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Two distinct conformational states of Mycobacterium tuberculosis virulent factor ESAT-6 are behind the discrepancy around its biological functions

Abstract : Early secreted antigenic target 6 kDa (ESAT-6) and culture filtrate protein 10 kDa (CFP-10) are complex proteins secreted by Mycobacterium tuberculosis that play a major role in the pathogenesis of tuberculosis. However, studies focusing on the biological functions of ESAT-6 led to discordant results and the role of ESAT-6 remains controversial. In the present study, we aim to address a potential explanation for this discrepancy and to highlight the physiological impact of two conformational states of ESAT-6. Analysis of a recombinant form of ESAT-6 by native gel electrophoresis, size exclusion chromatography and CD spectroscopy revealed that ESAT-6 forms dimers/multimers with higher molecular weight, which disappeared under the action of the detergent amidosulfobetaine-14 (ASB), giving rise to another conformational state of the protein. NMR has further indicated that ASB-treated versus nontreated ESAT-6 adopted distinct structural forms but with no well defined tertiary structure. However, protein-protein docking analysis favored a dimeric state of ESAT-6. Interestingly, the two preparations presented opposing effects on mycobacterial infectivity, as well as macrophage survival, interferon-γ secretion and membrane pore formation. Thereafter, we generated a recombinant form of the physiological heterodimer ESAT-6/CFP-10 that ASB was also able to dissociate and which showed functions similar to those of ESAT-6 dimers/multimers. Our data suggest that, in the absence of CFP-10, the hydrophobic regions of the ESAT-6 can form dimers/multimers, mimicking the ESAT-6/CFP-10 heterodimer, whereas their dissociation generates a protein presenting entirely different activities. Overall, the present study clarifies the intriguing divergences between reports that could be attributed to the ESAT-6 oligomeric state and sheds light on its importance for a better comprehension of the physiopathology of tuberculosis.
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https://hal-univ-rennes1.archives-ouvertes.fr/hal-01188227
Contributor : Laurent Jonchère <>
Submitted on : Friday, August 28, 2015 - 4:25:40 PM
Last modification on : Wednesday, July 8, 2020 - 12:42:50 PM

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Amira Refai, Meriam Haoues, Houcemeddine Othman, Mohamed-Ridha Barbouche, Philippe Moua, et al.. Two distinct conformational states of Mycobacterium tuberculosis virulent factor ESAT-6 are behind the discrepancy around its biological functions. FEBS Journal, Wiley, 2015, 282 (21), pp.4114-4129. ⟨10.1111/febs.13408⟩. ⟨hal-01188227⟩

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