Protein-protein- and protein-RNA-binding properties of the movement protein and VP25 coat protein of Apple latent spherical virus.
Protein-protein- and protein-RNA-binding properties of the movement protein and VP25 coat protein of Apple latent spherical virus.
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DOI：
10.1016/j.virol.2006.02.025
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发表时间：
2006-08
期刊：
影响因子：
3.7
通讯作者：
M. Isogai;Kentaro Watanabe;Yusuke Uchidate;N. Yoshikawa
中科院分区：
文献类型：
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作者：
M. Isogai;Kentaro Watanabe;Yusuke Uchidate;N. Yoshikawa
为了阐明苹果潜伏球形病毒（ALSV）运动的机制，分析了其细胞间运动蛋白（MP）的各种特性。 ELISA 和印迹重叠测定表明 MP 与 ALSV 病毒颗粒特异性结合，特别是与三种外壳蛋白之一 (VP25) 结合，但不与其他两种外壳蛋白（VP20 和 VP24）结合。突变分析表明 MP 包含两个具有独立 VP25 结合活性的结构域（氨基酸残基 1-188 和 189-281）。此外，核苷酸结合实验表明，MP和VP25与单链RNA（ssRNA）和ssDNA结合，没有任何序列特异性，但这两种蛋白不与双链RNA（dsRNA）和dsDNA结合。 MP 在氨基酸残基 95-188、189-281 和 277-376 之间包含三个潜在独立的单链核酸结合域。在凝胶延迟分析中，MP 表现出与 ssRNA 的合作性结合，VP25 表现出非合作性结合。当MP和VP25在竞争性结合实验中一起测试时，MP的协同RNA结合变得非合作性，即使提供了足够量的MP用于完全协同RNA结合MP。讨论了 MP 和 VP25 相互作用以及核酸结合活性在 ALSV 运动中的作用。
To elucidate the mechanism of Apple latent spherical virus (ALSV) movement, various properties of its cell-to-cell movement protein (MP) were analyzed. ELISA and blot overlay assays demonstrated that the MP bound specifically to ALSV virions and in particular to one of the three coat proteins (VP25) but not to the other two coat proteins (VP20 and VP24). Mutational analyses have revealed that the MP contains two domains with independent VP25-binding activity (amino acid residues 1–188 and 189–281). Furthermore, nucleotide-binding experiments showed that the MP and VP25 bound to single-stranded RNA (ssRNA) and ssDNA without any sequence specificity, but these two proteins did not bind to double-stranded RNA (dsRNA) and dsDNA. The MP contains three potentially independent single-stranded nucleic acid-binding domains between amino acid residues 95–188, 189–281 and 277–376. The MP demonstrated cooperative and VP25 demonstrated non-cooperative binding to ssRNA in gel-retardation analyses. The cooperative RNA binding of the MP became non-cooperative when MP and VP25 were tested together in competition binding experiments, even though a sufficient amount of the MP for fully cooperative RNA binding the MP was supplied. The roles of the MP and VP25 interactions and nucleic acid binding activities in ALSV movement are discussed.