Solid state NMR of prion peptides

朊病毒肽的固态核磁共振

• 批准号: 6578746
• 负责人: DAVID E WEMMER
• 金额: $ 28.58万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6578746
• 关键词: bioimaging /biomedical imaging; chemical aggregate; chemical models; conformation; nuclear magnetic resonance spectroscopy; prions; protein structure function; stable isotope; structural biology

The focal point of the efforts for the next years will be to apply solid state NMR to peptides from the prion protein to biologically active aggregate states. We will begin with the "beta" form of the mouse 89-143 peptide containing the P101L mutation, which has been found to stimulate disease in transgenic mice. The inactive helical and coil states of the same peptide will be used for comparison to help identify conformational differences that may relate to the activity. We will also investigate the internal deletion peptide sPrP61, which is myrstylated at the C-terminal residue to mimic the GPI anchor. This peptide also forms ordered aggregates, but unlike the P101L peptide becomes protease resistant, analogous to PrPSc. We will try to generate magnetically ordered samples of fibrils of the peptides (analogous to what has been done for fiber diffraction), which would provide more structural information than available in unordered "powder" samples. We will also explore block- labeled peptides (full labeling for a contiguous set of amino acids) for local structure analysis. We will finish ongoing calibrations for using chemical shielding for determination of phi and psi angles and analyze the extent to which x1 affects analysis and/or can be independently determined. If methods for preparing PrPSc in vitro become available we will prepare specifically labeled samples to compare with the peptide models that have been studied.

未来几年的工作重点将是将固态核磁共振应用于从Pron蛋白到具有生物活性的聚集态的多肽。我们将从含有P101L突变的小鼠89-143肽的“beta”形式开始，这种突变已被发现在转基因小鼠中会刺激疾病。相同多肽的非活性螺旋和卷曲状态将用于比较，以帮助确定可能与活性有关的构象差异。我们还将研究内部删除肽sPrP61，它在C-末端残基上被桃金娘酰化，以模拟GPI锚点。这种多肽也形成有序的聚集体，但与P101L不同的是，P101L多肽与PrPSc类似，具有蛋白酶抗性。我们将尝试生成多肽原纤维的磁性有序样本(类似于纤维衍射所做的那样)，这将提供比无序“粉末”样本更多的结构信息。我们还将探索用于局部结构分析的块标记多肽(对一组连续的氨基酸进行完全标记)。我们将完成正在进行的使用化学屏蔽来确定phi和psi角的校准，并分析x1影响分析和/或可以独立确定的程度。如果在体外制备PrPSc的方法可行，我们将准备专门标记的样品，与已研究的多肽模型进行比较。