Uncoupling Nitrogenase: Catalytic Reduction of Hydrazine to Ammonia by a MoFe Protein in the Absence of Fe Protein-ATP
- 2 September 2010
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Chemical Society
- Vol. 132 (38), 13197-13199
- https://doi.org/10.1021/ja1067178
Abstract
The catalytic reduction of hydrazine (N2H4) to ammonia by a β-98Tyr→His MoFe protein in the absence of the Fe protein or ATP is reported. The reduction of N2 or other substrates (e.g., hydrazine, protons, acetylene) by nitrogenase normally requires the transient association of the two nitrogenase component proteins, the Fe protein and the MoFe protein. The Fe protein, with two bound MgATP molecules, transfers one electron to the MoFe protein during each association, coupled to the hydrolysis of two MgATP. All substrate reduction reactions catalyzed by nitrogenase require delivery of electrons by the Fe protein coupled to the hydrolysis of MgATP. We report that when a single amino acid within the MoFe protein (β-98Tyr) is substituted by His, the resulting MoFe protein supports catalytic reduction of the nitrogenous substrate hydrazine (N2H4) to two ammonia molecules when provided with a low potential reductant, polyaminocarboxylate ligated EuII (Em −1.1 V vs NHE). The wild-type and a number of other MoFe proteins with amino acid substitutions do not show significant rates of hydrazine reduction under these conditions, whereas the β-98His MoFe protein catalyzes hydrazine reduction at rates up to 170 nmol NH3/min/mg MoFe protein. This rate of hydrazine reduction is 94% of the rate catalyzed by the β-98His or wild-type MoFe protein when combined with the Fe protein, ATP, and reductant under comparable conditions. The β-98His MoFe protein reduction of hydrazine in the absence of the Fe protein showed saturation kinetics for the concentration of reductant and substrate. The implications of these results in understanding the nitrogenase mechanism are discussed.Keywords
This publication has 16 references indexed in Scilit:
- Conformational Gating of Electron Transfer from the Nitrogenase Fe Protein to MoFe ProteinJournal of the American Chemical Society, 2010
- Mechanism of Mo-Dependent NitrogenaseAnnual Review of Biochemistry, 2009
- Climbing Nitrogenase: Toward a Mechanism of Enzymatic Nitrogen FixationAccounts of Chemical Research, 2009
- Nitrogenase MoFe-Protein at 1.16 Å Resolution: A Central Ligand in the FeMo-CofactorScience, 2002
- Catalytic and Biophysical Properties of a Nitrogenase Apo-MoFe Protein Produced by a nifB-Deletion Mutant of Azotobacter vinelandiiBiochemistry, 1998
- The Chatt cycle and the mechanism of enzymic reduction of molecular nitrogenJBIC Journal of Biological Inorganic Chemistry, 1996
- Mechanism of Molybdenum NitrogenaseChemical Reviews, 1996
- Involvement of the P Cluster in Intramolecular Electron Transfer within the Nitrogenase MoFe ProteinPublished by Elsevier BV ,1995
- Crystallographic Structure of the Nitrogenase Iron Protein from Azotobacter vinelandiiScience, 1992
- Electron-transfer reactions in nitrogen fixation. Part 2. The electrosynthesis of ammonia: identification and estimation of productsJ. Chem. Soc., Dalton Trans., 1986