A Structural and Functional Comparison Between Infectious and Non-Infectious Autocatalytic Recombinant PrP Conformers
A Structural and Functional Comparison Between Infectious and Non-Infectious Autocatalytic Recombinant PrP Conformers
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DOI：
10.1371/journal.ppat.1005017
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发表时间：
2015-06-01
期刊：
影响因子：
6.7
通讯作者：
Supattapone, Surachai
中科院分区：
文献类型：
--
作者：
Noble, Geoffrey P.;Wang, Daphne W.;Supattapone, Surachai
传染性朊病毒含有一种自我繁殖、错误折叠的朊病毒蛋白构象异构体，称为 PrPSc。仅蛋白质假说的一个关键预测是自催化 PrPSc 分子应该具有传染性。然而，一些自催化重组PrPSc分子在生物测定中具有低水平或不可检测的特异性感染性，并且重组朊病毒感染性的基本决定因素仍然不清楚。为了确定与感染性具体相关的结构和功能特征，我们比较了源自同一原始模板的两种自催化重组 PrP 构象异构体的特性，它们对野生型小鼠的特异性感染性相差 >10(5) 倍。在结构上，氢/氘交换质谱 (DXMS) 研究表明，除了包含残基 91-115 和 144-163 的两个结构域外，感染性和非感染性自催化重组 PrP 构象异构体的溶剂可及性特征在其整个蛋白酶抗性核心中非常相似。拉曼光谱和免疫沉淀研究证实，这些结构域在感染性与非感染性自催化重组 PrP 构象异构体中采用不同的构象。从功能上讲，体外朊病毒繁殖实验表明，非感染性构象异构体无法接种含有糖基磷脂酰肌醇 (GPI) 锚的小鼠 PrPC 底物，包括天然 PrPC。总而言之，这些结果表明，具有可被翻译后修饰的 PrPC 分子特异性采用的构象是重组朊病毒生物感染性的重要决定因素，并表明这种能力与 PrPSc 结构的离散特征相关。
Infectious prions contain a self-propagating, misfolded conformer of the prion protein termed PrPSc. A critical prediction of the protein-only hypothesis is that autocatalytic PrPSc molecules should be infectious. However, some autocatalytic recombinant PrPSc molecules have low or undetectable levels of specific infectivity in bioassays, and the essential determinants of recombinant prion infectivity remain obscure. To identify structural and functional features specifically associated with infectivity, we compared the properties of two autocatalytic recombinant PrP conformers derived from the same original template, which differ by >10(5)-fold in specific infectivity for wild-type mice. Structurally, hydrogen/deuterium exchange mass spectrometry (DXMS) studies revealed that solvent accessibility profiles of infectious and non-infectious autocatalytic recombinant PrP conformers are remarkably similar throughout their protease-resistant cores, except for two domains encompassing residues 91-115 and 144-163. Raman spectroscopy and immunoprecipitation studies confirm that these domains adopt distinct conformations within infectious versus non-infectious autocatalytic recombinant PrP conformers. Functionally, in vitro prion propagation experiments show that the non-infectious conformer is unable to seed mouse PrPC substrates containing a glycosylphosphatidylinositol (GPI) anchor, including native PrPC. Taken together, these results indicate that having a conformation that can be specifically adopted by post-translationally modified PrPC molecules is an essential determinant of biological infectivity for recombinant prions, and suggest that this ability is associated with discrete features of PrPSc structure.