Binding of herpes simplex virus-1 US11 to specific RNA sequences.
Binding of herpes simplex virus-1 US11 to specific RNA sequences.
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DOI：
10.1093/nar/gki919
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发表时间：
2005
影响因子：
14.9
通讯作者：
Coen DM
中科院分区：
文献类型：
--
作者：
Bryant KF;Cox JC;Wang H;Hogle JM;Ellington AD;Coen DM
单纯疱疹病毒1型US 11是一种具有新的RNA结合结构域的RNA结合蛋白。据报道，US 11表现出序列和构象特异性RNA结合，但对结合重要的序列和构象尚不清楚。US 11也被描述为双链RNA（dsRNA）结合蛋白。为了研究US 11-RNA相互作用，我们从由>1014个80个碱基序列组成的RNA文库中进行了结合US 11的RNA适体的体外选择，所述序列在30个碱基的随机化区域中不同。US 11以70 nM的亲和力特异性结合选定的适体。对23个选择的序列的分析揭示了强一致序列。US 11 RNA结合结构域和含有共有序列的所选RNA的≤46个碱基均足以结合。US 11结合保护共有基序免受羟基自由基裂解。所选适体的RNA酶双链断裂揭示了单链RNA和dsRNA的区域。我们观察到US 11以序列非特异性方式结合两种不同的dsRNA，但亲和力低于其结合所选适体的亲和力。结果定义了以高亲和力结合US 11的相对短的特异性序列，并表明单独的dsRNA不赋予高亲和力结合。
Herpes simplex virus-1 US11 is a RNA-binding protein with a novel RNA-binding domain. US11 has been reported to exhibit sequence- and conformation-specific RNA-binding, but the sequences and conformations important for binding are not known. US11 has also been described as a double-stranded RNA (dsRNA)-binding protein. To investigate the US11–RNA interaction, we performed in vitro selection of RNA aptamers that bind US11 from a RNA library consisting of >1014 80 base sequences which differ in a 30 base randomized region. US11 bound specifically to selected aptamers with an affinity of 70 nM. Analysis of 23 selected sequences revealed a strong consensus sequence. The US11 RNA-binding domain and ≤46 bases of selected RNA containing the consensus sequence were each sufficient for binding. US11 binding protected the consensus motif from hydroxyl radical cleavage. RNase digestions of a selected aptamer revealed regions of both single-stranded RNA and dsRNA. We observed that US11 bound two different dsRNAs in a sequence non-specific manner, but with lower affinity than it bound selected aptamers. The results define a relatively short specific sequence that binds US11 with high affinity and indicate that dsRNA alone does not confer high-affinity binding.
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