RNA is an extremely important target for the development of chemical probes of function or small molecule therapeutics. Aminoglycosides are the most well studied class of small molecules to target RNA. However, the RNA motifs outside of the bacterial rRNA A-site that are likely to be bound by these compounds in biological systems is largely unknown. If such information were known, it could allow for aminoglycosides to be exploited to target other RNAs and, in addition, could provide invaluable insights into potential bystander targets of these clinically used drugs. We utilized two-dimensional combinatorial screening (2DCS), a library-versus-library screening approach, to select the motifs displayed in a 3×3 nucleotide internal loop library and in a 6-nucleotide hairpin library that bind with high affinity and selectivity to six aminoglycoside derivatives. The selected RNA motifs were then analyzed using structure-activity relationships through sequencing (StARTS), a statistical approach that defines the privileged RNA motif space that binds a small molecule. StARTS allowed for the facile annotation of the selected RNA motif-aminoglycoside interactions in terms of affinity and selectivity. The interactions selected by 2DCS generally have nanomolar affinities, which is higher affinity than the binding of aminoglycosides to a mimic of their therapeutic target, the bacterial rRNA A-site.

译文

RNA是开发功能化学探针或小分子疗法的极其重要的靶标。氨基糖苷类是研究最充分的一类靶向RNA的小分子。然而,在生物系统中可能被这些化合物结合的细菌rRNA A位点之外的RNA基序在很大程度上是未知的。如果已知此类信息,则可以利用氨基糖苷类药物靶向其他rna,此外,还可以为这些临床使用的药物的潜在旁观者靶标提供宝贵的见解。我们利用二维组合筛选 (2DCS) (一种库对库筛选方法) 来选择在3 × 3核苷酸内环文库和6核苷酸发夹文库中显示的基序,这些基序以高亲和力和选择性结合六个氨基糖苷衍生物。然后通过测序 (ststarts) 使用结构-活性关系分析所选的RNA基序,这是一种统计方法,定义了结合小分子的特权RNA基序空间。开始允许在亲和力和选择性方面轻松注释所选RNA基序-氨基糖苷类相互作用。由2dc选择的相互作用通常具有纳摩尔亲和力,这比氨基糖苷类与其治疗靶标细菌rRNA a位点的模拟物结合的亲和力更高。

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