BIOCHEMICAL AND BIOPHYSICAL CHARACTERIZATION OF PRION PROTEIN 2D CRYSTALS

朊病毒蛋白二维晶体的生物化学和生物物理表征

• 批准号: 7957413
• 负责人: STANLEY B PRUSINER
• 金额: $ 0.01万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7957413
• 关键词: Benign; Biochemical; Classification; Computer Retrieval of Information on Scientific Projects Database; Cryoelectron Microscopy; Disease; Dose; Excision; Funding; Grant; Institution; Mass Spectrum Analysis; Molecular Conformation; Neurodegenerative Disorders; PrPSc Proteins; Preparation; Prions; Protein Isoforms; Proteins; Protocols documentation; Research; Research Personnel; Resources; Sampling; Scrapie; Source; Staining method; Stains; Structure; United States National Institutes of Health; base; particle; polyacrylamide gels; protein purification

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Prions cause transmissible, neurodegenerative diseases with invariably fatal progression. A prion is an infective particle consisting exclusively of the prion protein. Two forms are discriminated: the benign, cellular form PrPC, and the disease-causing "scrapie" isoform PrPSc, which is confirmationally rearranged. Since PrPC and PrPSc differ in structure, studies of each conformation are essential. Our apporach focuses on PrPSc. We developed a high-yield protein purification protocol. Our preparations contain prion protein 2D crystals, which are structurally analyzed with standard and low-dose cryo electron microscopy. However, to further the quality and purity of our samples, we need to identify contaminants/copurified proteins from our samples for their systematic removal. This is the task for mass spectrometry. On a collaborative basis, copurified proteins are to be identified by analysis of bands derived from Coomassie-stained polyacrylamide gels.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 prions会导致可传播的神经退行性疾病，其致命进展总是为致命的。 prion是一个仅由prion蛋白组成的感染粒子。两种形式有区别：良性，细胞形式PRPC和引起疾病的“ crapie”同工型PRPSC，这是确认重新排列的。由于PRPC和PRPSC的结构不同，因此对每个构象的研究都是必不可少的。我们的Apporach专注于PRPSC。我们制定了高收益蛋白纯化方案。我们的制剂含有prion蛋白2D晶体，并用标准和低剂量冷冻电子显微镜在结构上分析。 但是，为了进一步提高样品的质量和纯度，我们需要从样品中鉴定出污染物/共生的蛋白质，以进行系统的去除。这是质谱法的任务。 在协作的基础上，应通过分析源自Coomassie染色的聚丙烯酰胺凝胶的频带来识别共生蛋白。