Transferred NOE and Saturation Transfer Difference NMR Studies of Novobiocin Binding to EnvZ Suggest Binding Mode Similar to DNA Gyrase

Histidine protein kinases (HPKs) are a class of receptor proteins found in bacterial two‐component signal transduction systems, which allow bacteria to respond to changes in their external environment. To date, there are few potent inhibitors of histidine kinases, despite their potential ability to weaken bacteria against antibiotic treatment. EnvZ is a histidine protein kinase with osmoregulatory function in bacteria with sequence and topological similarity to DNA Gyrase B. DNA Gyrase B has several well‐characterized potent inhibitors, including novobiocin and clorobiocin which have detailed structures in complex. With fluorescence competition experiments, we have determined that novobiocin binds to EnvZ with a novoKD 120 ± 20 μm. NMR transferred NOE (trNOE) experiments, and saturation transfer difference (STD) experiments suggest that novobiocin binds to EnvZ in a conformation and orientation similar to its binding with DNA Gyrase B. These experiments suggest some similarity in the pocket despite weaker affinity for EnvZ by novobiocin.