Assembly of natural and recombinant prion protein into fibrils
Assembly of natural and recombinant prion protein into fibrils
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DOI：
10.1515/bc.2005.067
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发表时间：
2005-06-01
影响因子：
3.7
通讯作者：
Riesner, D
中科院分区：
文献类型：
--
作者：
Leffers, KW;Wille, H;Riesner, D
朊病毒蛋白（PrPC）的a-螺旋细胞异构体（PrPC）转化为不溶性的、富含p-sheet的、传染性的致病异构体（PrPSc）是朊病毒疾病的基本事件。PrPSc的c端片段（prp27 -30）通过有限的蛋白水解形成，并保持感染性。与全长PrPSc不同，PrP 27-30聚合成具有淀粉样蛋白超微结构和着色特性的棒状结构。为了研究PrP的折叠，包括PrPC形成PrPSc和PrP 27-30组装棒，我们在之前用于研究该蛋白结构转变的条件下，将叙利亚仓鼠（sol SHa） PrP 27-30溶解在低浓度（0.2%）十二烷基硫酸钠（SDS）中。在SIDS浓度为0.02% ~ 0.04%时，Sol SHaPrP 27-30呈现富β -sheet结构，保持可溶性。在这里，我们报道了NaCl将SHaPrP 27-30稳定在一个可溶的，富含β -薄片的状态，允许原纤维组装进行数周。在这些条件下，不仅溶胶PrP 27-30可以形成原纤维，而且天然的SHaPrP(C)也可以形成原纤维。鞘脂的添加似乎能促进纤维的生长。当重组（rec） SHaPrP（90-231）暴露在低浓度的SIDS中，类似于在250 mm NaCl存在下聚合溶胶SHaPrP 27-30时，纤维形成有规律。在过表达全长SHaPrP的转基因小鼠中对PrP 27-30或PrPC形成的原纤维进行生物检测，未发现原纤维具有感染性，而rec小鼠PrP（89-230）形成的annyloid原纤维具有感染性。目前，还不能确定缺乏传染性是由原纤维结构的差异还是生物测定条件的差异引起的。
The conversion of the a-helical, cellular isoform of the prion protein (PrPC) to the insoluble, p-sheet-rich, infectious, disease-causing isoform (PrPSc) is the fundamental event in the prion diseases. The C-terminal fragment of PrPSc (PrP 27-30) is formed by limited proteolysis and retains infectivity. Unlike full-length PrPSc, PrP 27-30 polymerizes into rod-shaped structures with the ultrastructural and tinctorial properties of amyloid. To study the folding of PrP, both with respect to the formation of PrPSc from PrPC and the assembly of rods from PrP 27-30, we solubilized Syrian hamster (sol SHa) PrP 27-30 in low concentrations (0.2%) of sodium dodecyl sulfate (SDS) under conditions previously used to study the structural transitions of this protein. Sol SHaPrP 27-30 adopted a beta-sheet-rich structure at SIDS concentrations between 0.02% and 0.04% and remained soluble. Here we report that NaCl stabilizes SHaPrP 27-30 in a soluble, beta-sheet-rich state that allows fibril assembly to proceed over several weeks. Under these conditions, fibril formation occurred not only with sol PrP 27-30, but also with native SHaPrP(C). Addition of sphingolipids seems to increase fibril growth. When recombinant (rec) SHaPrP(90-231) was exposed to low concentrations of SIDS, similar to those used to polymerize sol SHaPrP 27-30 in the presence of 250 mm NaCl, fibril formation occurred regularly. When fibrils formed from PrP 27-30 or PrPC were bioassayed in transgenic mice overexpressing full-length SHaPrP, no infectivity was obtained, whereas annyloid fibrils formed of rec mouse PrP(89-230) were infectious. At present, it cannot be determined whether the lack of infectivity is caused by a difference in the structure of the fibrils or in the bioassay conditions.