On the active site of hydrogenase from Chromatium vinosum

Albracht, S.P.J.; Albrecht-Ellmer, Krystina; Schmedding, D. J. M.; Slater, E.C.

doi:10.1016/0005-2728(82)90041-x

Cited by 44 publications

(16 citation statements)

References 17 publications

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“…elsdenii only C. pasteurianum hydrogenase is clearly of this type; many other spectra (see e.g. [36] in an increase of the EPR signal to an intensity which corresponds to about 0.25 spin, without an observable change in line shape of the 3 Fe-xS signal. The enzyme thus treated has about 50 % of its original hydrogenase activity.…”

Section: Oxidized Enzymesmentioning

confidence: 85%

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Grande

Dunham

Averill

et al. 1983

European Journal of Biochemistry

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The hydrogenases of DesuEfovibrio vulgaris and Megasphaera elsdenii are compared with respect to some of their physical properties. In addition to Fe the only metal ions that are present in significant amounts are Ni and Cu. From cluster extrusion experiments it follows that the D. vulgaris enzyme contains three 4 Fe-4 S clusters, while M . elsdenii hydrogenase only releases part of its Fe-S clusters. The resting D . vulgarisenzyme shows only a small 3 Fe-xS type of EPR signal (maximum 5 %electronequivalent). This amount can beincreased to approximately25 %by treatment with ferricyanide, with a concomitant large decrease in activity. The M. elsdenii enzyme shows in its oxidized state a normal Hipip (high-potential iron-sulphur protein) type of EPR spectrum. After a reduction/oxidation cycle the D. vulgaris enzyme also shows a weak Hipip type of EPR spectrum. In the reduced state both enzymes show complex spectra. By integration of those spectra it is shown that 1.5 electron equivalents are present. The complex spectra do not arise from nuclear hyperfine interactions but are partially due to electron spin interactions. It is proposed that the spectrum of reduced D. vulgaris hydrogenase consists of a sum of three different ferredoxin-like spectra.

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Section: Oxidized Enzymesmentioning

confidence: 85%

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Grande

Dunham

Averill

et al. 1983

European Journal of Biochemistry

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“…[7,19,42,43] The [21] A comprehensive analysis including simulations of the additional spin couplings can be found in earlier work. [36] The EPR spectra of the C81A, C81S, and C81V variants are displayed in Figure 2.…”

Section: Epr Spectroscopic Characterization Of the Oxidized Mbh Variamentioning

confidence: 99%

Impact of Amino Acid Substitutions near the Catalytic Site on the Spectral Properties of an O₂‐Tolerant Membrane‐Bound [NiFe] Hydrogenase

Saggu¹,

Ludwig²,

Friedrich³

et al. 2010

ChemPhysChem

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[NiFe] hydrogenases are widespread among microorganisms and catalyze the reversible cleavage of molecular hydrogen. However, only a few bacteria, such as Ralstonia eutropha H16 (Re), synthesize [NiFe] hydrogenases that perform H(2) cycling in the presence of O(2). These enzymes are of special interest for biotechnological applications. To gain further insight into the mechanism(s) responsible for the remarkable O(2) tolerance, we employ FTIR and EPR spectroscopy to study mutant variants of the membrane-bound hydrogenase (MBH) of Re-carrying substitutions of a particular cysteine residue in the vicinity of the [NiFe] active site that is characteristic of O(2)-tolerant membrane-bound [NiFe] hydrogenases. We demonstrate that these MBH variants, despite minor changes in the electronic structure and in the interaction behavior with the embedding protein matrix, display all relevant catalytic and noncatalytic states of the wild-type enzyme, as long as they are still located in the cytoplasmic membrane. Notably, in the oxidized Ni(r)-B state and the fully reduced forms, the CO stretching frequency increases with increasing polarity of the respective amino acid residue at the specific position of the cysteine residue. We purified the MBH mutant protein with a cysteine-to-alanine exchange to apparent homogeneity as dimeric enzyme after detergent solubilization from the membrane. This purified version displays increased oxygen sensitivity, which is reflected by detection of the oxygen-inhibited Ni(u)-A state, an irreversible inactive redox state, and the light-induced Ni(a)-L state even at room temperature.

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“…Some time later, inspired by the discovery of 3Fe clusters [59,111], Albracht et al [8] considered the possibility that the redox-induced changes might involve a 3Fe/4Fe cluster conversion. Subsequently it was recognized [9,10] vinosum strain DSM 185 {E~(pH 8.0) = + 150 mV [48].…”

Section: An Unknown Redox Component X: a Special Fe Ion?mentioning

confidence: 99%

Nickel hydrogenases: in search of the active site

Albracht

1994

Biochimica et Biophysica Acta (BBA) - Bioenergetics

470

479

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On the active site of hydrogenase from Chromatium vinosum

Cited by 44 publications

References 17 publications

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Impact of Amino Acid Substitutions near the Catalytic Site on the Spectral Properties of an O₂‐Tolerant Membrane‐Bound [NiFe] Hydrogenase

Nickel hydrogenases: in search of the active site

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On the active site of hydrogenase from Chromatium vinosum

Cited by 44 publications

References 17 publications

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Electron paramagnetic resonance and other properties of hydrogenases isolated from Desulfovibrio vulgaris (strain Hildenborough) and Megasphaera elsdenii

Impact of Amino Acid Substitutions near the Catalytic Site on the Spectral Properties of an O2‐Tolerant Membrane‐Bound [NiFe] Hydrogenase

Nickel hydrogenases: in search of the active site

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Impact of Amino Acid Substitutions near the Catalytic Site on the Spectral Properties of an O₂‐Tolerant Membrane‐Bound [NiFe] Hydrogenase