Structural similarities of molecules selectively binding the prfA thermosensor RNA

Abstract

In light of the "silent" AMR pandemic, new avenues to combat pathogenic bacteria are needed. In this work, we screened a large molecule library (n=35 684 unique compounds) with the aim of identifying molecules being able to bind and block translation of the prfA-thermosensor transcript in the bacterial pathogen Listeria monocytogenes. Using a thiazole-orange displacement approach, 468 ([~]1.3% of all molecules) showed the ability to reduce fluorescence. After dose response testing, 32 compounds remained promising and eight of them showed sufficient purity and availability to be further validated. Interestingly, four compounds, being structurally very similar, showed specificity for prfA at a varying degree. All four compounds carried 3 aromatic rings with one connecting amine between two of the rings and an amide linking an aliphatic amine side chain. The most selective compounds, M5, showed a Kd of [~]0.8 {micro}M for the prfA RNA at 35{degrees}C. However, none of the eight most efficient compounds were able to inhibit prfA translation in vitro, suggesting that the molecules are able to bind but not affect the stability of the overall structure. Through this work, we have been able to identify a set of molecules, able to bind the prfA thermosensor RNA selectively, but without affecting translation. These molecules could constitute an important scaffold for further drug development.