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Updated: Jun 8, 2025

Expression and Purification of Nuclease-Free Oxygen Scavenger Protocatechuate 3,4-Dioxygenase
Published on: November 8, 2019
Cross-Coupling of Mo- and V-Nitrogenases Permits Protein-Mediated Protection from Oxygen Deactivation
Daniel Ratcliff1,2, G C Danielle Sedoh1,2,3, Ross D Milton1,2
1Department of Inorganic and Analytical Chemistry, Faculty of Science, University of Geneva, Quai Ernest-Ansermet 30, 1205, Geneva, Switzerland.
Abstract:
Nitrogenases catalyze dinitrogen (N2) fixation to ammonia (NH3). While these enzymes are highly sensitive to deactivation by molecular oxygen (O2) they can be produced by obligate aerobes for diazotrophy, necessitating a mechanism by which nitrogenase can be protected from deactivation. In the bacterium Azotobacter vinelandii, one mode of such protection involves an O2-responsive ferredoxin-type protein ("Shethna protein II", or "FeSII") which is thought to bind with Mo-dependent nitrogenase's two component proteins (NifH and NifDK) to form a catalytically stalled yet O2-tolerant tripartite protein complex. This protection mechanism has been reported for Mo-nitrogenase, however, in vitro assays with V-nitrogenase suggest that this mechanism is not universal to the three known nitrogenase isoforms. Here we report that the reductase of the V-nitrogenase (VnfH) can engage in this FeSII-mediated protection mechanism when cross-coupled with Mo-nitrogenase NifDK. Interestingly, the cross-coupling of the Mo-nitrogenase reductase NifH with the V-nitrogenase VnfDGK protein does not yield such protection.
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