Prion protein insertional mutations increase aggregation propensity but not fiber stability.
Prion protein insertional mutations increase aggregation propensity but not fiber stability.
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DOI：
10.1186/1471-2091-9-7
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发表时间：
2008-03-17
期刊：
影响因子：
--
通讯作者：
True HL
中科院分区：
文献类型：
--
作者：
Kalastavadi T;True HL
PrNP基因突变约占所有Prion病病例的15%。关于PRNP中的一些突变如何导致蛋白质聚集成淀粉样纤维或导致疾病的机制，人们知之甚少。我们利用嵌合蛋白系统研究了PrP蛋白的寡肽重复结构域(ORD)的扩展及其对蛋白质聚集和淀粉样纤维形成的影响。我们用PrP野生型和扩增的Ord替换了酵母PrP蛋白Sup35p的Ord，并比较了它们的体外生化性质。我们先前已经确定，这些嵌合蛋白在体内保持了[PSI+]酵母蛋白的表型。有趣的是，我们注意到重复扩展的嵌合普里子似乎能够保持更强的[Psi+]菌株，并以更高的频率从[Psi-]转换为[Psi+]。在这项研究中，我们试图了解这些嵌合蛋白的生化性质，并建立一个系统来研究PrP的ORD在体内和体外的性质。对嵌合蛋白的体外研究表明，重复扩展增加了聚集倾向，纤维形成的动力学取决于重复的数量。纤维形成反应是混杂的，因为含有14个重复的嵌合蛋白可以很容易地与具有野生型重复数目的蛋白质的种子纤维形成。从形态上看，由重复扩大的蛋白质形成的淀粉样纤维相互关联，形成大团块，这在由含有野生型重复数的蛋白质形成的纤维中并不常见。尽管重复扩展蛋白的聚集倾向和侧向结合增加，但形成的纤维的稳定性并没有相应增加。因此，我们预测，不同重复长度的纤维形成的差异可能不是由于淀粉样蛋白核心的巨大变化造成的。这里提供的生化观察解释了以前在酵母中观察到的这些嵌合蛋白的性质。更重要的是，他们提出了一种机制，用于解释在人类中观察到的发病年龄和疾病严重性与ORD长度之间的相关性。
Mutations in the PRNP gene account for ~15% of all prion disease cases. Little is understood about the mechanism of how some of these mutations in PRNP cause the protein to aggregate into amyloid fibers or cause disease. We have taken advantage of a chimeric protein system to study the oligopeptide repeat domain (ORD) expansions of the prion protein, PrP, and their effect on protein aggregation and amyloid fiber formation. We replaced the ORD of the yeast prion protein Sup35p with that from wild type and expanded ORDs of PrP and compared their biochemical properties in vitro. We previously determined that these chimeric proteins maintain the [PSI+] yeast prion phenotype in vivo. Interestingly, we noted that the repeat expanded chimeric prions seemed to be able to maintain a stronger strain of [PSI+] and convert from [psi-] to [PSI+] with a much higher frequency. In this study we have attempted to understand the biochemical properties of these chimeric proteins and to establish a system to study the properties of the ORD of PrP both in vivo and in vitro. Investigation of the chimeric proteins in vitro reveals that repeat-expansions increase aggregation propensity and that the kinetics of fiber formation depends on the number of repeats. The fiber formation reactions are promiscuous in that the chimeric protein containing 14 repeats can readily cross-seed fiber formation of proteins that have the wild type number of repeats. Morphologically, the amyloid fibers formed by repeat-expanded proteins associate with each other to form large clumps that were not as prevalent in fibers formed by proteins containing the wild type number of repeats. Despite the increased aggregation propensity and lateral association of the repeat expanded proteins, there was no corresponding increase in the stability of the fibers formed. Therefore, we predict that the differences in fibers formed with different repeat lengths may not be due to gross changes in the amyloid core. The biochemical observations presented here explain the properties of these chimeric proteins previously observed in yeast. More importantly, they suggest a mechanism for the observed correlation between age of onset and disease severity with respect to the length of the ORD in humans.
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