Dynamic closure of the Mycobacterium tuberculosis InhA cofactor site reveals heterogeneous routes to isoniazid resistance
Year: 2026
Type: preprint
Abstract: Isoniazid remains a cornerstone of tuberculosis therapy, yet resistance-associated mutations in its target, InhA, are not fully explained by reduced NADH binding alone. Here, we combine X-ray crystallography, cryo-electron microscopy, calorimetry, thermal stability profiling, and multi-scale molecular dynamics to dissect ten clinically observed InhA missense variants. The structures show that many substitutions preserve the canonical cofactor-bound architecture, but thermodynamic and simulation analyses reveal divergent routes to resistance. Active-site variants remodel water and hydrophobic networks to weaken NADH binding, whereas I95P and several interface-proximal variants populate a coenzyme closed-loop state that occludes the nicotinamide/adduct-binding pocket. A190S and Y259H instead appear to preserve NADH affinity while rigidifying the Phe149–Asp150 region required to accommodate the larger INH–NAD adduct. These findings establish InhA coding-region resistance as a spectrum of local, allosteric, and adduct-selective mechanisms, providing a framework for interpreting clinical variants and designing next-generation InhA inhibitors.
Source: ChemRxiv
Language: English
Subfield: Infectious Diseases
Field: Medicine
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W7165004950