MASS AND MOLECULAR COMPOSITION OF VESICULAR STOMATITIS-VIRUS - A SCANNING-TRANSMISSION ELECTRON-MICROSCOPY ANALYSIS
MASS AND MOLECULAR COMPOSITION OF VESICULAR STOMATITIS-VIRUS - A SCANNING-TRANSMISSION ELECTRON-MICROSCOPY ANALYSIS
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DOI：
10.1128/jvi.54.2.598-607.1985
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发表时间：
1985-01-01
影响因子：
5.4
通讯作者：
STEVEN, AC
中科院分区：
文献类型：
--
作者：
THOMAS, D;NEWCOMB, WW;STEVEN, AC
暗场扫描透射电镜用于对纯化的水泡性口炎病毒粒子、链霉蛋白酶处理的病毒粒子和核衣壳进行质量分析，从而对水泡性口炎病毒的分子组成进行了完全自洽的描述。获得的质量为265.6±。天然病毒颗粒为 13.3 兆道尔顿 (MDa)，197.5 .+-。链霉蛋白酶处理的病毒颗粒的 MDa 为 8.4，MDa 为 69.4.+-。 4.9 核衣壳的 MDa。链霉蛋白酶处理导致的质量减少，对应于 G 蛋白外部结构域（尖峰）的切除，导致平均 1205 个 G 蛋白/病毒粒子分子。在补偿 RNA (3.7 MDa) 和残留量的其他蛋白质后，核衣壳质量产生了 1258 个 N 蛋白拷贝的补体。通过生化定量校准M、NS和L蛋白相对于N蛋白的量，每个病毒粒子分别产生1826、466和50个分子的值。假设剩余的病毒粒子质量由病毒包膜中的脂质贡献，则获得其脂质含量的值为 56.1 MDa。应用四种不同的 EM 程序来确定核衣壳长度，得出的结论为 3.5-3.7 μm。核衣壳包含重复单元链，沿着链中部测量，其中心间距为3.3nm。这些重复单元代表N蛋白的单体，每个单体与9.+-相关。 1个碱基的单链RNA。从负染色核衣壳的扫描透射电镜图像推断，N 蛋白具有楔形双叶结构，尺寸为 .apprx。 9.0 nm（长度），约5.0 nm（深度）和.apprx。 3.3 nm（宽度，在其长轴的中点处）。在原位核衣壳的卷曲构型中，N蛋白的长轴呈放射状，其深度对应于核衣壳螺旋的螺距。
Dark-field scanning transmission EM was used to perform mass analyses of purified vesicular stomatitis virions, pronase-treated virions and nucleocapsids, leading to a complete self-consistent account of the molecular composition of vesicular stomatitis virus. The masses obtained were 265.6 .+-. 13.3 megadaltons (MDa) for the native virion, 197.5 .+-. 8.4 MDa for the pronase-treated virion and 69.4 .+-. 4.9 MDa for the nucleocapsid. The reduction in mass effected by pronase treatment, which corresponds to excision of the external domains (spikes) of G protein, leads to an average of 1205 molecules of G protein/virion. The nucleocapsid mass, after compensation for the RNA (3.7 MDa) and residual amounts of other proteins, yielded a complement of 1258 copies of N protein. Calibration of the amounts of M, NS and L proteins relative to N protein by biochemical quantitation yielded values of 1826, 466 and 50 molecules, respectively, per virion. Assuming that the remaining virion mass is contributed by lipids in the viral envelope, a value of 56.1 MDa for its lipid content was obtained. Four different EM procedures were applied to determine the nucleocapsid length, which was concluded to be 3.5-3.7 .mu.m. The nucleocapsid comprises a strand of repeating units which have a center-to-center spacing of 3.3 nm as measured along the middle of the strand. These repeating units represent monomers of N protein, each of which is associated with 9 .+-. 1 bases of single-stranded RNA. From scanning transmission EM images of negatively stained nucleocapsids, it is inferred that N protein has a wedge-shaped, bilobed structure with dimensions of .apprx. 9.0 nm (length), .apprx. 5.0 nm (depth) and .apprx. 3.3 nm (width, at the midpoint of its long axis). In the coiled configuration of the in situ nucleocapsid, the long axis of N protein is directed radially, and its depth corresponds to the pitch of the nucleocapsid helix.