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A Small Molecule Microarray Platform To Select RNA Internal Loop−Ligand Interactions

Supporting Info

Jessica L. Childs-Disney, Meilan Wu, Alexei Pushechnikov, Olga Aminova and Matthew D. Disney*

Department of Chemistry and Center of Excellence in Bioinformatics and Life Sciences, University at Buffalo, 657 Natural Sciences Complex, Buffalo, New York 14260

ACS Chem. Biol., 2007, 2 (11), pp 745–754

DOI: 10.1021/cb700174r

Publication Date (Web): November 2, 2007

* Corresponding author, mddisney@buffalo.edu.

Abstract

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 × 3 nucleotide loops, there is a preference for purines, and for 2 × 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 gel 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 × 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.

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