The pH dependence of proton-deuterium exchange, hydrogen production and uptake catalyzed by hydrogenases from sulfate-reducing bacteria

Lespinat, Paul A.; Berlier, Y.; Fauque, Guy; Czechowski, Melvin H.; Dimon, Bernard; Gall, Jean Le

doi:10.1016/s0300-9084(86)81068-9

Cited by 53 publications

(53 citation statements)

References 28 publications

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“…This indicates either that enzyme-bound H-might exchange with H20 as well, or that the observation could be due to a molecular cage effect, by which formed HD reacts again to form DD, before diffusion of HD from the enzyme into the bulk water [117]. A low HD/H 2 ratio, comparable to the value found by Krasna [117] for the D. desulfuricans enzyme, was also reported by Lespinat et al [128] in D2/H20 exchange experiments at pH 7 with the selenium-containing enzyme from D. balculatus. These investigators found a strong pH dependence for the HD/H 2 ratio in the exchange reaction.…”

Section: Reaction With Hydrogen Without Apparent Redox Changessupporting

confidence: 73%

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Nickel hydrogenases: in search of the active site

Albracht

1994

Biochimica et Biophysica Acta (BBA) - Bioenergetics

470

479

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Section: Reaction With Hydrogen Without Apparent Redox Changessupporting

confidence: 73%

“…In order to explain the effects of Se and pH, it was assumed [128,198] that the hydride and proton acceptor sites can independently exchange with the solvent. In that case, the relative amounts of HD and H 2 produced would depend on the relative rates of exchange of the two sites.…”

Section: M+d2+b-~ ~ Md-+ Dbmentioning

confidence: 99%

Nickel hydrogenases: in search of the active site

Albracht

1994

Biochimica et Biophysica Acta (BBA) - Bioenergetics

470

479

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“…4). A strong pH dependence for the uH2/uHD ratio has been reported for the D. gigas and D. baculatus enzymes (Lespinat et al, 1986;Teixeira et al, 1987). Such results support a mechanism of hydrogenase action in which the H-and the H' resulting from the heterolytic splitting of the H, can exchange with the solvent independently and at a different exchange rate (Yagi et al, 1973;Vignais et al, 1982).…”

Section: Discussionmentioning

confidence: 61%

“…However, extended-X-ray-absorption-fine-structure analyzes of the 7: roseopersicincz hydrogenase suggested a mixed-ligand coordination with 2 S-(C1-), and 3 N (0) atoms in the first coordination sphere of Ni (Maroney et al, 1991). Therefore, it appeared of interest to study the H-D-exchange reaction of T. roseopersicina enzyme as a function of pH, similarly to what had been performed with D. gigas hydrogenase (Lespinat et al, 1986), not only to investigate the immediate environment of the metal ion(s) at the active site but also to further understand the catalytic mechanism of H, activation. It is shown here that the hydrogenase from 7: roseopersicina, once reductively activated, becomes irreversibly inhibited by iodoacetamide.…”

mentioning

confidence: 96%

Inhibition by Iodoacetamide and Acetylene of the H‐D‐Exchange Reaction Catalyzed by Thiocapsa Roseopersicina Hydrogenase

Зорин

Dimon

Gagnon

et al. 1996

European Journal of Biochemistry

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The kinetics of H-D isotope exchange catalyzed by the thermostable hydrogenase from Thiocapsa roseopersicina have been studied by analysis of the exchange between D, and H,O. The pH dependence of the exchange reaction was examined between pH 2.5 and pH 11. Over the whole pH range, HD was produced at a higher initial velocity than H,, with a marked optimum at pH 5.5; a second peak in the pH profile was observed at around pH 8.5. The rapid formation of H, with respect to HD in the D,/H,O system is consistent with a heterolytic cleavage of D, into D' and an enzyme hydride that can both exchange with the solvent. The H-D-exchange activity was lower in the H,/D,O system than in the D,/ H,O system. The other reactions catalyzed by the hydrogenase, H, oxidation and H, evolution, are pH dependent; the optimal pH were 9.5 for H, uptake and 4.0 for H, production. Treatment of the active form of hydrogenase by iodoacetamide led to a slow and irreversible inhibition of the H-D exchange. When i~do[l-'~C]acetamide was incubated with hydrogenase, the radioactive labeling of the large subunit was higher for the enzyme activated under H, than for the inactive oxidized form. Cysteine residues were identified as the alkylated derivative by amino acid analysis. Acetylene, which inhibits H-D exchange and abolishes the Ni-C EPR signal, protected the enzyme from irreversible inhibition by iodoacetamide. These data indicate that iodoacetamide can reach the active site of the H,-activated hydrogenase from 7: roseopersicina. This was not found to be the case with the seleno hydrogenase from Desulfovibrio baculatus (now Desulfornicrobiurn baculatus). Cysteine modification by iodoacetamide upon activation of the enzyme concomitant with loss of H-D exchange indicates that reductive activation makes at least one Cys residue of the active site available for alkylation.

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“…The specific hydrogen-uptake activity of the M. voltae F,,,-non-reducing enzyme [9] is 50-to lOO-fold greater than the activities found in ordinary nickel hydrogenases. The presence of selenium in the D. baculatus enzyme apparently considerably changed the HJHD ratio in D2/H20 exchange measurements [10,40]. One of the possibilities therefore is that the R-Se-ligand acts as a base in order to help in the heterolytic cleavage of H,.…”

Section: The Effect Of "Co On the Epr Spectrum Of Nickelmentioning

confidence: 99%

Influence of illumination on the electronic interaction between ⁷⁷Se and nickel in active F₄₂₀‐non‐reducing hydrogenase from Methanococcus voltae

Sorgenfrei¹,

Klein²,

Albracht³

1993

FEBS Letters

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The selenium-containing F ,,-non-reducing hydrogenase from Methanococcus voltae was anaerobically purified. The enzyme as isolated showed an EPR spectrum with g,,_ = 2.21, 2.15 and 2.01. Upon illumination this spectrum disappeared and a new signal with the lowest g value at 2.05 arose. EPR studies were carried out either with the enzyme containing natural selenium or enriched in the nuclear isotope "Se. The hype&e splitting caused by "Se in the 'dark' signal is shown to be highly anisotropic. In contrast the splitting is nearly isotropic after illumination. A new model for the nickel site is proposed to explain these observations.

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The pH dependence of proton-deuterium exchange, hydrogen production and uptake catalyzed by hydrogenases from sulfate-reducing bacteria

Cited by 53 publications

References 28 publications

Nickel hydrogenases: in search of the active site

Nickel hydrogenases: in search of the active site

Inhibition by Iodoacetamide and Acetylene of the H‐D‐Exchange Reaction Catalyzed by Thiocapsa Roseopersicina Hydrogenase

Influence of illumination on the electronic interaction between ⁷⁷Se and nickel in active F₄₂₀‐non‐reducing hydrogenase from Methanococcus voltae

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The pH dependence of proton-deuterium exchange, hydrogen production and uptake catalyzed by hydrogenases from sulfate-reducing bacteria

Cited by 53 publications

References 28 publications

Nickel hydrogenases: in search of the active site

Nickel hydrogenases: in search of the active site

Inhibition by Iodoacetamide and Acetylene of the H‐D‐Exchange Reaction Catalyzed by Thiocapsa Roseopersicina Hydrogenase

Influence of illumination on the electronic interaction between 77Se and nickel in active F420‐non‐reducing hydrogenase from Methanococcus voltae

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Influence of illumination on the electronic interaction between ⁷⁷Se and nickel in active F₄₂₀‐non‐reducing hydrogenase from Methanococcus voltae