A small molecule microarray platform to select RNA internal loop-ligand interactions
A small molecule microarray platform to select RNA internal loop-ligand interactions
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DOI：
10.1021/cb700174r
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发表时间：
2007-11-01
影响因子：
4
通讯作者：
Disney, Matthew D.
中科院分区：
文献类型：
--
作者：
Childs-Disney, Jessica L.;Wu, Meilan;Disney, Matthew D.
在此，我们报告了一个微阵列平台的发展，以选择RNA基序配体的相互作用，允许同时筛选RNA和化学空间。我们使用该平台来鉴定结合6 '-N-5-己炔酸卡那霉素A的RNA内环（1）。我们详细研究了结合1的选定内环，通常显示一个腺嘌呤与一个胞嘧啶交叉，与环的大小无关。还观察到其他偏好。对于3 × 3核苷酸环，优先选择嘌呤，对于2 × 2核苷酸环，优先选择由腺嘌呤与胞嘧啶交叉连接的嘧啶。该技术具有用于选择RNA基序-配体相互作用的几个有利特征：（1）通过从较低配体加载量收获结合的RNA来鉴定较高亲和力的RNA基序-配体相互作用;（2）通过凝胶提取从阵列收获结合的RNA，减轻选择中的动力学偏差;以及（3）在单个阵列表面上完成多重选择。为了进一步证明多重选择可以在同一阵列表面上平行完成，我们从结合四元氨基糖苷文库的4096元RNA内环文库中选择RNA内环。这些实验在单个实验中探测了16，384（4个氨基糖苷X 4096-成员RNA文库）相互作用。这些研究允许化学和RNA空间的平行筛选，以提高我们对RNA-配体相互作用的理解。这些信息可能有助于合理和模块化设计的小分子靶向RNA。
Herein, we report the development of a microarray platform to select RNA motif-ligand interactions that allows simultaneous screening of both RNA and chemical space. We used this platform to identify the RNA internal loops that bind 6'-N-5-hexynoate kanamycin A (1). Selected internal loops that bind 1 were studied in detail and commonly display an adenine across from a cytosine independent of the size of the loop. Additional preferences are also observed. For 3 X 3 nucleotide loops, there is a preference for purines, and for 2 X 2 nucleotide loops there is a preference for pyrimidines neighbored by an adenine across from a cytosine. This technique has several advantageous features for selecting RNA motif-ligand interactions: (1) higher affinity RNA motif-ligand interactions are identified by harvesting bound RNAs from lower ligand loadings; (2) bound RNAs are harvested from the array via get extraction, mitigating kinetic biases in selections; and (3) multiple selections are completed on a single array surface. To further demonstrate that multiple selections can be completed in parallel on the same array surface, we selected the RNA internal loops from a 4096-member RNA internal loop library that bound a four-member aminoglycoside library. These experiments probed 16,384 (4 aminoglycoside X 4096-member RNA library) interactions in a single experiment. These studies allow for parallel screening of both chemical and RNA space to improve our understanding of RNA-ligand interactions. This information may facilitate the rational and modular design of small molecules targeting RNA.