Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F

Naoki Yahata, Takashi Saitoh, Yuki Takayama, Kiyoshi Ozawa, Hideaki Ogata, Yoshiki Higuchi, Hideo Akutsu

Research output: Contribution to journalArticle

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Abstract

Cytochrome c3 isolated from a sulfate-reducing bacterium, Desulfo vibrio vulgaris Miyazaki F, is a tetraheme protein. Its physiological partner, [NiFe] hydrogenase, catalyzes the reversible oxidoreduction of molecular hydrogen. To elucidate the mechanism of electron transfer between cytochrome c3 and [NiFe] hydrogenase, the transient complex formation by these proteins was investigated by means of NMR. All NH signals of uniformly 15N-labeled ferric cytochrome c3 except N-terminus, Pro, and Gly73 were assigned. 1H-15N HSQC spectra were recorded for 15N-labeled ferric and ferrous cytochrome c3, in the absence and presence of hydrogenase. Chemical shift perturbations were observed in the region around heme 4 in both oxidation states. Additionally, the region between hemes 1 and 3 in ferrous cytochrome c3 was affected in the presence of hydrogenase, suggesting that the mode of interaction is different in each redox state. Heme 3 is probably the electron gate for ferrous cytochrome c3. To investigate the transient complex of cytochrome c3 and hydrogenase in detail, modeling of the complex was performed for the oxidized proteins using a docking program, ZDOCK 2.3, and NMR data. Furthermore, the roles of lysine residues of cytochrome c3 in the interaction with hydrogenase were investigated by site-directed mutagenesis. When the lysine residues around heme 4 were replaced by an uncharged residue, methionine, one by one, the Km of the electron-transfer kinetics increased. The results showed that the positive charges of Lys60, Lys72, Lys95, and Lys101 around heme 4 are important for formation of the transient complex with [NiFe] hydrogenase in the initial stage of the cytochrome c3 reduction. This finding is consistent with the most possible structure of the transient complex obtained by modeling.

Original languageEnglish
Pages (from-to)1653-1662
Number of pages10
JournalBiochemistry
Volume45
Issue number6
DOIs
Publication statusPublished - 14-02-2006

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Desulfovibrio vulgaris
Oxidation-Reduction
Heme
Hydrogenase
Electrons
Lysine
Nuclear magnetic resonance
Proteins
Vibrio
Mutagenesis
Site-Directed Mutagenesis
cytochrome c(3)
nickel-iron hydrogenase
Chemical shift
Methionine
Sulfates
Hydrogen
Bacteria
Oxidation

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Yahata, N., Saitoh, T., Takayama, Y., Ozawa, K., Ogata, H., Higuchi, Y., & Akutsu, H. (2006). Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F. Biochemistry, 45(6), 1653-1662. https://doi.org/10.1021/bi0514360
Yahata, Naoki ; Saitoh, Takashi ; Takayama, Yuki ; Ozawa, Kiyoshi ; Ogata, Hideaki ; Higuchi, Yoshiki ; Akutsu, Hideo. / Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F. In: Biochemistry. 2006 ; Vol. 45, No. 6. pp. 1653-1662.
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abstract = "Cytochrome c3 isolated from a sulfate-reducing bacterium, Desulfo vibrio vulgaris Miyazaki F, is a tetraheme protein. Its physiological partner, [NiFe] hydrogenase, catalyzes the reversible oxidoreduction of molecular hydrogen. To elucidate the mechanism of electron transfer between cytochrome c3 and [NiFe] hydrogenase, the transient complex formation by these proteins was investigated by means of NMR. All NH signals of uniformly 15N-labeled ferric cytochrome c3 except N-terminus, Pro, and Gly73 were assigned. 1H-15N HSQC spectra were recorded for 15N-labeled ferric and ferrous cytochrome c3, in the absence and presence of hydrogenase. Chemical shift perturbations were observed in the region around heme 4 in both oxidation states. Additionally, the region between hemes 1 and 3 in ferrous cytochrome c3 was affected in the presence of hydrogenase, suggesting that the mode of interaction is different in each redox state. Heme 3 is probably the electron gate for ferrous cytochrome c3. To investigate the transient complex of cytochrome c3 and hydrogenase in detail, modeling of the complex was performed for the oxidized proteins using a docking program, ZDOCK 2.3, and NMR data. Furthermore, the roles of lysine residues of cytochrome c3 in the interaction with hydrogenase were investigated by site-directed mutagenesis. When the lysine residues around heme 4 were replaced by an uncharged residue, methionine, one by one, the Km of the electron-transfer kinetics increased. The results showed that the positive charges of Lys60, Lys72, Lys95, and Lys101 around heme 4 are important for formation of the transient complex with [NiFe] hydrogenase in the initial stage of the cytochrome c3 reduction. This finding is consistent with the most possible structure of the transient complex obtained by modeling.",
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Yahata, N, Saitoh, T, Takayama, Y, Ozawa, K, Ogata, H, Higuchi, Y & Akutsu, H 2006, 'Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F', Biochemistry, vol. 45, no. 6, pp. 1653-1662. https://doi.org/10.1021/bi0514360

Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F. / Yahata, Naoki; Saitoh, Takashi; Takayama, Yuki; Ozawa, Kiyoshi; Ogata, Hideaki; Higuchi, Yoshiki; Akutsu, Hideo.

