Fragment Screening Hit Draws Attention to a Novel Transient Pocket Adjacent to the Recognition Site of the tRNA-Modifying Enzyme TGT

Fragment-based
lead discovery was applied to tRNA-guanine transglycosylase,
an enzyme modifying post-transcriptionally tRNAs in Shigella, the causative agent of shigellosis. TGT
inhibition prevents translation of Shigella’s virulence factor VirF, hence reducing pathogenicity. One
discovered fragment opens a transient subpocket in the preQ1-recognition site by pushing back an aspartate residue. This step
is associated with reorganization of further amino acids structurally
transforming a loop adjacent to the recognition site by duplicating
the volume of the preQ1-recognition pocket. We synthesized
6-carboxamido-, 6-hydrazido-, and 4-guanidino-benzimidazoles to target
the opened pocket, including a dihydro-imidazoquinazoline with a propyn-1-yl
exit vector pointing into the transient pocket and displacing a conserved
water network. MD simulations and hydration-site analysis suggest
water displacement to contribute favorably to ligand binding. A cysteine
residue, exclusively present in bacterial TGTs, serves as gatekeeper
of the transient subpocket. It becomes accessible upon pocket opening
for selective covalent attachment of electrophilic ligands in eubacterial
TGTs.