STRUCTURAL CHARACTERIZATION OF PRION PROTEINS

朊病毒蛋白的结构表征

• 批准号: 6308820
• 负责人: STANLEY B PRUSINER
• 金额: $ 0.99万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2002-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6308820
• 关键词: Mammalia; aging; animal tissue; biological products; biomedical resource; congenital disorders; infection; nervous system; proteins; spectrometry; structural biology

Introduction: Prion diseases are unique fatal neurodgenerative diseases caused by novel pathogens termed prions. The major or only component of prions is an abnormal form of a normal cellular protein termed the prion protein (PrP). The normal form (PrPC) is converted to the pathogenic form (PrPSc) by an as yet unidentified posttranslational process. Mass spectrometry is essential for the detailed analysis of small-scale samples of both forms of the prion protein isolated from the brains of hamsters. Methods: Several novel methods have been refined for the analysis of PrPSc using LSIMS, ESIMS, MALDI and MS/MS for the characterization of peptides and oligosaccharides from enzyme digests. These analyses are complicated by the hydrophobic, insoluble and strongly aggregating propoerties, particularly of PrPSc. PrPC analyses have been delayed by problems of purifying the protein from brain. Preliminary results by MALDI show that the two forms are similar, but microheterogeneity has prevented any definitive conclusions being drawn. Comparisons are also required between PrPSc from different prion strains. Strain behavior could be due to differences in the N-linked oligosaccharides; perhaps cell specific lectins cause targetting of certain PrP species to a limited range of cell types, resulting in regional distribution of PrPSc. We plan to use LC/MS for much of this characterization. Discussion: Results obtained so far suggest that the modification of PrPC involved in the formation of PrPSc may be conformational rather than chemical. It is essential to confirm that PrpCand PrPSc either are or are not identical in order to develop hypotheses for the molecular mechanisms of prion diseases.

简介：朊病毒病是一种独特的致命性神经退行性疾病 由称为朊病毒的新型病原体引起的疾病。 主要或唯一 朊病毒的成分是正常细胞蛋白质的异常形式 称为朊病毒蛋白（PrP）。 转换范式（PrPC） 致病性形式 (PrPSc) 是由一种尚未鉴定的 翻译后过程。 质谱分析对于 对两种形式的朊病毒的小规模样本进行详细分析 从仓鼠大脑中分离出的蛋白质。 方法：几种新颖的 使用 LSIMS 改进了 PrPSc 分析方法， ESIMS、MALDI 和 MS/MS 用于表征肽和 来自酶消化的寡糖。 这些分析很复杂 通过疏水性、不溶性和强聚集特性， 特别是 PrPSc。 PrPC 分析因以下问题而被推迟 从大脑中纯化蛋白质。 MALDI 初步结果显示 这两种形式是相似的，但微观异质性阻碍了 得出任何明确的结论。 还需要比较 来自不同朊病毒株的 PrPSc 之间的差异。 应变行为可能是 由于N-连接寡糖的差异；也许是细胞 特定的凝集素导致某些 PrP 物种的靶向性有限 细胞类型的范围，导致 PrPSc 的区域分布。 我们 计划使用 LC/MS 进行大部分表征。 讨论： 迄今为止获得的结果表明 PrPC 的修饰涉及 PrPSc 的形成过程可能是构象作用而非化学作用。 必须确认 PrpCand PrPSc 是或不是 相同以发展分子机制的假设 朊病毒疾病。