Spectroscopic studies of biological dinitrogen reduction

The nitrogenase family of enzymes is responsible for nearly all biological nitrogen fixation and has attracted intense interest for decades due to its ability to reduce N2 to NH3 under ambient conditions. The active site of nitrogenase is a MoFe7S9C cluster, the so-called iron molybdenum cofactor (or FeMoco), which is the site of substrate activation. In recent years, our understanding of the mechanism has advanced substantially, however, many open questions remain as to the specific site of reduction and protonation events during the catalytic cycle. Recently, we have shown that selective Se substitution combined with selenium Kα high-energy resolution fluorescence detected X-ray absorption spectroscopy (Se Kα HERFD XAS) can reveal a more detailed picture of the electronic structure of the active site. In the present study, we utilize a combination of electron paramagnetic resonance (EPR) and Se Kα HERFD XAS, together with extended X-ray absorption fine structure (EXAFS), to understand the mechanism of Se insertion and the nature of early intermediates in the catalytic cycle. In addition, recent spectroelectrochemical studies of a nitrogenase biohybrid system will be presented.