Two-Dimensional Combinatorial Screening Identifies Specific 6′-Acylated Kanamycin A- and 6′-Acylated Neamine-RNA Hairpin Interactions
Two-Dimensional Combinatorial Screening Identifies Specific 6′-Acylated Kanamycin A- and 6′-Acylated Neamine-RNA Hairpin Interactions
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DOI：
10.1021/bi8012615
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发表时间：
2008-12-02
期刊：
影响因子：
2.9
通讯作者：
Disney, Matthew D.
中科院分区：
文献类型：
--
作者：
Aminova, Olga;Paul, Dustin J.;Disney, Matthew D.
本文中，我们报告了通过二维组合筛选（2DCS）鉴定的结合6 '-酰化卡那霉素A（1）和6'-酰化新霉胺（2）的六核苷酸发夹文库中的RNA发夹环。选择结合1的发夹具有235至1035 nM的K-d，平均K-d为618 nM。对于2，所选发夹结合的Kd范围为135至2300 nM，平均Kd为1010 nM。所选择的RNA发夹-配体相互作用也对它们选择结合的配体具有特异性，与其他阵列配体相比。例如，选择为1的发夹的混合物与1的结合平均比与2的结合紧密33倍，而选择为2的发夹的混合物与2的结合平均比与1的结合紧密11倍。完成所选序列的二级结构预测以确定每个配体结合的基序，并计算1和2的发夹环偏好。对于1，优选的发夹环含有由至少两个核苷酸与胞嘧啶分开的腺嘌呤，例如，ANNCNN（双尾p值= 0.0010）和ANNNCN（双尾p值< 0.0001）。对于2，优选的发夹环含有5 'GC和5' CG步骤（双尾p值< 0.0001）。这些结果扩展了有关结合小分子的RNA发夹环的可用信息，并可能被证明对靶向RNA有用。
Herein, we report the RNA hairpin loops from a six-nucleotide hairpin library that bind 6'-acylated kanamycin A (1) and 6'-acylated neamine (2) identified by two-dimensional combinatorial screening (2DCS). Hairpins selected to bind 1 have K-d's ranging from 235 to 1035 nM, with an average K-d of 618 nM. For 2, the selected hairpins bind with K-d's ranging from 135 to 2300 nM, with an average K-d of 1010 nM. The selected RNA hairpin-ligand interactions are also specific for the ligand that they were selected to bind compared with the other arrayed ligand. For example, the mixture of hairpins selected for 1 on average bind 33-fold more tightly to 1 than to 2, while the mixtures of hairpins selected for 2 on average bind 11-fold more tightly to 2 than to 1. Secondary structure prediction of the selected sequences was completed to determine the motifs that each ligand binds, and the hairpin loop preferences for 1 and 2 were computed. For 1, the preferred hairpin loops contain an adenine separated by at least two nucleotides from a cytosine, for example, ANNCNN (two-tailed p-value = 0.0010) and ANNNCN (two-tailed p-value < 0.0001). For 2, the preferred hairpin loops contain both 5'GC and 5'CG steps (two-tailed p-value < 0.0001). These results expand the information available on the RNA hairpin loops that bind small molecules and could prove useful for targeting RNA.