Abstract
Bacterial histidine kinases (HKs) are quintessential regulatory enzymes found ubiquitously in bacteria. Apart from their regulatory roles, they are also involved in the production of virulence factors and conferring resistance to various antibiotics in pathogenic microbes. We have previously reported compounds that inhibit multiple HKs by targeting the conserved catalytic and ATP-binding (CA) domain. Herein, we conduct a detailed structure-activity relationship assessment of adenine-based inhibitors using biochemical and docking methods. These studies have resulted in several observations. First, interaction of an inhibitor's amine group with the conserved active-site Asp is essential for activity and likely dictates its orientation in the binding pocket. Second, a N-NH-N triad in the inhibitor scaffold is highly preferred for binding to conserved Gly:Asp:Asn residues. Lastly, hydrophobic electron-withdrawing groups at several positions in the adenine core enhance potency. The selectivity of these inhibitors was tested against heat shock protein 90 (HSP90), which possesses a similar ATP-binding fold. We found that groups that target the ATP-lid portion of the catalytic domain, such as a six-membered ring, confer selectivity for HKs.