堤岸田鼠(Bank Vole)重组朊蛋白在RT-QuIC反应上的特点及其关键位点的作用探索

Reactivities of the Recombinant Prion Protein of Myodes Glareolus (Bank Vole) and Effects of Two Amino-acid Alternations in RT-QuIC

堤岸田鼠对很多种属的朊病毒易感。在堤岸田鼠的朊蛋白氨基酸序列中,只有两个位点是同仓鼠和小鼠都不相同的,分别是位于C端的227位和230位。为了评估这两个位点对堤岸田鼠RT-QuIC反应的影响,我们构建了田鼠野生型（Vole-WT）,两种突变型（Vole-E227D,Vole-S230R）,以及仓鼠野生型（Ha-WT）蛋白用于RT-QuIC反应。结果显示田鼠野生型及两种突变型具有极强的RT-QuIC反应效率。即使在较弱的反应条件下,田鼠野生型及突变型也表现出比仓鼠更高的反应效率。其中Vole-E227D在三种田鼠蛋白中是效率最高的,随后为野生型和Vole-S230R。我们的结果表明堤岸田鼠朊蛋白氨基酸227位和230位不是造成堤岸田鼠RT-QuIC反应如此高的原因,但特定氨基酸位点的改变会在一定程度上影响朊蛋白的RT-QuIC反应效率。

The mature PrP protein（aa23-231）of Myodes glareolus（bank vole）shows two amino-acid differences at the C-terminus of aa227 and aa230 compared with that of the mouse and hamster.To assess the potential influence of these two amino acids on the reactivity in RT-QuIC,two PrP mutants in Myodes glareolus,Vole-E227 Dand Vole-S230 R,were generated,in which the amino acids of PrP at aa227 and aa230 were replaced with those of the hamster and mouse.Together with the PrP of Myodes glareolus（Vole-WT）and PrP from a hamster（Ha-WT）,the reactivities of Vole-E227 Dand Vole-S230 R in RTQuIC using a strain（263 K）adapted from a hamster suffering from scrapie as the seed were tested.All three PrPs from Myodes glareolus showed much stronger reactivities in RT-QuIC than the PrP from a hamster,even under attenuated reaction conditions.Moreover,compared with Vole-WT,the RT-QuIC reactivity of Vole-E227 Dwas stronger,whereas that of Vole-S230 Rwas slightly weaker.Our results indicate that exchange of amino acids at positions aa227 and aa230 in the PrP of Myodes glareolus with hamster-and mouse-derived ones have little influence on their strong RT-QuIC reactivities.Nevertheless,special amino-acid changes at these two positions offer the constructs limited（but stable）RT-QuIC reactivity.