Mechanism of N2 Reduction Catalyzed by Fe-Nitrogenase Involves Reductive Elimination of H2

Harris, Derek F.; Lukoyanov, Dmitriy A.; Shaw, Sudipta; Compton, Phil; Tokmina-Lukaszewska, Monika; Bothner, Brian; Kelleher, Neil; Dean, Dennis R.; Hoffman, Brian M.; Seefeldt, Lance C.

doi:10.1021/acs.biochem.7b01142

Mechanism of N₂ Reduction Catalyzed by Fe-Nitrogenase Involves Reductive Elimination of H₂

Journal Article · Wed Dec 27 23:00:00 EST 2017 · Biochemistry

DOI:https://doi.org/10.1021/acs.biochem.7b01142· OSTI ID:1470112

^[1]; Lukoyanov, Dmitriy A. ^[2]; Shaw, Sudipta ^[1]; Compton, Phil ^[2]; Tokmina-Lukaszewska, Monika ^[3]; Bothner, Brian ^[3]; ^[2]; Dean, Dennis R. ^[4]; ^[2]; ^[1]

Utah State Univ., Logan, UT (United States)
Northwestern Univ., Evanston, IL (United States)
Montana State Univ., Bozeman, MT (United States)
Virginia Polytechnic Inst. and State Univ. (Virginia Tech), Blacksburg, VA (United States)

Of the three forms of nitrogenase (Mo-nitrogenase, V-nitrogenase, and Fe-nitrogenase), Fe-nitrogenase has the poorest ratio of N₂ reduction relative to H₂ evolution. Current work on the Mo-nitrogenase has revealed that reductive elimination of two bridging Fe–H–Fe hydrides on the active site FeMo-cofactor to yield H₂ is a key feature in the N₂ reduction mechanism. The N₂ reduction mechanism for the Fe-nitrogenase active site FeFe-cofactor was unknown. Here, we have purified both component proteins of the Fe-nitrogenase system, the electron-delivery Fe protein (AnfH) plus the catalytic FeFe protein (AnfDGK), and established its mechanism of N₂ reduction. Inductively coupled plasma optical emission spectroscopy and mass spectrometry illustrate that the FeFe protein component does not contain significant amounts of Mo or V, thus ruling out a requirement of these metals for N₂ reduction. The fully functioning Fe-nitrogenase system was found to have specific activities for N₂ reduction (1 atm) of 181 ± 5 nmol NH₃ min^–1 mg^–1 FeFe protein, for proton reduction (in the absence of N₂) of 1085 ± 41 nmol H₂ min^–1 mg^–1 FeFe protein, and for acetylene reduction (0.3 atm) of 306 ± 3 nmol C₂H₄ min^–1 mg^–1 FeFe protein. Under turnover conditions, N₂ reduction is inhibited by H₂ and the enzyme catalyzes the formation of HD when presented with N₂ and D₂. These observations are explained by the accumulation of four reducing equivalents as two metal-bound hydrides and two protons at the FeFe-cofactor, with activation for N₂ reduction occurring by reductive elimination of H₂.

View Accepted Manuscript (DOE)

Research Organization:: Montana State Univ., Bozeman, MT (United States). Energy Frontier Resarch Center (EFRC) Center for Biological Electron Transfer and Catalysis (BETCy)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)

Grant/Contract Number:: SC0012518

OSTI ID:: 1470112

Journal Information:: Biochemistry, Journal Name: Biochemistry Journal Issue: 5 Vol. 57; ISSN 0006-2960

Publisher:: American Chemical Society (ACS)Copyright Statement

Country of Publication:: United States

Language:: English

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37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
Alternative nitrogenase
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