Structural and functional characterization of the Geobacillus copper nitrite reductase: Involvement of the unique N-terminal region in the interprotein electron transfer with its redox partner |
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Authors: | Yohta Fukuda Hiroyasu Koteishi Ryohei Yoneda Taro Tamada Hideto Takami Tsuyoshi Inoue Masaki Nojiri |
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Affiliation: | 1. Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan;2. Department of Materials Chemistry, Graduate School of Engineering, Osaka University, Suita, Osaka 565-0871, Japan;3. Molecular Biology Research Center, Quantum Beam Science Directorate, Japan Atomic Energy Agency, Tokai, Ibaraki 319-1195, Japan;4. Microbial Genome Research Group, Japan Agency of Marine-Earth Science and Technology, Yokosuka, Kanagawa 237-0061, Japan;5. RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo, Hyogo 679-5148, Japan |
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Abstract: | The crystal structures of copper-containing nitrite reductase (CuNiR) from the thermophilic Gram-positive bacterium Geobacillus kaustophilus HTA426 and the amino (N)-terminal 68 residue-deleted mutant were determined at resolutions of 1.3 Å and 1.8 Å, respectively. Both structures show a striking resemblance with the overall structure of the well-known CuNiRs composed of two Greek key β-barrel domains; however, a remarkable structural difference was found in the N-terminal region. The unique region has one β-strand and one α-helix extended to the northern surface of the type-1 copper site. The superposition of the Geobacillus CuNiR model on the electron-transfer complex structure of CuNiR with the redox partner cytochrome c551 in other denitrifier system led us to infer that this region contributes to the transient binding with the partner protein during the interprotein electron transfer reaction in the Geobacillus system. Furthermore, electron-transfer kinetics experiments using N-terminal residue-deleted mutant and the redox partner, Geobacillus cytochrome c551, were carried out. These structural and kinetics studies demonstrate that the region is directly involved in the specific partner recognition. |
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Keywords: | CuNiR, copper-containing nitrite reductase GkNiR, CuNiR from Geobacillus kaustophilus AxNiR, CuNiR from Achromobacter xylosoxidans Cyt c551, cytochrome c551 GkCyt c551, Cyt c551 from Geobacillus kaustophilus AxCyt c551, Cyt c551 from Achromobacter xylosoxidans T1Cu, type 1 copper site T2Cu, type 2 copper site wt, wild-type ET, electron transfer SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis EDTA, ethylenediaminetetraacetic acid Tris, tris (hydroxymethyl) aminomethane RMS, root-mean-square |
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