Characterization of prion protein conformational changes

描述（由申请方提供）：传染性海绵状脑病（TSE）由朊病毒蛋白（PrP）的构象变化引起，朊病毒蛋白是一种膜相关糖蛋白。TSE，或朊病毒疾病，包括克雅氏病，人类的致命性家族性失眠症和库鲁病，绵羊的瘙痒病，牛的牛海绵状脑病，以及鹿和麋鹿的慢性消耗病。虽然罕见，但这些疾病总是致命的。考虑到它们也是可传播的，它们不一定只影响受折磨的病人或动物：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。 朊病毒病的核心假设是它是一种仅蛋白质的疾病，朊病毒蛋白是繁殖和传播所必需的唯一媒介。在疾病过程中，朊病毒蛋白从其主要的螺旋细胞形式（PrPC）转化为富含β结构的构象异构体，保留大部分螺旋结构（PrPSc）。PrPSc聚集，引起疾病，并且具有传染性。虽然大多数研究只关注蛋白质，但PrP的N-连接聚糖影响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，它已被证明是不可能获得高分辨率的结构信息的转换过程和相应的疾病相关的构象从实验。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Byron Caughey博士，NIH RML合作测试）的转换过程，物种障碍，感染性低聚物，糖基化和脂质膜的影响的过程。具体而言，我们将： ?在中性和低pH（低pH触发体内和体外PrPSc的构象变化）条件下，对来自不同物种（人、牛、仓鼠、绵羊、小鼠、鸡、龟、猪和麋鹿）的细胞朊病毒蛋白（PrPC）进行多重分子动力学（MD）模拟 ?通过在中性和低pH下加入聚糖、GPI锚和膜环境，对生物学相关性更高的蛋白质构建体进行MD模拟，以评估其对转化的影响 ?构建PrPSc的感染性、毒性原纤维寡聚体（非原纤维）的分子模型，并对这些构建体以及异源寡聚体（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.