Functional analysis of thiamine pyrophosphate-responsive riboswitches in human bacterial pathogens of the ESKAPE group using a dual-luciferase reporter gene assay

Riboswitches are RNA molecules that control important processes in bacteria. Infections with pathogens of the ESKAPE group are common, and we are trying to find new ways to fight these bacteria. Small molecules can be designed to bind to riboswitches and optimally block their activity. In the present work, we have analyzed the thiamine pyrophosphate (TPP) riboswitches of ESKAPE pathogens with respect to small molecule binding. For this purpose, we developed a dual-luciferase reporter gene assay. Most of the predicted TPP riboswitches were indeed functional regulators and are thus targets for new anti-infectives. The small molecule pyrithiamine does not block all TPP riboswitches tested, and we found a structural basis for this behavior.