Detailed characterization of protein, peptide or drug interactions with natural membrane is still a challenge. This review focuses on the use of nuclear magnetic resonance (NMR) for the analysis of interaction of molecules with small unilamellar vesicles (SUV). These phospholipid vesicles are often used as model membranes for fluorescence or circular dichroism experiments. The various NMR approaches for studying molecule-lipid association are reviewed. After a brief survey of the SUV characterization, the use of heteronuclear NMR (phosphorous, carbon, fluorine) is described. Applications of proton NMR through transferred nuclear Overhauser effect to perform structural determination of peptide are presented. Special care is finally given to the influence of the kinetic of the interactions for the proton NMR of bound molecules in SUV, which can constitute a good model for the study of dynamical processes at the membrane surface.

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.