title: Redox-coupled structural changes in nitrite reductase revealed by serial femtosecond and microfocus crystallography.
abstract: Serial femtosecond crystallography (SFX) has enabled the damage-free structural determination of metalloenzymes and filled the gaps of our knowledge between crystallographic and spectroscopic data. Crystallographers, however, scarcely know whether the rising technique provides truly new structural insights into mechanisms of metalloenzymes partly because of limited resolutions. Copper nitrite reductase (CuNiR), which converts nitrite to nitric oxide in denitrification, has been extensively studied by synchrotron radiation crystallography (SRX). Although catalytic Cu (Type 2 copper (T2Cu)) of CuNiR had been suspected to tolerate X-ray photoreduction, we here showed that T2Cu in the form free of nitrite is reduced and changes its coordination structure in SRX. Moreover, we determined the completely oxidized CuNiR structure at 1.43 Å resolution with SFX. Comparison between the high-resolution SFX and SRX data revealed the subtle structural change of a catalytic His residue by X-ray photoreduction. This finding, which SRX has failed to uncover, provides new insight into the reaction mechanism of CuNiR.
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan;
authors.forename: Yohta
authors.initials: Y
authors.lastname: Fukuda
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Ka Man
authors.initials: KM
authors.lastname: Tse
authors.affiliation: Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan; RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp.
authors.forename: Mamoru
authors.initials: M
authors.lastname: Suzuki
authors.affiliation: Department of Biology, University of Konstanz, D-78457 Konstanz, Germany; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Kay
authors.initials: K
authors.lastname: Diederichs
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Kunio
authors.initials: K
authors.lastname: Hirata
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Takanori
authors.initials: T
authors.lastname: Nakane
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Michihiro
authors.initials: M
authors.lastname: Sugahara
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Eriko
authors.initials: E
authors.lastname: Nango
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5198, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Kensuke
authors.initials: K
authors.lastname: Tono
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5198, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Yasumasa
authors.initials: Y
authors.lastname: Joti
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5198, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kameshima
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; Department of Physics, Pohang University of Science and Technology, Pohang 790-784, Korea; and mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Changyong
authors.initials: C
authors.lastname: Song
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Takaki
authors.initials: T
authors.lastname: Hatsui
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Makina
authors.initials: M
authors.lastname: Yabashi
authors.affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan; mamoru.suzuki@protein.osaka-u.ac.jp inouet@chem.eng.osaka-u.ac.jp mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Osamu
authors.initials: O
authors.lastname: Nureki
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan;
authors.forename: Hiroyoshi
authors.initials: H
authors.lastname: Matsumura
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; inouet@chem.eng.osaka-u.ac.jp.
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Inoue
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo-cho, Sayo-gun, Hyogo 679-5148, Japan; Department of Cell Biology, Graduate School of Medicine, Kyoto University, Yoshidakonoe-cho, Sakyo-ku, Kyoto, 606-8501, Japan.
authors.forename: So
authors.initials: S
authors.lastname: Iwata
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; mizohata@chem.eng.osaka-u.ac.jp.
authors.forename: Eiichi
authors.initials: E
authors.lastname: Mizohata
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D020134:Catalytic Domain; D003300:Copper / Q000737:chemistry*; D056506:Geobacillus / Q000201:enzymology*; D009572:Nitrite Reductases / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D003300:Copper; D009572:Nitrite Reductases
keywords: X-ray free-electron laser; copper; electron transfer; enzyme; serial femtosecond crystallography