De Dios Krisel, Kumar Sachin, Alvandi Ehsan, Adhikari Utpal Kumar, David Monique Amtoinette, Tayebi Mourad
School of Medicine, Western Sydney University, Sydney, NSW 2560, Australia.
Brain Sci. 2025 Feb 26;15(3):250. doi: 10.3390/brainsci15030250.
: The normal cellular prion protein (PrP) is a cell-surface glycoprotein, mainly localised in neurons of the central nervous system (CNS). The human gene encodes 253 amino acid residues of precursor PrP. Several studies that investigated the role of and PrP in placental mammals, such as humans and mice, failed to reveal its exact function. : In this study, we sequenced and characterised the gene and PrP of the marsupial, , as a strategy to gain molecular insights into its structure and physicochemical properties. Placentals are separated from marsupials by approximately 125 million years of independent evolution. : Standard Western blotting analysis of PrP phascogale displayed the typical un-, mono-, and di-glycosylated bands recognized in placentals. Furthermore, we showed that phascogale gene has two exons, similar to all the marsupials and placentals of the genes studied. Of note, the phascogale gene contained distinctive repeats in the PrP tail region comparable to the closely related Tasmanian devil () and more distantly related to the grey short-tailed opossum (), common wombat (), and Tammar wallaby (); however, its specific composition and numbers were different from placentals. Of importance, comparisons of the phascogale's PrP physicochemical properties with other monotremes, marsupials, and placentals confirmed the Monotremata-Marsupialia-Placentalia evolutionary distance. We found that the protein instability index, a method used to predict the stability of a protein in vivo (Stable: <40; Instable >40), showed that the PrP of all marsupials tested, including phascogale, were highly stable compared with the birds, reptiles, amphibians, and fish that were shown to be highly unstable. However, the instability index predicted that all placental species, including human (), mouse (), bank vole (), rhinoceros (), dog (), flying fox (), whale (), cattle (), and sheep (), were either slightly unstable or nearly unstable. Further, our analysis revealed that despite their predicted high PrP stability, exhibited substantial N-terminal disorder (53.76%), while species with highly unstable PrPs based on their instability index, such as Danio rerio, Oryzias latipes, and Astyanax mexicanus, displayed even higher levels of N-terminal disorder (up to 75.84%). These findings highlight a discrepancy between overall predicted stability and N-terminal disorder, suggesting a potential compensatory role of disorder in modulating prion protein stability and function. : These results suggest that the high stability of marsupial prion proteins indicates a vital role in maintaining protein homeostasis; however more work is warranted to further depict the exact function.
正常细胞朊病毒蛋白（PrP）是一种细胞表面糖蛋白，主要定位于中枢神经系统（CNS）的神经元中。人类基因编码前体PrP的253个氨基酸残基。多项研究调查了PrP在胎盘哺乳动物（如人类和小鼠）中的作用，但未能揭示其确切功能。：在本研究中，我们对有袋动物袋鼬的基因和PrP进行了测序和表征，以此作为深入了解其结构和物理化学性质的分子策略。胎盘哺乳动物与有袋动物经历了约1.25亿年的独立进化。：对袋鼬PrP的标准蛋白质印迹分析显示出在胎盘哺乳动物中识别出的典型的未糖基化、单糖基化和双糖基化条带。此外，我们发现袋鼬基因有两个外显子，这与所研究的所有有袋动物和胎盘哺乳动物的基因相似。值得注意的是，袋鼬基因在PrP尾部区域包含独特的重复序列，与亲缘关系较近的袋獾以及亲缘关系较远的灰短尾负鼠、毛鼻袋熊和塔马尔沙袋鼠类似；然而，其具体组成和数量与胎盘哺乳动物不同。重要的是，可以通过比较袋鼬PrP的物理化学性质与其他单孔目动物、有袋动物和胎盘哺乳动物来证实单孔目动物 - 有袋动物 - 胎盘哺乳动物的进化距离。我们发现，用于预测蛋白质在体内稳定性的蛋白质不稳定指数（稳定：<40；不稳定：>40）表明，与显示出高度不稳定的鸟类、爬行动物、两栖动物和鱼类相比，包括袋鼬在内的所有测试有袋动物的PrP都高度稳定。然而，不稳定指数预测所有胎盘物种，包括人类、小鼠、社鼠、犀牛、狗、狐蝠、鲸鱼、牛和绵羊，要么略微不稳定，要么几乎不稳定。此外，我们的分析表明，尽管预测袋鼬的PrP具有高稳定性，但其N端无序程度较高（53.76%），而基于不稳定指数PrP高度不稳定的物种，如斑马鱼、青鳉和墨西哥丽脂鲤，其N端无序程度更高（高达75.84%）。这些发现突出了总体预测稳定性与N端无序之间的差异，表明无序在调节朊病毒蛋白稳定性和功能方面可能具有补偿作用。：这些结果表明，有袋动物朊病毒蛋白的高稳定性表明其在维持蛋白质稳态中起着至关重要的作用；然而，需要更多的工作来进一步描述其确切功能。
