The spread of drug-resistant bacterial infections has increased the need for novel antimicrobial agents. One potential but unexplored target is the de novo purine biosynthetic pathway. N5-CAIR synthetase, found only in bacteria, yeast and fungi, catalyzes the sixth step in purine biosynthesis and has no human homolog. N5-CAIR synthetase is a member of the ATP-grasp superfamily of enzymes and recently, nanomolar inhibitors of biotin carboxylase, a related enzyme, were published. We hypothesized that those inhibitors, which target the ATP-binding site, could also inhibit N5-CAIR synthetase. To explore this hypothesis, a collection of twelve compounds consisting of three scaffolds, pyridopyrimidines, aminooxazoles and dihydropyridopyrimidinones were synthesized and tested against Aspergillus N5-CAIR synthetase. Compound 1 had an IC50 of 180 μM. Further optimization has resulted in compound 24 with an IC50 of 15 µM, a 12-fold increase in activity. Kinetic assays have confirmed that these are ATP-competitive inhibitors. Compound 24 is the most potent ATP-binding site inhibitor against N5-CAIR synthetase described to date. To our surprise, we have found that these compounds are weak inhibitors of E. coli N5-CAIR synthetase, with IC50 values greater than 200 µM. This indicated that even for the same enzyme from different organisms, the ATP binding sites are distinct. Such a result has important implications for the selectivity of compounds which target the ATP binding site.

Design, synthesis, and antimicrobial activity of ATP-binding site inhibitors of N5-CAIR synthetase

251st National Meeting (2016)

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