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Crystal structure of quinol-dependent nitric oxide reductase from Geobacillus stearothermophilus

Nature Structural & Molecular Biology volume 19, pages 238245 (2012) | Download Citation

Abstract

The structure of quinol-dependent nitric oxide reductase (qNOR) from G. stearothermophilus, which catalyzes the reduction of NO to produce the major ozone-depleting gas N2O, has been characterized at 2.5 Å resolution. The overall fold of qNOR is similar to that of cytochrome c–dependent NOR (cNOR), and some structural features that are characteristic of cNOR, such as the calcium binding site and hydrophilic cytochrome c domain, are observed in qNOR, even though it harbors no heme c. In contrast to cNOR, structure-based mutagenesis and molecular dynamics simulation studies of qNOR suggest that a water channel from the cytoplasm can serve as a proton transfer pathway for the catalytic reaction. Further structural comparison of qNOR with cNOR and aerobic and microaerobic respiratory oxidases elucidates their evolutionary relationship and possible functional conversions.

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Acknowledgements

We thank Y. Shimomura for support in the preparation of qNOR and the staff of the SPring-8 beamlines for their help with diffraction measurements. This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology (21245041; T.T., H.S., S.N., A.V.P., Y. Sugita and Y. Shiro).

Author information

Author notes

    • Yushi Matsumoto
    • , Tomoya Hino
    •  & Shingo Nagano

    Present addresses: Division of Protein Chemistry, Post-Genome Science Center, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan (Y.M.); Department of Chemistry and Biotechnology, Graduate School of Engineering, Tottori University, Tottori, Japan (T.H. and S.N.).

Affiliations

  1. Biometal Science Laboratory, RIKEN SPring-8 Center, Sayo, Hyogo, Japan.

    • Yushi Matsumoto
    • , Takehiko Tosha
    • , Tomoya Hino
    • , Hiroshi Sugimoto
    • , Shingo Nagano
    •  & Yoshitsugu Shiro
  2. Theoretical Biochemistry Laboratory, RIKEN Advanced Science Institute, Wako, Saitama, Japan.

    • Andrei V Pisliakov
    •  & Yuji Sugita

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Contributions

Y.M. was responsible for cloning the qNOR gene and constructing the expression system as well as for purifying, characterizing and crystallizing the protein. T.T. carried out the enzyme assays and the ICP-AES metal analysis. T.H. purified P. aeruginosa cNOR. Y.M. and H.S. collected, processed and refined the X-ray data. A.V.P. carried out the molecular dynamics simulation. T.H., T.T., Y.M., S.N., Y. Sugita and Y. Shiro designed the study. Y.M., T.T., A.V.P., S.N. and Y. Shiro prepared the manuscript. All authors analyzed the data and discussed the results.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Shingo Nagano or Yoshitsugu Shiro.

Supplementary information

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  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–7 and Supplementary Methods

Image files

  1. 1.

    Supplementary Movie 1

    Dynamics in the water channel in MD simulation. Time-dependent dynamics, such as side chains fluctuations, moving water molecules, and transient hydrogen-bond networks can be seen. Water molecules in the water channel are shown in yellow, and bulk water around the channel entrance is shown as red and white lines. Hydrogen-bonds are shown as dashed green lines.

  2. 2.

    Supplementary Movie 2

    Motion of selected water molecules along the water channel in the MD simulation. Eight individual waters are shown as large spheres and are highlighted in different colors. Other water molecules inside the water channel and in the bulk around the channel entrance are shown as yellow sticks. The selected water molecules come into the water channel via the entry site, exchange with waters inside the channel, travel up to the binuclear active center, and eventually return to the bulk. The water molecules are highly mobile and move easily along the channel on a short timescale. There are several stable positions (“traps”) where water molecules stay near residues for a long time (typically, nanoseconds), before moving further along the channel.

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DOI

https://doi.org/10.1038/nsmb.2213

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