High-throughput competitive binding assay for targeting RNA tertiary structures with small molecules: application to pseudoknots and G-quadruplexes.

ABSTRACT: There is an indisputable need for new screening methodologies to identify small molecules that target RNA tertiary structures, such as pseudoknots or G-quadruplexes. Here, we present a high-throughput competitive binding antisense assay designed to identify ligands for complex RNA tertiary structures. In this assay, initially customized for the bacterial PreQ1-I riboswitch pseudoknot, ligands compete with a rationally designed quencher-labelled antisense oligonucleotide for binding to a fluorophore-labelled riboswitch. The method is validated for four PreQ1-I riboswitches, using the natural riboswitch ligand PreQ1 and various analogues. A commercial RNA-focused library consisting of ∼15 000 compounds was then screened against the Fusobacterium nucleatum riboswitch, leading to the identific

SUBMITTER: Wintermans SEL 

PROVIDER: S-EPMC12397912 | biostudies-literature | 2025 Aug

REPOSITORIES: biostudies-literature