the process. Specifically, we will: ?Perform multiple molecular dynamics (MD) simulations of the cellular prion protein (PrPC) from different species (human, bovine, hamster, ovine, mouse, chicken, turtle, pig and elk) in water at neutral and low pH (low pH triggers the conformational change to PrPSc both in vivo and in vitro) ?Perform MD simulations of more biologically relevant protein constructs by including glycans, the GPI anchor, and the membrane environment at neutral and low pH to assess their effect on conversion ?Construct molecular models for the infectious, toxic protofibrillar oligomers (not fibrils) of PrPSc and perform MD simulations of these constructs as well as hetero-oligomers (for example bovine PrPSc + human PrPC) to investigate species barriers.