The Hha and TomB proteins from Escherichia coli form an oxygen-dependent toxin–antitoxin (TA) system. Here we show that YmoB, the Yersinia orthologue of TomB, and its single cysteine variant [C117S]YmoB can replace TomB as antitoxins in E. coli. In contrast to other TA systems, [C117S]YmoB transiently interacts with Hha (rather than forming a stable complex) and enhances the spontaneous oxidation of the Hha conserved cysteine residue to a -SOxH-containing species (sulfenic, sulfinic or sulfonic acid), which destabilizes the toxin. The nuclear magnetic resonance structure of [C117S]YmoB and the homology model of TomB show that the two proteins form a four-helix bundle with a conserved buried cysteine connected to the exterior by a channel with a diameter comparable to that of an oxygen molecule. The Hha interaction site is located on the opposite side of the helix bundle.

The interaction of low-molecular-weight ligands with proteins in the search for new selective drugs is driving great efforts in the pharmaceutical industry and academic research. The same principles can be applied to solve an equally challenging task: selective monitoring of protein interactions by NMR at concentrations close to physiological ones in unlabelled samples. In this review, we discuss different approaches to this problem through the measurement of either relaxation rates or chemical shifts of free ligand spies in equilibrium with their macromolecular targets.