A Conformational Switch Triggers Nitrogenase Protection from Oxygen Damage by Shethna Protein II (FeSII)

The two-component metalloprotein nitrogenase catalyzes the reductive fixation of atmospheric dinitrogen into bioavailable ammonium in diazotrophic prokaryotes. The process requires an efficient energy metabolism, so that although the metal clusters of nitrogenase rapidly decompose in the presence of dioxygen, many free-living diazotrophs are obligate aerobes. In order to retain the functionality of the nitrogen-fixing enzyme, some of these are able to rapidly switch-off'' nitrogenase, by shifting the enzyme into an inactive but oxygen-tolerant state. Under these conditions the two components of nitrogenase form a stable, ternary complex with a small [2Fe:25] ferredoxin termed FeSII or the Shethna protein II. Here we have produced and isolated Azotobacter vinelandii FeS II and have determined its three-dimensional structure to 2.1 angstrom resolution by X-ray diffraction. In the crystals, the dimeric protein was present in two distinct states that differ in the conformation of an extended loop in close proximity to the iron sulfur cluster. We show that this rearrangement is redox-dependent and forms the molecular basis for oxygen-dependent conformational protection of nitrogenase. Protection assays highlight that FeSII binds to a preformed complex of MoFe and Fe protein upon activation, primarily through electrostatic interactions. The surface properties and known complexes of nitrogenase component proteins allow us to propose a model of the conformationally protected ternary complex of nitrogenase.