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Title: Structural characterization of the P1+ intermediate state of the P-cluster of nitrogenase

Abstract

Nitrogenase is the enzyme that reduces atmospheric dinitrogen (N2) to ammonia (NH3) in biological systems. It catalyzes a series of single-electron transfers from the donor iron protein (Fe protein) to the molybdenum–iron protein (MoFe protein) that contains the iron–molybdenum cofactor (FeMo-co) sites where N2 is reduced to NH3. The P-cluster in the MoFe protein functions in nitrogenase catalysis as an intermediate electron carrier between the external electron donor, the Fe protein, and the FeMo-co sites of the MoFe protein. Previous work has revealed that the P-cluster undergoes redox-dependent structural changes and that the transition from the all-ferrous resting (PN) state to the two-electron oxidized P2+ state is accompanied by protein serine hydroxyl and backbone amide ligation to iron. In this work, the MoFe protein was poised at defined potentials with redox mediators in an electrochemical cell, and the three distinct structural states of the P-cluster (P2+, P1+, and PN) were characterized by X-ray crystallography and confirmed by computational analysis. These analyses revealed that the three oxidation states differ in coordination, implicating that the P1+ state retains the serine hydroxyl coordination but lacks the backbone amide coordination observed in the P2+ states. Lastly, these results provide a complete picture of themore » redox-dependent ligand rearrangements of the three P-cluster redox states.« less

Authors:
; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Pacific Northwest National Laboratory (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1772834
Alternate Identifier(s):
OSTI ID: 1461545
Report Number(s):
PNNL-SA-132325
Journal ID: ISSN 0021-9258; S0021925820399488; PII: S0021925820399488
Grant/Contract Number:  
AC05-76RL01830; MCB-1330807
Resource Type:
Published Article
Journal Name:
Journal of Biological Chemistry
Additional Journal Information:
Journal Name: Journal of Biological Chemistry Journal Volume: 293 Journal Issue: 25; Journal ID: ISSN 0021-9258
Publisher:
Elsevier
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; oxidation-reduction (redox); metalloprotein; nitrogenase; enzyme structure; computational biology; nitrogen fixation; nitrogen reduction; P-cluster of MoFe protein; poised states; redox mediators; redox-dependent ligand exchange; [8Fe-7S] cluster

Citation Formats

Keable, Stephen M., Zadvornyy, Oleg A., Johnson, Lewis E., Ginovska, Bojana, Rasmussen, Andrew J., Danyal, Karamatullah, Eilers, Brian J., Prussia, Gregory A., LeVan, Axl X., Raugei, Simone, Seefeldt, Lance C., and Peters, John W. Structural characterization of the P1+ intermediate state of the P-cluster of nitrogenase. United States: N. p., 2018. Web. doi:10.1074/jbc.RA118.002435.
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Keable, Stephen M., Zadvornyy, Oleg A., Johnson, Lewis E., Ginovska, Bojana, Rasmussen, Andrew J., Danyal, Karamatullah, Eilers, Brian J., Prussia, Gregory A., LeVan, Axl X., Raugei, Simone, Seefeldt, Lance C., & Peters, John W. Structural characterization of the P1+ intermediate state of the P-cluster of nitrogenase. United States. https://doi.org/10.1074/jbc.RA118.002435
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Keable, Stephen M., Zadvornyy, Oleg A., Johnson, Lewis E., Ginovska, Bojana, Rasmussen, Andrew J., Danyal, Karamatullah, Eilers, Brian J., Prussia, Gregory A., LeVan, Axl X., Raugei, Simone, Seefeldt, Lance C., and Peters, John W. Fri . "Structural characterization of the P1+ intermediate state of the P-cluster of nitrogenase". United States. https://doi.org/10.1074/jbc.RA118.002435.
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@article{osti_1772834,
title = {Structural characterization of the P1+ intermediate state of the P-cluster of nitrogenase},
author = {Keable, Stephen M. and Zadvornyy, Oleg A. and Johnson, Lewis E. and Ginovska, Bojana and Rasmussen, Andrew J. and Danyal, Karamatullah and Eilers, Brian J. and Prussia, Gregory A. and LeVan, Axl X. and Raugei, Simone and Seefeldt, Lance C. and Peters, John W.},
abstractNote = {Nitrogenase is the enzyme that reduces atmospheric dinitrogen (N2) to ammonia (NH3) in biological systems. It catalyzes a series of single-electron transfers from the donor iron protein (Fe protein) to the molybdenum–iron protein (MoFe protein) that contains the iron–molybdenum cofactor (FeMo-co) sites where N2 is reduced to NH3. The P-cluster in the MoFe protein functions in nitrogenase catalysis as an intermediate electron carrier between the external electron donor, the Fe protein, and the FeMo-co sites of the MoFe protein. Previous work has revealed that the P-cluster undergoes redox-dependent structural changes and that the transition from the all-ferrous resting (PN) state to the two-electron oxidized P2+ state is accompanied by protein serine hydroxyl and backbone amide ligation to iron. In this work, the MoFe protein was poised at defined potentials with redox mediators in an electrochemical cell, and the three distinct structural states of the P-cluster (P2+, P1+, and PN) were characterized by X-ray crystallography and confirmed by computational analysis. These analyses revealed that the three oxidation states differ in coordination, implicating that the P1+ state retains the serine hydroxyl coordination but lacks the backbone amide coordination observed in the P2+ states. Lastly, these results provide a complete picture of the redox-dependent ligand rearrangements of the three P-cluster redox states.},
doi = {10.1074/jbc.RA118.002435},
journal = {Journal of Biological Chemistry},
number = 25,
volume = 293,
place = {United States},
year = {Fri Jun 01 00:00:00 EDT 2018},
month = {Fri Jun 01 00:00:00 EDT 2018}
}
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https://doi.org/10.1074/jbc.RA118.002435
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