Abstract
This paper describes the rational design, synthesis, and biological evaluation of a new generation of inhibitors of the bacterial enzyme tRNA‐guanine transglycosylase (TGT), which has been identified as a new target in the fight against bacillary dysentery (Shigellosis). The enzyme catalyzes the exchange of guanine in the anticodon wobble position of tRNA by the modified base preQ~1~, a guanine derivative, according to a ping‐pong mechanism involving a covalent TGT‐tRNA intermediate (Fig. 2). Based on computer modeling (Fig. 3), lin‐benzoguanine (6‐aminoimidazol[4,5‐g]quinazolin‐8(7__H__)‐one (2)) was selected as an extended central scaffold, to form up to seven in‐plane intermolecular H‐bonds with the protein while sandwiching between Tyr106 and Met260. Versatile synthetic protocols were developed for the synthesis of 2, and derivatives with phenyl, benzyl, and 2‐phenylethyl side chains (i.e., 16, 17a, and 12a, 12b, 13, 17, resp.) to reach into the lipophilic pocket lined by Val282, Val45, and Leu68 (Schemes 1–3). To account for the limited solubility of the new ligands and in consequence of a recently developed detailed understanding of the mechanism of TGT catalysis (Fig. 2), the enzyme kinetic assay was completely redesigned, providing competitive (K~ic~) and uncompetitive (K~iu~) inhibition constants with respect to tRNA binding by TGT. The modifications of the various parameters in the new assay are described in detail. Binding affinities of the new inhibitors were found to be in the single‐digit micromolar range (K~ic~ values, Fig. 8). Decoration of the lin‐benzoguanine scaffold with lipophilic residues only gave a modest improvement in biological activity which was explained on structural grounds with the help of four crystal structures (Fig. 10) obtained by soaking the protein with inhibitors 2 and 12a–12c. Both biochemical and biostructural analyses reported in this paper provide a fertile basis for the development of more potent future generations of TGT inhibitors.