Aptamers targeted to an RNA hairpin show improved specificity compared to that of complementary oligonucleotides
Aptamers targeted to an RNA hairpin show improved specificity compared to that of complementary oligonucleotides
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DOI：
10.1021/bi0606344
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发表时间：
2006-10-03
期刊：
影响因子：
2.9
通讯作者：
Toulme, Jean-Jacques
中科院分区：
文献类型：
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作者：
Darfeuille, Fabien;Reigadas, Sandrine;Toulme, Jean-Jacques
核酸适配体通过环环（所谓的接吻）相互作用与RNA发夹相互作用，被描述为识别RNA发夹的反义寡聚物的替代选择。R06，一种RNA适体，先前被证明与HIV-1 RNA的TAR（反式激活反应性）发夹形成接吻复合物（Duconge和Toulme (1999) RNA 5,1605）。我们从R06中获得了一个嵌合锁定核酸(LNA)/DNA适体，它保留了最初选择的R06适体的结合特性。我们证明了这种rna /DNA适体与逆转录病毒蛋白Tat的肽竞争以结合TAR，即使这两个配体的结合位点彼此不重叠。这表明，在结合时，适体TAR采用不再适合Tat结合的构象。相反，与嵌合适体具有相同结合常数和相同碱基配对潜力的LNA/DNA反义寡聚体不会与Tat竞争。此外，我们发现LNA/DNA适体是比LNA/DNA反义序列更特异的TAR结合物。这些结果证明了读取RNA靶标的三维形状而不是其初级序列对于设计高度特异性寡核苷酸的好处。
Aptamers interacting with RNA hairpins through loop-loop ( so-called kissing) interactions have been described as an alternative to antisense oligomers for the recognition of RNA hairpins. R06, an RNA aptamer, was previously shown to form a kissing complex with the TAR (trans-activating responsive) hairpin of HIV-1 RNA (Duconge and Toulme (1999) RNA 5, 1605). We derived a chimeric locked nucleic acid (LNA)/DNA aptamer from R06 that retains the binding properties of the originally selected R06 aptamer. We demonstrated that this LNA/DNA aptamer competes with a peptide of the retroviral protein Tat for binding to TAR, even though the binding sites of the two ligands do not overlap each other. This suggests that upon binding, the aptamer TAR adopts a conformation that is no longer appropriate for Tat association. In contrast, a LNA/DNA antisense oligomer, which exhibits the same binding constant and displays the same base-pairing potential as the chimeric aptamer, does not compete with Tat. Moreover, we showed that the LNA/DNA aptamer is a more specific TAR binder than the LNA/DNA antisense sequence. These results demonstrate the benefit of reading the three-dimensional shape of an RNA target rather than its primary sequence for the design of highly specific oligonucleotides.