Dethiobiotin synthetase fromMycobacterium tuberculosis(MtDTBS) is a promising antituberculosis drug target. Small-molecule inhibitors that targetMtDTBS provide a route towards new therapeutics for the treatment of antibiotic-resistant tuberculosis. Adenosine diphosphate (ADP) is an inhibitor ofMtDTBS; however, structural studies into its mechanism of inhibition have been unsuccessful owing to competitive binding to the enzyme by crystallographic precipitants such as citrate and sulfate. Here, a crystallographic technique termed precipitant–ligand exchange has been developed to exchange protein-bound precipitants with ligands of interest. Proof of concept for the exchange method was demonstrated using cytidine triphosphate (CTP), which adopted the same binding mechanism as that obtained with traditional crystal-soaking techniques. Precipitant–ligand exchange also yielded the previously intractable structure ofMtDTBS in complex with ADP solved to 2.4 Å resolution. This result demonstrates the utility of precipitant–ligand exchange, which may be widely applicable to protein crystallography.
