Aggregates of scrapie-associated prion protein induce the cell-free conversion of protease-sensitive prion protein to the protease-resistant state
Aggregates of scrapie-associated prion protein induce the cell-free conversion of protease-sensitive prion protein to the protease-resistant state
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DOI：
10.1016/1074-5521(95)90087-x
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发表时间：
1995-12-01
影响因子：
--
通讯作者：
Lansbury, PT
中科院分区：
文献类型：
--
作者：
Caughey, B;Kocisko, DA;Lansbury, PT
简介：羊瘙痒病感染通过一种未知的机制促使正常的蛋白酶敏感性朊病毒蛋白（PrPC）在体内转化为蛋白酶抗性形式（PrPSc）。体外研究表明，PrPSc可以诱导这种转化，这与PrPSc本身可能是传染性羊瘙痒病病原体的提议一致。使用这种无细胞模型的PrPC到PrPSc的转换，我们研究了反应物浓度的依赖性的转换，和性质的PrPSc衍生的物种，具有转换activity.Results：S-35 PrPC的蛋白酶K抗性形式的无细胞转换是依赖于反应时间和初始浓度的PrPSc（以上的表观最低阈值浓度）和S-35 PrPC。转化活性的物理大小的分析表明，可检测的转化活性仅与聚集体相关。在温和的离液条件下，部分解聚PrPSc和提高转换活性的活性物种是异质性的大小，但大于有效溶解的PrP或分子量：标准的类似2000 KDa.Conclusions：负责转换活性的实体是许多倍大于可溶性PrP单体，并需要一个阈值浓度的PrPSc。这些结果与PrPSc形成的成核聚合机制一致，并且与异源二聚体机制不一致。
Introduction: Scrapie infection instigates the in vivo conversion of normal, protease-sensitive prion protein (PrPC) into a protease-resistant form (PrPSc) by an unknown mechanism. In vitro studies have indicated that PrPSc can induce this conversion, consistent with proposals that PrPSc itself might be the infectious scrapie agent. Using this cell-free model of the PrPC to PrPSc conversion, we have studied the dependence of conversion on reactant concentration, and the properties of the PrPSc-derived species that has converting activity.Results: The cell-free conversion of S-35 PrPC to the proteinase K-resistant form was dependent on the reaction time and initial concentrations of PrPSc (above an apparent minimum threshold concentration) and S-35 PrPC. Analysis of the physical size of the converting activity indicated that detectable converting activity was associated only with aggregates. Under mildly chaotropic conditions, which partially disaggregated PrPSc and enhanced the converting activity the active species were heterogeneous in size, hut larger than either effectively solubilized PrP or molecular weight: standards of similar to 2000 kDa.Conclusions: The entity responsible for the converting activity was many times larger than a soluble PrP monomer and required a threshold concentration of PrPSc. These results are consistent with a nucleated polymerization mechanism of PrPSc formation and inconsistent with a heterodimer mechanism.