EXPRESSION OF GLYCOLIPID ANCHORED PRIONS IN TRYPANOSOMES

布氏锥虫引起动物和人类的疾病，通常称为非洲锥虫病，Nagana或睡眠病。 锥虫可以在感染的宿主中无限期地持续存在，通过抗原变异的过程逃避免疫系统，该过程由在锥虫表面上形成可降解的“涂层”的变体表面糖蛋白（VSG）的顺序表达介导。 每个锥虫表达1千万个单一的糖基磷脂酰肌醇（GPI）锚定的VSG分子，占总细胞蛋白的10%。T.布氏杆菌的数量与红细胞的数量大致相同，并且在大鼠或小鼠中感染高峰时，锥虫的产量约为红细胞数量的三分之一。 VSG的研究提供了明确的化学证据，随后，第一个完整的结构的GPI膜锚和阐明了许多细节的复杂途径GPI合成。 糖脂锚随后在广泛的真核细胞中的数百种蛋白质中发现，它们具有共同的核心结构。 T.布氏杆菌是内源性GPI锚定蛋白表面表达的“工厂”：VSG和前环酸性重复蛋白（PARP）分别占血液和昆虫（Glossina，the Tesetse）“前环”形式的总锥虫蛋白的10%。大多数哺乳动物GPI锚定蛋白占总细胞蛋白的不到0.1%。尽管积累的知识涉及朊病毒或突变型朊病毒基因在几种脑病，核心理论-朊病毒传播涉及的内源性细胞蛋白（PrPC）的自我催化转化为传染性羊瘙痒症朊病毒（PrPSc）-不能容易地进行测试，因为不能产生大量的感染性PrPSc或天然PrPC从天然来源或重组技术。关于PrPC结构的几个问题仍然存在，尽管最近发表的NMR研究，必须进行复性非糖基化非GPI锚定的重组蛋白。大量的GPI锚定的VSG，是由锥虫和方法的发展，使锥虫稳定转化表达异位基因，提供了可信的前景，可以快速评估，生产高兴趣的高价值GPI锚定的哺乳动物蛋白一般和PrP特别。 该提案将集中于在T中生产不同形式的PrP。布鲁塞。

Trypanosoma brucei causes a disease of animals and humans that is commonly called African Trypanosomiasis, Nagana or Sleeping Sickness. Trypanosomes can persist indefinitely in the infected host, evading the immune system through a process of antigenic variation, mediated by the sequential expression of variant surface glycoproteins (VSGs) that form an impregnable 'coat' on the trypanosome surface. Each trypanosome expresses 10 million molecules of a single glycosylphosphatidylinositol- (GPI) anchored VSG, representing 10 percent of the total cell protein. The surface area of T. brucei is about the same as that of an erythrocyte and the yield of trypanosomes, at the peak of infection in a rat or mouse, is about one third of the number of erythrocytes. Studies of VSGs provided the definitive chemical evidence and, subsequently, the first complete structure of a GPI membrane anchor and the elucidation of many details of a complex pathway for GPI synthesis. The glycolipid anchors subsequently found in hundreds of proteins in a wide range of eukaryotic cells share a common core structure. T. brucei is a 'factory' for the surface expression of endogenous GPI-anchored proteins: VSG and the procyclic acidic repetitive protein (PARP) each comprise 10 percent of the total trypanosome protein, in the bloodstream and insect (Glossina, the Tesetse) 'procyclic' forms, respectively. Most mammalian GPI-anchored proteins comprise less than 0.1 percent of the total cellular protein. Despite the accumulated knowledge implicating prions or mutant prion genes in several encephalopathies, the central theory - that prion propagation involves a self-catalysed conversion of the endogenous cellular protein (PrPC) to the infectious scrapie prion (PrPSc) - cannot readily be tested because of the inability to produce infective PrPSc or native PrPC in large amounts from natural sources or by recombinant techniques. Several questions about the structure of PrPC remain, despite recently published NMR studies that were necessarily performed on renatured non-glycosylated non-GPI-anchored recombinant protein. The vast amount of GPI-anchored VSG that is made by trypanosomes and the development of methods that allow trypanosomes to be stably transformed to express ectopic genes, offer credible prospects, which can be rapidly evaluated, for producing high-interest high-value GPI-anchored mammalian proteins in general and PrP in particular. This proposal will focus on producing different forms of PrP in T. brucei.