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Cyclin dependent kinase 2 (CDK2) is an established target protein for therapeutic intervention in various diseases, including cancer. Reported inhibitors of CDK2 target the ATP-binding pocket to inhibit the kinase activity. Many small molecule CDK2 inhibitors have been discovered and their crystal structure with CDK2 or CDK2/Cyclin A complex have been published. NU6140 is a CDK2 inhibitor with moderate potency and selectivity. Herein we report the co-crystal structure determination of NU6140 in complex with CDK2 and confirmation of the binding using various biophysical methods. Our data shows that NU6140 bind to CDK2 with a Kd of 800 nM as determined by SPR and stabilizes the protein against thermal denaturation (ΔTm- 5 ℃). The co-crystal structure determined in our study shows that NU6140 binds in the ATP binding pocket as expected for this class of compounds and interacts with Leu83 and Glu81 with regular hydrogen bonds and with Asp145 via water mediated H-bond. Based on these data, we propose structural modifications of NU6140 to introduce new interactions with CDK2 that can improve its potency while retaining the selectivity.

Acinetobacter baumannii is an opportunistic Gram-negative bacterial pathogen, associated mostly with hospital-acquired infections. The emergence of drug resistance strains made it necessary to explore new pathways for the development of more effective antibiotics. Enoyl CoA reductase (FabI), a key enzyme in the fatty acid biosynthesis (FAS) pathway, has emerged as a potential target for antibacterial drug development. Earlier reports show that the lead SaFabI inhibitor AFN-1252 can inhibit FabI from other organisms including Escherichia coli and Burkholderia pseudomallei, but with differential potency. In the present work, we show that AFN-1252 is a moderate inhibitor of AbFabI with an IC50 of 216 nM. AFN-1252 stabilized AbFabI with a 4.2°C increase in the melting temperature (Tm ) and, interestingly, the stabilization effect was significantly increased in presence of the cofactor NADH (∆Tm = 17°C), suggesting the formation of a ternary complex AbFabI: AFN-1252: NADH. X-ray crystallography studies of AbFabI co-crystalized with AFN-1252 and NADH confirmed the ternary complex formation. The critical interactions of AFN-1252 with AbFabI and NADH identified from the co-crystal structure may facilitate the design and development of new drugs against A. baumannii infections by targeting the FAS pathway.