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System for Efficacy and Cytotoxicity Screening of Inhibitors Targeting Intracellular Mycobacterium tuberculosis
Published on: April 5, 2017
Structural basis of substrate accommodation and thiourea-mediated inhibition of the Mycobacterium tuberculosis
Soumya Sarathi Ganguly1, Rituparna Saha1, Bina Kumari Singh1
1Department of Bioscience and Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal, India.
Abstract:
Mycobacterium tuberculosis relies on the fatty acid synthase II (FAS II) pathway for the biosynthesis of mycolic acids, which are essential components of its lipid-rich cell envelope and major contributors to intrinsic drug resistance. The HadAB dehydratase complex catalyzes a critical dehydration step during early meromycolate chain elongation; however, the structural basis governing substrate accommodation and inhibitor recognition has remained unclear. Here, we report the first substrate-bound crystal structure of M. tuberculosis HadAB in complex with Palmitoyl-CoA at 1.69 Å resolution, revealing the architecture and dynamic remodeling of the acyl housing tunnel. The structure reveals that the tunnel is formed at the HadA-HadB interface, with HadA contributing primarily to substrate accommodation and HadB providing the catalytic machinery. Comparison with the previously determined apo-structure reveals substrate-induced displacement of the α4 hot-dog helix of HadA, resulting in elongation and reshaping of the tunnel to accommodate fatty acyl intermediates. The structure further reveals the positioning of Cys61, at the distal end of the acyl housing tunnel, and Cys105, which mediate covalent inhibition by thiourea-derived compounds. Biochemical analyses demonstrate selective activation of Thioacetazone and SPA0355 by mycobacterial Baeyer-Villiger monooxygenases, EthA and MymA, leading to cysteine-dependent modification and inhibition of HadAB. A cell viability assay against Mycobacterium smegmatis demonstrated promising antimycobacterial activity for SPA0355, similar to Thioacetazone. Furthermore, modelling of HadAB-AcpM provided insights into the interaction between HadAB and its physiological acyl carrier protein AcpM. Together, these findings establish the molecular basis of substrate recognition, tunnel plasticity, and prodrug-mediated inhibition of HadAB, providing a framework for structure-guided development of therapeutics that target mycobacterial lipid biosynthesis.
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