老鼠普立昂蛋白自發性結構轉變

普立昂疾病為一類致命且具傳染性的神經退化疾病，其致病因子為結構不正常且易於聚集的普立昂蛋白(prion protein)。此一具致病力的形式，或稱PrPSc，係由細胞中正常表現的PrPC改變其三維立體結構而來；而此一由α螺旋為主的結構轉變為β摺板為主的結構，會導致蛋白質本身之聚集，進而形成易沉澱的澱粉樣纖維構造。因此，欲釐清普立昂疾病之由來與其傳播方式，就必須視蛋白質不正常之摺疊機制為一重要的探討方向。而近年來有越來越多的證據顯示一種結構以β摺板為主卻可溶的蛋白寡聚體 (β-PrP)，可能是澱粉樣纖維形成過程中的前驅物，此一寡聚體便成為探討普立昂蛋白的不正常摺疊中十分重要的部分。然而，目前文獻中所探討的β-寡聚體多係由還原反應、酸化、或是具變性條件之中所產生，且此蛋白中的雙硫鍵對於結構穩定之貢獻亦仍無定論。在這項研究中，我們由全長老鼠之普立昂蛋白得到一種新穎之β寡聚體，且其結構轉變之過程可單純由移除雙硫鍵而觀察到。有趣的是，這些過程可在中性緩衝液中發生。此外電子自旋共振(EPR)圖譜暗示蛋白之helix 2可能參與其中。我們的發現指出雙硫鍵之還原在普立昂蛋白結構轉變中扮演重要地位。

關鍵字

普立昂疾病；錯誤摺疊； β寡聚體；雙硫鍵；結構轉變

並列摘要

Prion diseases are fatal transmissible neurodegenerative disorder. The infectious agent, “PRION”, is disease-associated β-rich aggregates (PrPSc) which is converted from the normal cellular conformer (PrPC) of the prion protein. Since the α->β conformational transition leads to protein aggregation and formation of toxic amyloid fibrils, the mechanism of protein misfolding is a key subject to elucidate the pathogenic pathway of prion diseases which is still unclear. Recently, several publications have shown that the soluble state of β-rich oligomers (β-PrP) can be a possible precursor of amyloid fibrils, therefore β-PrP is an important part on elucidating the mechanism of prion refolding. However, in previous reports β-PrP was generated under reduced, acidic, and denaturing conditions, and the role of disulfide bond involved in prion structure stabilization is still in doubt. In this study, we show that a novel β-PrP was generated from the full-length recombinant mouse prion protein, and its structural conversion can be observed simply by removing the disulfide bond. Interestingly, slow, spontaneous structural conversion from α- to β- structure can be monitored in a neutral buffer by CD spectroscopy. Moreover, EPR spectra suggest that unfolding of helix 2 is involved in the structural change. Our findings strongly indicate the important role of disulfide bond reduction on the structural conversion of prion protein.

並列關鍵字

prion diseases ； misfolding ； oligomer ； disulfide bond ； structural conversion

參考文獻

Aguzzi A, Heikenwalder M, Polymenidou M (2007) Insights into prion strains and neurotoxicity. Nat Rev Mol Cell Biol 8: 552-561

Aguzzi A, Polymenidou M (2004) Mammalian prion biology: one century of evolving concepts. Cell 116: 313-327

Baskakov IV (2007) The reconstitution of mammalian prion infectivity de novo. FEBS J 274: 576-587

Baskakov IV, Legname G, Baldwin MA, Prusiner SB, Cohen FE (2002) Pathway complexity of prion protein assembly into amyloid. J Biol Chem 277: 21140-21148

Bocharova OV, Breydo L, Parfenov AS, Salnikov VV, Baskakov IV (2005) In vitro conversion of full-length mammalian prion protein produces amyloid form with physical properties of PrP(Sc). J Mol Biol 346: 645-659

國際替代計量

老鼠普立昂蛋白自發性結構轉變

全文下載

主題瀏覽