Thiamine Pyrophosphate Riboswitches Are Targets for the Antimicrobial Compound Pyrithiamine

Abstract

Thiamine metabolism genes are regulated in numerous bacteria by a riboswitch class that binds the coenzyme thiamine pyrophosphate (TPP). We demonstrate that the antimicrobial action of the thiamine analog pyrithiamine (PT) is mediated by interaction with TPP riboswitches in bacteria and fungi. For example, pyrithiamine pyrophosphate (PTPP) binds the TPP riboswitch controlling the tenA operon in Bacillus subtilis. Expression of a TPP riboswitch-regulated reporter gene is reduced in transgenic B. subtilis or Escherichia coli when grown in the presence of thiamine or PT, while mutant riboswitches in these organisms are unresponsive to these ligands. Bacteria selected for PT resistance bear specific mutations that disrupt ligand binding to TPP riboswitches and derepress certain TPP metabolic genes. Our findings demonstrate that riboswitches can serve as antimicrobial drug targets and expand our understanding of thiamine metabolism in bacteria.

Structural features of metabolite-sensing riboswitches

“…The pyrimidine ring of TPP is recognized by one helix whereas binding to the pyrophosphate moiety occurs within an interhelical region between coaxially stacked helices located parallel to the pyrimidine-binding helix. Surprisingly, the thiazole moiety of TPP is not specifically recognized by RNA through hydrogen bonding or base stacking interactions, accounting for the prior observation that TPP analogs containing alterations in this region were still capable of associating with the RNA [16,42,50]. Similar to the SAM sensor, the TPP riboswitch class also faces the electrostatic challenge of accommodating the negative charge of the pyrophosphate linkage.…”

Ribozymes, riboswitches and beyond: regulation of gene expression without proteins

“…Recent studies have identified several metabolite-like antimicrobial compounds that function by targeting riboswitches. Among them are the TPP analogue pyrithiamine pyrophosphate (PTPP), which differs from TPP by the central ring 59 (FIG. 4b), two analogues of lysine, L-aminoethylcysteine (AEC) and DL-4-oxalysine, which contain carbon substitutions in the lysine side chain 60 (FIG.…”

Structural analysis of a class III preQ 1 riboswitch reveals an aptamer distant from a ribosome-binding site regulated by fast dynamics

“…Although present in all domains of life, they are prominent in bacteria where they typically reside in the 5′-leader sequences of mRNA (2). Broad interest in riboswitches originates from the discovery that they can be targeted by antimicrobials (3)(4)(5), and the observation that they use complex scaffolds to achieve gene regulation without the need for protein partners. In the latter respect, riboswitches typically exhibit bipartite sequence organization comprising a conserved aptamer linked to a downstream expression platform (2).…”