In: Biochemistry, Vol. 45, No. 6, 14.02.2006, p. 1653-1662.

Research output: Contribution to journalArticle

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T1 - Redox interaction of cytochrome c3 with [NiFe] hydrogenase from desulfovibrio vulgaris Miyazaki F

AU - Yahata, Naoki

AU - Saitoh, Takashi

AU - Takayama, Yuki

AU - Ozawa, Kiyoshi

AU - Ogata, Hideaki

AU - Higuchi, Yoshiki

AU - Akutsu, Hideo

PY - 2006/2/14

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N2 - Cytochrome c3 isolated from a sulfate-reducing bacterium, Desulfo vibrio vulgaris Miyazaki F, is a tetraheme protein. Its physiological partner, [NiFe] hydrogenase, catalyzes the reversible oxidoreduction of molecular hydrogen. To elucidate the mechanism of electron transfer between cytochrome c3 and [NiFe] hydrogenase, the transient complex formation by these proteins was investigated by means of NMR. All NH signals of uniformly 15N-labeled ferric cytochrome c3 except N-terminus, Pro, and Gly73 were assigned. 1H-15N HSQC spectra were recorded for 15N-labeled ferric and ferrous cytochrome c3, in the absence and presence of hydrogenase. Chemical shift perturbations were observed in the region around heme 4 in both oxidation states. Additionally, the region between hemes 1 and 3 in ferrous cytochrome c3 was affected in the presence of hydrogenase, suggesting that the mode of interaction is different in each redox state. Heme 3 is probably the electron gate for ferrous cytochrome c3. To investigate the transient complex of cytochrome c3 and hydrogenase in detail, modeling of the complex was performed for the oxidized proteins using a docking program, ZDOCK 2.3, and NMR data. Furthermore, the roles of lysine residues of cytochrome c3 in the interaction with hydrogenase were investigated by site-directed mutagenesis. When the lysine residues around heme 4 were replaced by an uncharged residue, methionine, one by one, the Km of the electron-transfer kinetics increased. The results showed that the positive charges of Lys60, Lys72, Lys95, and Lys101 around heme 4 are important for formation of the transient complex with [NiFe] hydrogenase in the initial stage of the cytochrome c3 reduction. This finding is consistent with the most possible structure of the transient complex obtained by modeling.

AB - Cytochrome c3 isolated from a sulfate-reducing bacterium, Desulfo vibrio vulgaris Miyazaki F, is a tetraheme protein. Its physiological partner, [NiFe] hydrogenase, catalyzes the reversible oxidoreduction of molecular hydrogen. To elucidate the mechanism of electron transfer between cytochrome c3 and [NiFe] hydrogenase, the transient complex formation by these proteins was investigated by means of NMR. All NH signals of uniformly 15N-labeled ferric cytochrome c3 except N-terminus, Pro, and Gly73 were assigned. 1H-15N HSQC spectra were recorded for 15N-labeled ferric and ferrous cytochrome c3, in the absence and presence of hydrogenase. Chemical shift perturbations were observed in the region around heme 4 in both oxidation states. Additionally, the region between hemes 1 and 3 in ferrous cytochrome c3 was affected in the presence of hydrogenase, suggesting that the mode of interaction is different in each redox state. Heme 3 is probably the electron gate for ferrous cytochrome c3. To investigate the transient complex of cytochrome c3 and hydrogenase in detail, modeling of the complex was performed for the oxidized proteins using a docking program, ZDOCK 2.3, and NMR data. Furthermore, the roles of lysine residues of cytochrome c3 in the interaction with hydrogenase were investigated by site-directed mutagenesis. When the lysine residues around heme 4 were replaced by an uncharged residue, methionine, one by one, the Km of the electron-transfer kinetics increased. The results showed that the positive charges of Lys60, Lys72, Lys95, and Lys101 around heme 4 are important for formation of the transient complex with [NiFe] hydrogenase in the initial stage of the cytochrome c3 reduction. This finding is consistent with the most possible structure of the transient complex obtained by modeling.

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