STRUCTURAL STUDIES ON PRION PROTEINS

朊病毒蛋白的结构研究

• 批准号: 6267176
• 负责人: STANLEY B PRUSINER
• 金额: $ 22.24万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-02-01 至 1998-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6267176
• 关键词: conformation; posttranslational modifications; prions; protein folding; protein purification; protein reconstitution; protein structure; radiotracer; scrapie; spectrometry; synthetic peptide; virulence; zinc

A wealth of experimental data argues persuasively that infectious prion particles are composed largely, if not entirely, of the scrapie isoform of the prior protein (PrPSc). Additional investigations have shown that PrPSc is formed from the cellular prion protein (PrPc) by a posttranslational process, which occurs after PrPC reaches the cell surface where it is anchored by a glycosyl phosphatidyl-inositol (GPI) moiety. No candidate chemical modifications have been identified that might distinguish PrPSc from PrPC, to date. These experimental results argue that it is likely that the conversion from PrPC into PrPSc involves a conformational change in the protein. In the studies proposed here, we plan to refine the purification of PrPc and establish whether the primary structure of PrPC including the posttranslational modifications is identical to that already determined for PrPSc. The secondary, tertiary and quaternary structures of PrPC and PrPSc as well as synthetic PrP peptides will be determined using spectroscopic methods. These studies should help elucidate the conformational changes involved in the various "strains" of scrapie are due to detailed differences in the posttranslational modifications of PrPSc including the arrays of heterogeneous asparagine-linked oligosaccharides and the GPI anchor. We propose to investigate the possibility of generating scrapie infectivity in vitro through refolding of denatured PrSc as well as modifying the folding of PrPc, recombinant PrP90-228 and synthetic PrP peptides. We hope to establish the size and composition of the smallest infectious prion particle using Zn2+ or diethylpyrocarbonate (DEPC) to perturb reversibly the structure of PrPSc. The results with Zn2+ ions and DEPC as well as those obtained with synthetic PrP peptides implicate His111 in the synthesis of PrPSc and the propagation of scrapie prion infectivity. Radiolabeled DEPC will be used to test the hypothesis that scrapie infectivity is diminished when His111 is carbethoxylated. Studies of the molecular mechanisms responsible for prion diseases should give new insights into the etiology, pathogenesis and treatment of the more prevalent neurodegenerative disorders such as Alzheimer's disease.

大量的实验数据令人信服地证明传染性朊病毒