Mertens P P, Burroughs J N, Anderson J
Virology. 1987 Apr;157(2):375-86. doi: 10.1016/0042-6822(87)90280-7.
Effective purification methods have been developed for virus particles, infectious subviral particles (ISVP), and virus cores of bluetongue virus (BTV) serotypes 1 and 4. The purified particles were analysed by indirect ELISA or PAGE using either silver staining, or fluorography of [35S]methionine-labelled preparations. No significant contamination with host cell proteins, or with the majority of BTV nonstructural proteins was detectable in any of the particle preparations. In addition to the two major outer capsid and five core proteins previously described, the purified virus particles of both serotypes were consistently found to contain small amounts of BTV protein NS2, previously regarded as exclusively nonstructural. This protein could be removed from the particle surface by treatment with a combination of chymotrypsin and sodium N-lauroyl sarcosinate, which also resulted in the cleavage of the larger of the two major outer capsid components (protein VP2). Two of the cleavage products of VP2 and the whole of the other major outer capsid component (protein VP5) formed a modified outer capsid layer in the resultant ISVP. These subviral particles were as or more infectious than the intact virus particles but had lost haemagglutinating activity. The core-associated RNA polymerase remained inactive in ISVP.
已开发出针对蓝舌病毒（BTV）血清型1和4的病毒颗粒、感染性子病毒颗粒（ISVP）及病毒核心的有效纯化方法。通过间接ELISA或PAGE对纯化颗粒进行分析，PAGE采用银染或对[35S]甲硫氨酸标记制剂进行荧光自显影。在任何颗粒制剂中均未检测到宿主细胞蛋白或大多数BTV非结构蛋白的明显污染。除了先前描述的两种主要外衣壳蛋白和五种核心蛋白外，两种血清型的纯化病毒颗粒均始终被发现含有少量BTV蛋白NS2，该蛋白先前被认为完全是非结构性的。用胰凝乳蛋白酶和N-月桂酰肌氨酸钠组合处理可从颗粒表面去除该蛋白，这也导致两种主要外衣壳成分中较大的一种（蛋白VP2）被切割。VP2的两种切割产物和另一种主要外衣壳成分（蛋白VP5）的全部在所得的ISVP中形成了修饰的外衣壳层。这些亚病毒颗粒与完整病毒颗粒具有相同或更高的感染性，但失去了血凝活性。核心相关的RNA聚合酶在ISVP中仍无活性。
