Protein-Cadmium Interactions in Crowded Biomolecular Environments Probed by In-cell and Lysate NMR Spectroscopy

One of the mechanisms by which toxic metal ions interfere with cellular functions is ionic mimicry, where they bind to protein sites in lieu of native metals Ca ²⁺ and Zn ²⁺ . The influence of crowded intracellular environments on these interactions is not well understood. Here, we demonstrate the application of in-cell and lysate NMR spectroscopy to obtain atomic-level information on how a potent environmental toxin cadmium interacts with its protein targets. The experiments, conducted in intact E. coli cells and their lysates, revealed that Cd ²⁺ can profoundly affect the quinary interactions of its protein partners, and can replace Zn ²⁺ in both labile and non-labile protein structural sites without significant perturbation of the membrane binding function. Surprisingly, in crowded molecular environments Cd ²⁺ can effectively target not only all-sulfur and mixed sulfur/nitrogen but also all-oxygen coordination sites. The sulfur-rich coordination environments show significant promise for bioremedial applications, as demonstrated by the ability of the designed protein scaffold α ₃ DIV to sequester intracellular cadmium. Our data suggests that in-cell NMR spectroscopy is a powerful tool for probing interactions of toxic metal ions with their potential protein targets, and for the assessment of potency of sequestering agents.