Edenhofer F, Rieger R, Famulok M, Wendler W, Weiss S, Winnacker E L
Laboratorium Für Molekulare Biologie-Genzentrum-Institute Für Biochemie der Ludwig-Maximilians-Universität Munchen, Germany.
J Virol. 1996 Jul;70(7):4724-8. doi: 10.1128/JVI.70.7.4724-4728.1996.
Prions mediate the pathogenesis of certain neurodegenerative diseases, including bovine spongiform encephalopathy in cattle and Creutzfeldt-Jakob disease in humans. The prion particle consists mainly, if not entirely, of PrPSc, a posttranslationally modified isoform of the cellular host-encoded prion protein (PrPc). It has been suggested that additional cellular factors might be involved in the physiological function of PrPc and in the propagation of PrPSc. Here we employ a Saccharomyces cerevisiae two-hybrid screen to search for proteins which interact specifically with the Syrian golden hamster prion protein. Screening of a HeLa cDNA library identified heat shock protein 60 (Hsp60), a cellular chaperone as a major interactor for PrPc. The specificity of the interaction was confirmed in vitro for the recombinant proteins PrPc23-231 and rPrP27-30 fused to glutathione S-transferase with recombinant human Hsp60 as well as the bacterial GroEL. The interaction site for recombinant Hsp60 and GroEL proteins was mapped between amino acids 180 and 210 of the prion protein by screening with a set of recombinant PrPc fragments. The binding of Hsp60 and GroEL occurs within a region which contains parts of the putative alpha-helical domains H3 and H4 of the prion protein.
朊病毒介导某些神经退行性疾病的发病机制，包括牛的牛海绵状脑病和人类的克雅氏病。朊病毒颗粒主要（如果不是完全）由PrPSc组成，PrPSc是细胞宿主编码的朊病毒蛋白（PrPc）的翻译后修饰异构体。有人提出，其他细胞因子可能参与PrPc的生理功能和PrPSc的传播。在这里，我们利用酿酒酵母双杂交筛选来寻找与叙利亚金仓鼠朊病毒蛋白特异性相互作用的蛋白质。对HeLa cDNA文库的筛选确定热休克蛋白60（Hsp60），一种细胞伴侣蛋白，是PrPc的主要相互作用蛋白。用重组人Hsp60以及细菌GroEL对与谷胱甘肽S-转移酶融合的重组蛋白PrPc23-231和rPrP27-30进行体外实验，证实了这种相互作用的特异性。通过用一组重组PrPc片段进行筛选，确定了重组Hsp60和GroEL蛋白与朊病毒蛋白氨基酸180至210之间的相互作用位点。Hsp60和GroEL的结合发生在一个包含朊病毒蛋白假定的α-螺旋结构域H3和H4部分的区域内。
