Elucidating the structural basis of Fgd1-mediated resistance to Delamanid in Mycobacterium tuberculosis.

Delamanid and Pretomanid are nitro-dihydro-imidazooxazole derivatives essential for the treatment of multidrug-resistant tuberculosis (MDR-TB). The emergence of tuberculosis resistance to Delamanid necessitates understanding the molecular mechanisms underlying this resistance. This study focuses on the structural analysis of the fgd1 protein in both wild-type and mutant forms to explain the molecular basis of Delamanid resistance. Since the fgd1 complex with the F420 cofactor is responsible for the activation of Delamanid, we investigated the impact of mutations on the structural stability, primarily using molecular dynamics simulations, and electronic properties using quantum mechanics/molecular mechanics (QM/MM) studies of the fgd1-F420 complex. Among the pool of available mutants, we selected 12 fgd1 mutants (K9A, L70R, G71D, Q88E, M93R, N112K, A176D, G191D, K198A, E230K, W284S, and G314E) that exhibited resistance to Delamanid. A significant change in the protein and cofactor conformation was observed in some mutants compared to that in the wild-type, with increased RMSD and RMSF. To further analyze the binding affinity between fgd1-F420 WT and the mutants in detail, we performed MM-GBSA calculations and observed reduced F420 binding for K9A, G71D, N112K, K198A, and W284S mutations. Additionally, QM/MM calculations demonstrated how mutations influenced the electronic environment of fgd1, indicating that all mutations, except K9A, led to reduced stability. These alterations suggest that mutations may alter the local electronic environment, potentially affecting the cofactor binding and catalytic processes. Overall, these results provide new structural insights and vital clues for designing novel inhibitors to combat nitroimidazole resistance.