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The crystal structure of a reduced [NiFeSe] hydrogenase provides an image of the activated catalytic center

Background: [NiFeSe] hydrogenases are metalloenzymes that catalyze the reaction H 2↔2H ++2e -.They are generally heterodimeric, contain three iron–sulfur clusters in their small subunit and a nickel–iron-containing active site in their large subunit that includes a selenocysteine (SeCys) ligand. Res...

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Published in:Structure (London) 1999-05, Vol.7 (5), p.557-566
Main Authors: Garcin, E, Vernede, X, Hatchikian, EC, Volbeda, A, Frey, M, Fontecilla-Camps, JC
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creator Garcin, E
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Hatchikian, EC
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Frey, M
Fontecilla-Camps, JC
description Background: [NiFeSe] hydrogenases are metalloenzymes that catalyze the reaction H 2↔2H ++2e -.They are generally heterodimeric, contain three iron–sulfur clusters in their small subunit and a nickel–iron-containing active site in their large subunit that includes a selenocysteine (SeCys) ligand. Results: We report here the X-ray structure at 2.15 Å resolution of the periplasmic [NiFeSe] hydrogenase from Desulfomicrobium baculatum in its reduced, active form. A comparison of active sites of the oxidized, as-prepared, Desulfovibrio gigas and the reduced D. baculatum hydrogenases shows that in the reduced enzyme the nickel-iron distance is 0.4 Å shorter than in the oxidized enzyme. In addition, the putative oxo ligand, detected in the as-prepared D.gigas enzyme, is absent from the D.baculatum hydrogenase. We also observe higher-than-average temperature factors for both the active site nickel–selenocysteine ligand and the neighboring Glu18 residue, suggesting that both these moieties are involved in proton transfer between the active site and the molecular surface. Other differences between [NiFeSe] and [NiFe] hydrogenases are the presence of a third [4Fe4S] cluster replacing the [3Fe4S] cluster found in the D.gigas enzyme, and a putative iron center that substitutes the magnesium ion that has already been described at the C terminus of the large subunit of two [NiFe] hydrogenases. Conclusions: The heterolytic cleavage of molecular hydrogen seems to be mediated by the nickel center and the selenocysteine residue. Beside modifying the catalytic properties of the enzyme, the selenium ligand might protect the nickel atom from oxidation. We conclude that the putative oxo ligand is a signature of inactive ‘unready’ [NiFe] hydrogenases.
doi_str_mv 10.1016/S0969-2126(99)80072-0
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Results: We report here the X-ray structure at 2.15 Å resolution of the periplasmic [NiFeSe] hydrogenase from Desulfomicrobium baculatum in its reduced, active form. A comparison of active sites of the oxidized, as-prepared, Desulfovibrio gigas and the reduced D. baculatum hydrogenases shows that in the reduced enzyme the nickel-iron distance is 0.4 Å shorter than in the oxidized enzyme. In addition, the putative oxo ligand, detected in the as-prepared D.gigas enzyme, is absent from the D.baculatum hydrogenase. We also observe higher-than-average temperature factors for both the active site nickel–selenocysteine ligand and the neighboring Glu18 residue, suggesting that both these moieties are involved in proton transfer between the active site and the molecular surface. Other differences between [NiFeSe] and [NiFe] hydrogenases are the presence of a third [4Fe4S] cluster replacing the [3Fe4S] cluster found in the D.gigas enzyme, and a putative iron center that substitutes the magnesium ion that has already been described at the C terminus of the large subunit of two [NiFe] hydrogenases. Conclusions: The heterolytic cleavage of molecular hydrogen seems to be mediated by the nickel center and the selenocysteine residue. Beside modifying the catalytic properties of the enzyme, the selenium ligand might protect the nickel atom from oxidation. 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Results: We report here the X-ray structure at 2.15 Å resolution of the periplasmic [NiFeSe] hydrogenase from Desulfomicrobium baculatum in its reduced, active form. A comparison of active sites of the oxidized, as-prepared, Desulfovibrio gigas and the reduced D. baculatum hydrogenases shows that in the reduced enzyme the nickel-iron distance is 0.4 Å shorter than in the oxidized enzyme. In addition, the putative oxo ligand, detected in the as-prepared D.gigas enzyme, is absent from the D.baculatum hydrogenase. We also observe higher-than-average temperature factors for both the active site nickel–selenocysteine ligand and the neighboring Glu18 residue, suggesting that both these moieties are involved in proton transfer between the active site and the molecular surface. Other differences between [NiFeSe] and [NiFe] hydrogenases are the presence of a third [4Fe4S] cluster replacing the [3Fe4S] cluster found in the D.gigas enzyme, and a putative iron center that substitutes the magnesium ion that has already been described at the C terminus of the large subunit of two [NiFe] hydrogenases. Conclusions: The heterolytic cleavage of molecular hydrogen seems to be mediated by the nickel center and the selenocysteine residue. Beside modifying the catalytic properties of the enzyme, the selenium ligand might protect the nickel atom from oxidation. 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Results: We report here the X-ray structure at 2.15 Å resolution of the periplasmic [NiFeSe] hydrogenase from Desulfomicrobium baculatum in its reduced, active form. A comparison of active sites of the oxidized, as-prepared, Desulfovibrio gigas and the reduced D. baculatum hydrogenases shows that in the reduced enzyme the nickel-iron distance is 0.4 Å shorter than in the oxidized enzyme. In addition, the putative oxo ligand, detected in the as-prepared D.gigas enzyme, is absent from the D.baculatum hydrogenase. We also observe higher-than-average temperature factors for both the active site nickel–selenocysteine ligand and the neighboring Glu18 residue, suggesting that both these moieties are involved in proton transfer between the active site and the molecular surface. 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source BACON - Elsevier - GLOBAL_SCIENCEDIRECT-OPENACCESS
subjects Amino Acid Sequence
anaerobic techniques
Catalytic Domain
cryocrystallography
Crystallography, X-Ray
hydrogen activation
Hydrogenase - chemistry
Hydrogenase - metabolism
Models, Molecular
Molecular Sequence Data
nickel–iron–selenium hydrogenase
Protein Conformation
Sequence Homology, Amino Acid
title The crystal structure of a reduced [NiFeSe] hydrogenase provides an image of the activated catalytic center
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