In vitro cleavage at or near the N-terminus of the helper component protein in the tobacco vein mottling virus polyprotein.
In vitro cleavage at or near the N-terminus of the helper component protein in the tobacco vein mottling virus polyprotein.
复制标题
烟草静脉斑驳病毒多蛋白中辅助成分蛋白 N 末端或附近的体外裂解。
DOI：
10.1016/0042-6822(91)90543-k
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发表时间：
1991
期刊：
影响因子：
3.7
通讯作者：
Rhoads,RE
中科院分区：
文献类型：
--
作者：
Mavankal,G;Rhoads,RE
烟草静脉斑驳病毒 (TVMV) RNA 在小麦胚芽系统中的翻译产生了两种在兔网织红细胞系统中未观察到的产物。其中之一是 N 端蛋白，因为它是最丰富的产物，并且在 SDS-PAGE 上的迁移约为 34 kDa。基于 SDS-PAGE 上与 HC 的共迁移以及抗 HC 抗体的免疫沉淀，第二种产物与从 TVMV 感染植物中分离的辅助成分 (HC) 相似或相同。通过放射化学Edman降解确定该产物的N末端为多蛋白的Ser-257。该分配得到了色氨酸特异性试剂的肽图谱的支持。当烟草蚀刻病毒在小麦胚芽系统中翻译时，观察到类似的裂解。与其他五种马铃薯病毒组的同源区域的比较表明，所提议的切割位点两侧的氨基酸残基是保守的。通过 TVMV RNA 转录模板的定点诱变将 Phe-256 转化为 Met、Pro、Arg、His 或 Trp 可抑制小麦胚芽系统中的裂解。这些结果表明体外裂解发生在Phe-256和Ser-257之间，并且该裂解与释放HC N-末端的体内裂解相同。
Translation of tobacco vein mottling virus (TVMV) RNA in a wheat germ system resulted in two products that were not observed in a rabbit reticulocyte system. One of these was the N-terminal protein, based on its being the most abundant product and its migration on SDS-PAGE at about 34 kDa. The second product was similar or identical to helper component (HC) isolated from TVMV-infected plants, based on co-migration with HC on SDS-PAGE and immunoprecipitation with anti-HC antibodies. The N-terminus of this product was determined by radiochemical Edman degradation to be Ser-257 of the polyprotein. This assignment was supported by peptide mapping with a tryptophan-specific reagent. A similar cleavage was observed when tobacco etch virus was translated in a wheat germ system. Comparison with homologous regions in five other potyviruses indicated conservation of amino acid residues on both sides of the proposed cleavage site. Conversion of Phe-256 to Met, Pro, Arg, His, or Trp by site-directed mutagenesis of a TVMV RNA transcription template inhibited cleavage in the wheat germ system. These results suggest thatin vitrocleavage occurs between Phe-256 and Ser-257 and that this cleavage is the same as thein vivocleavage which liberates the N-terminus of HC.