Open Access Dissertation
Date of Award
Steven M. Firestine
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. PurK, found only in bacteria, yeast, and fungi, catalyzes the sixth step in purine biosynthesis and has no human homolog. Herein we disclose the discovery of the first PurK inhibitor with submicromolar potency.
PurK 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 PurK. To explore this hypothesis, four compounds possessing different scaffolds were synthesized and tested against PurK from A. clavatus. Only one compound, 1 containing a pyridopyrimidine core, weakly inhibits PurKAsp with an IC50 of 167 μM. Kinetic assays have confirmed that 1 is an ATP-competitive inhibitor. With the guidance of computational modeling, further optimizations have been focused on the ribose and phosphate positions within the ATP-binding site. Both positions gave compounds with improved activities. A double-modified compound, 56, merging the best compound from each position, inhibits PurKAsp with an IC50 of 1.7 µM, a 100-fold improvement in potency. The binding affinity was validated by thermal shift assays. 56 is the most potent ATP-binding site inhibitor against PurKAsp described to date.
Lin, Qian, "Design, Synthesis And Biological Evaluation Of Pyrido[2,3-D]pyrimidines As Inhibitors Of N5-Cair Synthetase" (2018). Wayne State University Dissertations. 2112.