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Structural and redox plasticity in the heterodimeric periplasmic nitrate reductase

Nature Structural & Molecular Biology volume 10, pages 928–934 (2003)Cite this article

Abstract

The structure of the respiratory nitrate reductase (NapAB) from Rhodobacter sphaeroides, the periplasmic heterodimeric enzyme responsible for the first step in the denitrification process, has been determined at a resolution of 3.2 Å. The di-heme electron transfer small subunit NapB binds to the large subunit with heme II in close proximity to the [4Fe-4S] cluster of NapA. A total of 57 residues at the N- and C-terminal extremities of NapB adopt an extended conformation, embracing the NapA subunit and largely contributing to the total area of 5,900 Å2 buried in the complex. Complex formation was studied further by measuring the variation of the redox potentials of all the cofactors upon binding. The marked effects observed are interpreted in light of the three-dimensional structure and depict a plasticity that contributes to an efficient electron transfer in the complex from the heme I of NapB to the molybdenum catalytic site of NapA.

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Figure 1: Stereo views of the overall fold of the NapAB complex.
Figure 2: Comparisons of NapAB complex with the structures of the uncomplexed subunits.
Figure 3: Details of the intersubunit interface between NapA and NapB.
Figure 4: Kinetics of nitrate reduction and estimation of the binding affinity between NapA and NapB.
Figure 5: Midpoint redox potentials of the cofactors in both the free (red, NapB; green, NapA) and bound subunits (black) measured by optical spectroscopy and EPR titration as a function of the distances between the cofactors.

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Acknowledgements

We would like to thank the European Synchrotron Radiation Facility and especially the staff of the BM30 and ID14 beamlines (Grenoble, France) for the provision of data collection and processing facilities, the staff of the Laboratoire de Cristallographie et de Cristallogenèse des Protéines of the Institut de Biologie Structurale for encouragement, the Progamme de Toxicologie Nucléaire of the Comissariat à l'Energie atomique for financial support, and J. Lavergne and A. Verméglio for discussions and reading of the manuscript.

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  1. CEA/Cadarache, DSV, DEVM, Laboratoire de Bioénergétique Cellulaire, 13108 St Paul lez, Durance, Cedex, France

    Pascal Arnoux, Monique Sabaty, Jean Alric, Jean-Marc Adriano & David Pignol

  2. Laboratoire de Bioénergétique et Ingénierie des Protéines, CNRS, IBSM, 31 Chemin Joseph Aiguier, Marseille, 13402, Cedex 9, France

    Bettina Frangioni & Bruno Guigliarelli

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  1. Pascal Arnoux
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Correspondence to Pascal Arnoux or David Pignol.

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Arnoux, P., Sabaty, M., Alric, J. et al. Structural and redox plasticity in the heterodimeric periplasmic nitrate reductase. Nat Struct Mol Biol 10, 928–934 (2003). https://doi.org/10.1038/nsb994

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