ANNOUNCEMENTS
The emergence of the multi- and extensively drug-resistant (MDR and XDR) strains of Mycobacterium tuberculosis (M.tb) necessitates innovative therapeutic approaches to mitigate Tuberculosis (TB) infection. The impermeable thick, waxy lipid layer, composed of a unique mycolic acid, is a key factor contributing to bacterial pathogenicity and conferring resistance to conventional drugs. In this study, novel strategies were employed to combat the drug resistance by targeting the critical methyltransferase enzymes- Mycolic acid methyltransferase 1 (MmaA1), and Methoxy mycolic acid synthase 3 (MmaA3), which are involved in the biosynthesis of mycolic acids. The MmaA1 is chiefly responsible for the conversion of cis-olefin into trans-olefin and the introduction of an allylic methyl branch as a precursor to both the methoxy and ketone-containing mycolic acids. MmaA3 catalyzes the O-methylation of the hydroxy group of hydroxy mycolate to form methyl ether. Here, two approaches have been employed for the structural characterization of methyltransferase proteins. In the first approach, four crystal structures of MmaA1 were reported in the apo-form and in complex with the ligands S-adenosyl-L-methionine (SAM), S-adenosyl-L-homocysteine (SAH), and Sinefungin (SFG) at 1.4-1.9 Å resolution. Structural characterization revealed a dynamic loop (between α7 and α8) and its critical role in structural integrity and efficient catalysis. Additionally, a C268 exhibits dual behavior, exists in a reduced state in the SAM-bound structure, and transits to an oxidized state in the SAH-bound form. The conserved cysteine residue might have a role in the regulation of the substrate binding pocket in MmaA1. In the second approach, a threedimensional structure of MmaA3 was generated using homolog-based modeling. High-throughput virtual screening was performed against the MmaA3 model, utilizing diverse libraries. Rigorous computational analyses revealed the putative binding partners with MmaA3. The potent binder DG70 was screened based on the molecular dynamics (MD) simulations and binding free energy calculations. Later, the protein-ligand (DG70) interactions and functional activity of the protein(s) were validated utilizing in-vitro studies. Thus, these findings aimed to contribute to the development of therapeutic strategies and overcoming the adverse effects of MDR and XDR-TB.
