In vivo detection of nitrogenase cofactor synthesis intermediates

Engineering nitrogen fixation in crops is an essential step towards reducing the overuse of expensive and polluting nitrogen fertilizers. While substantial efforts have been done towards transferring many of the proteins involved in nitrogenase synthesis, information on how they collaborate to synthesize nitrogenase cofactors in planta is lagging. This is due to to cofactor synthesis intermediates not having any enzymatic activity to trace in vitro. As an alternative, we propose to take advantage of ID21 imaging and Fe and S near-edge spectroscopy capabilitiesto detect in vivo the signature of some of the unique intermediate cofactors synthesized. In a first step, we will detect these molecules in a model nitrogen fixing bacteria and in mutants where these intermediates accumulate, and then, we will attempt to detect them in a simple eukaryote that already expresses some of these components.