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Structural and Functional Characterization of Proteins involved in PDIM biosynthesis in Mycobacterium tuberculosis

par Yang, Dong
Président du jury Delporte, Cédric
Promoteur Fontaine, Véronique
Co-Promoteur Vandenbussche, Guy
Publication Non publié, 2020-04-27

Thèse de doctorat

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Résumé :

Tuberculosis is an infectious disease caused by the pathogen Mycobacterium tuberculosis which is still a public health threat. Due to the appearance of multidrug-resistant M. tuberculosis strains, it is urgent to find new drugs to fight against tuberculosis, especially against the extensively drug resistant tuberculosis. The unique impermeable mycobacteria cell wall, rich in lipids, is a key barrier to anti-tuberculosis drug uptake. One of these lipids, the phthiocerol dimycocerosate (PDIM), is related to mycobacteria virulence, drug resistance and mycobacteria cell wall integrity. Therefore, research focusing on PDIM biosynthesis potentially provides a new route for the development of new anti-tuberculosis drugs.In this context, we investigated two proteins, the thioesterase TesA and the chaperonin GroEL1, that are involved in the PDIM biosynthesis, and studied their structural and functional properties. Firstly, using various substrates, we demonstrated that TesA displays thioesterase and esterase activities and showed that TesA undergoes substrate-dimerization. A new TesA inhibitor, methyl arachidonyl fluorophosphonate (MAFP), was identified that irreversibly binds to the TesA catalytic serine residue. Furthermore, MAFP can increase the susceptibility to vancomycin in a TesA inhibition dependent manner while it can also affect the M. bovis BCG biofilm formation through broader action mechanisms. Secondly, we observed that the mycobacterial GroEL1, unlike the other chaperonins, E. coli GroEL and mycobacteria GroEL2, increases copper tolerance. We demonstrated that the copper can specifically bind to the GroEL1 with a strong affinity for its natural carboxyl terminal histidine-rich region. The classical feature of chaperonins is also increased in the presence of copper. These results suggest that GroEL1 could help mycobacteria to tolerate high bactericidal metal concentrations during macrophage infection, potentially through a metal storage mechanism. Last but not the least, we identified new mycobacterial GroEL1 and GroEL2 activities. In one word, the biochemical properties of two essential proteins, GroEL1 and TesA, involved in PDIM biosynthesis in M. tuberculosis have been further elucidated. The implications of these results are important not only to develop new antituberculous drugs but also to fight drug resistance.