Journal: Chemical Science

Article Title: Potent inhibitors of the human RNA ligase Rlig1 highlights its role in RNA integrity maintenance under oxidative cellular stress †

doi: 10.1039/d4sc06542e

Figure Lengend Snippet: (A) General principle of the Rlig1 screen. A nicked RNA hairpin was labelled with FAM and BHQ1 at the 5′ and 3′ terminus, respectively. After ligation with Rlig1-AMP, EDTA is added to induce RNA unfolding, resulting in increased fluorescence intensity for the non-ligated substrate. In contrast, the fluorescence of the ligation product remains quenched, as the quencher and fluorophore are still in close proximity. RNA is illustrated in blue. (B) Schematic workflow of the high-throughput screening assay. Compound libraries were screened in a 384-well plate format to identify inhibitors of Rlig1-AMP. The data was evaluated using KNIME, followed by data confirmation and characterization. (C) Structure of SGI-1027 (1), which was identified as the most promising hit from the HTS, demonstrating over 70% inhibition at a 10 μM concentration. The indicated R 1 and R 2 residues were further investigated to understand their influence on the inhibitory effect on Rlig1-AMP activity.

Article Snippet: With this principle, we screened a library comprising 13 026 biologically active compounds in a 384-well plate format (HY-L001, MedChemExpress) for their inhibition of Rlig1-AMP.

Techniques: Ligation, Fluorescence, High Throughput Screening Assay, Screening Assay, Inhibition, Concentration Assay, Activity Assay