Evidence that the RNA methylation and poly(A) polymerase stimulatory activities of vaccinia virus protein VP39 do not impinge upon one another.
Evidence that the RNA methylation and poly(A) polymerase stimulatory activities of vaccinia virus protein VP39 do not impinge upon one another.
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有证据表明痘苗病毒蛋白 VP39 的 RNA 甲基化和聚腺苷酸聚合酶刺激活性不会相互影响。
DOI：
10.1006/viro.1998.9209
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Hodel,AE
中科院分区：
文献类型：
--
作者：
Gershon,PD;Shi,X;Hodel,AE
痘苗病毒蛋白VP 39具有两种RNA修饰活性。在单体形式中，它作为mRNA帽特异性2′-O-甲基转移酶，特异性修饰m7 G加帽mRNA的第一个转录核苷酸的核糖部分。与VP 55（牛痘多聚腺苷酸聚合酶的催化亚基）结合，VP 39促进长度已经大于1035 nt的多聚腺苷酸尾的快速延伸。引入新的检测方法，我们提供的证据表明，VP 39的两种活性中的每一种的底物都不会可检测地调节匡威反应，并且VP 39的2′-O-甲基转移酶活性不会受到其与VP 55的关联的显著影响。在电泳迁移率变动分析中，VP 39与短（5个核苷酸）RNA相互作用，只有当后者是m7 G-加帽。具有较长（22个核苷酸）RNA的复合物更稳定（即，不依赖于帽），但通过m7 G帽的存在进一步稳定。在RNA：蛋白质摩尔比升高时观察到额外的复合物，表明每个VP 39分子存在两个RNA结合位点。在这些网站之一的相互作用是稳定的帽结构。另外的实验表明，通过VP 55-VP 39异二聚体经历聚（A）尾延伸的RNA分子不适合作为帽甲基化底物。
Vaccinia protein VP39 has two RNA modifying activities. In monomeric form, it acts as an mRNA cap-specific 2′-O-methyltransferase, specifically modifying the ribose moiety of the first transcribed nucleotide of m7G-capped mRNA. In association with VP55, the catalytic subunit of the vaccinia poly(A) polymerase, VP39 facilitates the rapid elongation of poly(A) tails that are already greater than ∼35 nt in length. Introducing new assays, we provide evidence that substrates for each of VP39's two activities do not detectably modulate the converse reaction and that VP39's 2′-O-methyltransferase activity is not significantly affected by its association with VP55. In an electrophoretic mobility shift assay, VP39 interacted with a short (5 nucleotide) RNA only when the latter was m7G-capped. Complexes with longer (22 nucleotide) RNAs were more stable (i.e., cap-independent) but were further stabilized by the presence of an m7G cap. An additional complex was observed at elevated RNA:protein molar ratios, indicating the presence of two RNA binding sites per VP39 molecule. Interaction at one of these sites was stabilized by the cap structure. Additional experiments indicated that RNA molecules undergoing poly(A) tail elongation by the VP55-VP39 heterodimer are not favored as cap-methylation substrates.