A useful guide for medicinal chemists to design PknG inhibitors: structure-activity relationships, structural requirements and molecular dynamics simulations

Although treatable, tuberculosis remains a leading cause of death, and multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains have intensified its burden. The serine/threonine kinase G (PknG) of Mycobacterium tuberculosis is an attractive target because it enables immune evasion. More than 100 PknG inhibitors have been reported, yet none has progressed to regulatory approval and explicit design rules are lacking. A comprehensive chemoinformatic analysis was performed on reported PknG inhibitors. Principal component analysis (PCA) was used to explore chemical space. Furthermore, docking studies were used to conduct a structure-activity relationship (SAR) assessment and identify key molecular features associated with enhanced inhibition. Molecular dynamics simulations of representative PknG–inhibitor complexes were then carried out to characterize the stability and persistence of key binding interactions and to compute per-residue binding free energy contributions. Based on these trajectories, hydration analyses using the V_4S index were applied to identify regions of the binding site where water removal is energetically favorable, thereby enabling direct ligand–protein interactions. Four interaction regions were identified: the hinge region (HR), catalytic region (CR), core and hydrophobic region (HyR). Engagement of the HR was found essential for binding, while interactions within the CR significantly enhanced inhibitory activity, improving potency by up to one order of magnitude (A9 IC₅₀:0.01 µM vs. AX20017:0.3 µM). These findings define a simple structure-based strategy for designing PknG inhibitors, providing a rule of thumb that, together with computational methods, may guide rational therapeutic development.