- 學位論文
研究普立昂感染屏障與胺基酸序列的關係
Exploring the relationship of prion transmission barrier and amino acid sequence
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摘要
傳播性海綿樣腦病是一種致死性的神經退化疾病會影響許多哺乳類動物，造成此病的原因是由非傳統性的感染病原普立昂（prion）所引起。Prion的傳播在同物種間比較容易發生，然而在不同物種之間則受限於「種族屏障」，也就是當外來的普立昂胺基酸序列若是與宿主的胺基酸序列相似度差異大的話就會影響對prion的形成並且不容易跨種傳染。為研究普立昂感染屏障與胺基酸序列的關係，我們截取普遍認為普立昂蛋白最重要的一段胺基酸序列108-144為模型，合成數條普立昂胜肽比較其類澱粉形成的特性，此外更利用晶種滴定方法量化出不同物種間普立昂疾病的感染屏障並檢測不同之胺基酸序列如何影響感染效率。 實驗結果顯示，PrP第112號殘基若為甲硫胺酸，會促進類澱粉纖維形成時的成核作用，若第139號殘基是甲硫胺酸、第143號殘基為絲胺酸則會延遲成核作用，但若同一條胜肽存在上述條件時，類澱粉纖維形成的效果會被中和。此外，若是以已經形成類澱粉纖維的牛（bPrP）為晶種進行引晶實驗會發現，第129號殘基若是白胺酸、第135號殘基為天門冬胺酸、第139號殘基是甲硫胺酸、第143號殘基為絲胺酸皆會使引晶屏障變高，但若138號殘基為甲硫胺酸則會降低引晶屏障，若是增加引晶屏障與降低引晶屏障兩者影響因子同時出現在一條普立昂胜肽上，則兩者的影響會相互抵銷。整體而言，bPrP對於catPrP、mPrP與bvPrP屬於低度引晶屏障，對於huPrP、haPrP與lionPrP則屬於中度引晶屏障，而對於mdPrP、dogPrP與pigPrP屬於高度引晶屏障。 若是以已經形成類澱粉纖維的北美大角鹿（mdPrP）為晶種進行引晶實驗，對bvPrP來說屬於低度種族屏障，而對於catPrP、lionPrP與mPrP則屬於中度引晶屏障，而對於bPrP、pigPrP、dogPrP、haPrP以及huPrP則屬於高度引晶屏障。
並列摘要
Transmissible spongiform encephalopathies (TSE) are a group of fatal neurodegenerative diseases of mammals, caused by unconventional infectious agents known as “prion”. Transmission of prion between species is limited by a so called “species barrier”. Prion formation could be affected dramatically if the amino acid sequences between the host PrPC (cellular prion protein) and the inoculated prion are different, which results in the resistance of cross-species TSE transmission. In order to explore the correlation between the prion transmission barrier and the amino acid sequence, several prion peptides (sequences 108-144) were synthesized（the sequence 108-144 based on the numbering of human prion protein sequence） and the amyloidogenic properties of these peptides were compared. We used seeding titration method to quantify the seeding barrier and to examine how sequence difference affected the seeding efficiency. Our results showed that while residue 112 with methionine accelerated nucleation, residues 139 with methionine or 143 with serine retarded the nucleation process. But these effects could be neutralized when the substitutions with opposite effects had shown on the same peptide. Moreover, using the amyloid fibrils prepared from bPrP as seed, residues 129 with leucine, 135 with asparagine, 139 with methionine or 143 with serine substitution could increase seeding barrier, but residue 138 with methionine decreased seeding barrier. When both decreased and increased factors were appeared in the same peptide, their effects tended to cancel each other out. Overall, the relative seeding barrier is low for catPrP, mPrP, and bvPrP; moderate for huPrP, haPrP, and lionPrP; high for mdPrP, dogPrP, and pigPrP. When the amyloid fibrils prepared from mdPrP were used as seed, the relative seeding barriers for all these peptides were different from the bPrP seeding results: the relative seeding barrier is low for bvPrP; moderate for catPrP, lionPrP, and mPrP; high for bPrP, pigPrP, dogPrP, haPrP, and huPrP.
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