Basic studies on the biosynthesis of pathogenic prion protein

致病性朊病毒蛋白生物合成的基础研究

• 批准号: 12460130
• 负责人: MAKINO Sou-ichi
• 金额: $ 9.09万
• 依托单位: Obihiro University of Agriculture and Veterinary Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12460130/
• 关键词: prion; scrapie; PrP; conformational transformation; glycosaminoglycan; synthetic peptide; BSE; 伝達性海綿状脳症; 種特異性

An abnormal isoform of prion protein (PrPSc) is a major component of the infectious agent "prion" and the biosynthesis of PrPSc play a central role in pathogenesis of prion diseases. Once prion enters the host, PrPSc binds to normal prion protein (PrPC) and converts PrPC into PrPSc. During the conformational transformation, PrPSc acts as a template or seed. In this study, we analyzed inter-molecular interaction between PrPC and PrPSc that is a key event in PrPSc generation.We found that PrPC bound to heterologous PrPSc but did not convert to PrPSc and that the conversion reaction could be interfered by heterologous PrPC. These finding indicate that the conversion of PrPC to PrPSc is controlled more by the conformational transformation step. Analysis using mutant PrPC revealed that three amino acids locating central part of PrP is major determinant of the conversion reaction. We also examined the micro-environment where the conversion reaction takes place and found that sulfated glycans … More , a constituent of glycosaminoglycan, facilitated the PrPSc generation. Glycosaminoglycan is a major component of extracellular matrix so that environment in the outer surface of cells may be involved in the interaction between PrPC and PrPSc. Furthermore we found PrP synthetic peptide, aa119-141, 167-179 and 200-223 inhibited the PrPSc formation. These peptides formed high beta-sheet aggregates and bound to PrPC, implying the PrPC binding domain on the PrPSc molecule. In this study we have succeeded to produce an extensive panel of monoclonal antibodies against PrP molecule. The mAbs in this panel could be divided into at least 11 groups based on their epitope. Seven of them recognized linear epitopes on PrPC and/or denatured PrP molecule, while 3 of them recognized discontinuous epitopes on PrP molecule. The remaining one group of mAb, mAb 6H10 could differentiate PrPSc from PrPC. This extensive panel of mAb will be invaluable for further analyses of interaction between PrPSc and PrPC. Less

Prion蛋白（PRPSC）的异常同工型是传染药“ Prion”的主要组成部分，PRPSC的生物合成在prion病的发病机理中起着核心作用。 Prion进入宿主后，PRPSC与正常的Prion蛋白（PRPC）结合，并将PRPC转换为PRPSC。在构象转化期间，PRPSC充当模板或种子。在这项研究中，我们分析了PRPC和PRPSC之间的分子间相互作用，这是PRPSC生成中的一个关键事件。我们发现PRPC与异源性PRPSC结合，但未转化为PRPSC，并且转化反应可以由异源PRPC干扰。这些发现指标表明，PRPC转换为PRPSC的转化是通过构象转换步骤控制的。使用突变体PRPC的分析表明，定位PRP中心部分的三个氨基酸是转化反应的主要决定剂。我们还检查了发生转化反应发生的微环境，发现硫化的聚糖……更多，构成糖胺聚糖的一个支持PRPSC的一代。糖胺聚糖是细胞外基质的主要组成部分，因此细胞外表面的环境可能与PRPC和PRPSC之间的相互作用有关。此外，我们发现PRP合成肽AA119-141、167-179和200-223抑制了PRPSC的形成。这些胡椒体形成了高β-片骨料并与PRPC结合，这意味着PRPSC分子上的PRPC结合域。在这项研究中，我们成功地生产了针对PRP分子的大量单克隆抗体。该面板中的mab可以根据其epoisodeTope将至少11组分为11组。他们中的七个识别PRPC和/或变性的PRP分子上的线性表位，而其中3个识别PRP分子上的不连续表位。其余一组MAB MAB 6H10可以将PRPSC与PRPC区分开。对于PRPSC和PRPC之间的进一步分析，这一广泛的MAB将是无价的。较少的

项目成果

Horiuchi,M., et al.,: "Inhibition of interactions and interconversions of prion protein isoforms by peptide fragments from the C-terminal folded domain"Journal of Biological Chemistry. (in press). (2001)

Horiuchi，M.，等人：“C 端折叠结构域的肽片段对朊病毒蛋白亚型的相互作用和相互转化的抑制”生物化学杂志。

Takekida, K.et al.: "Quantitative analysis of prion protein by immunoblottiing"J.Health Science. 48. 288-291 (2002)

Takekida, K.等人：“通过免疫印迹对朊病毒蛋白进行定量分析”J.健康科学。

Horiuchi, M., et al.: "Inhibition of interactions and interconversions of prion protein isoforms by peptide fragments from the C-terminal folded domain"J. Biol. Chem.. 276. 15489-15497 (2001)

Horiuchi, M., 等人：“C 端折叠结构域的肽片段对朊病毒蛋白亚型的相互作用和相互转化的抑制”J.

Horiuchi, M.et al.: "Interactions between heterologous forms of prion protein : binding, inhibition of conversion, and species barrier"Proc.Natl Acad.Sci.USA. 97. 5836-5841 (2000)

Horiuchi, M.等人：“异源形式朊病毒蛋白之间的相互作用：结合、转化抑制和物种屏障”Proc.Natl Acad.Sci.USA。

Ikeda, T., et al.: "Outline of inspection system for bovine spongiform encephalopathy in Japan and future correspondence (in Japanese)"Food Sanitation Research. 52. 33-42 (2001)

Ikeda, T., et al.：“日本牛海绵状脑病检查系统概要和未来对应（日语）”食品卫生研究。