Electron Paramagnetic Resonance (EPR) Studies on Hydrogenase-1 (HYD1) Purified from a Mutant Strain (AP6) ofEscherichia coliEnhanced in HYD1

Dervartanian, Marie; Menon, N; Przybyla, Alan; Peck, Harry D.; Dervartanian, D.V.

doi:10.1006/bbrc.1996.1491

Cited by 5 publications

(1 citation statement)

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“…The rhombic signals characteristic of both Ni-A and Ni-B show identical g values (to two decimal places) for Hyd-1 and Hyd-2 (Ni-A, g x ϭ 2.31, g y ϭ 2.24, and g z ϭ 2.01; Ni-B, g x ϭ 2.31, g y ϭ 2.16, and g z ϭ 2.01). These values are in good agreement with those previously reported for [NiFe]-hydrogenases (6), although we consistently observed relatively more Ni-B than was evident in the data presented by DerVartanian et al (47). The exact ratio of Ni-A to Ni-B varies between preparations: in the spectra shown, the ratio is ϳ1:3 for Hyd-1 and 1:4 for Hyd-2.…”

Section: Cyclic Voltammetry Experiments On Hyd-1 and Hyd-2-bothsupporting

confidence: 82%

How Escherichia coli Is Equipped to Oxidize Hydrogen under Different Redox Conditions

Lukey

Parkin

Roessler

et al. 2010

Journal of Biological Chemistry

215

271

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The enterobacterium Escherichia coli synthesizes two H 2 uptake enzymes, Hyd-1 and Hyd-2. We show using precise electrochemical kinetic measurements that the properties of Hyd-1 and Hyd-2 contrast strikingly, and may be individually optimized to function under distinct environmental conditions. Hyd-2 is well suited for fast and efficient catalysis in more reducing environments, to the extent that in vitro it behaves as a bidirectional hydrogenase. In contrast, Hyd-1 is active for H 2 oxidation under more oxidizing conditions and cannot function in reverse. Importantly, Hyd-1 is O 2 tolerant and can oxidize H 2 in the presence of air, whereas Hyd-2 is ineffective for H 2 oxidation under aerobic conditions. The results have direct relevance for physiological roles of Hyd-1 and Hyd-2, which are expressed in different phases of growth. The properties that we report suggest distinct technological applications of these contrasting enzymes.Hydrogenases catalyze the reversible cleavage of H 2 into protons and electrons, and play an important role in the energy metabolism of a broad range of microorganisms (1). Hydrogenases are classified according to their active site metal ion content, and three phylogenetically distinct classes have so far been identified: di-iron [FeFe]-, nickel-iron [NiFe]-, and mono-iron [Fe]-hydrogenases (1). Nickel-iron hydrogenases are the most abundant of the three types (1), and many members of this class are membrane bound, with the membrane-extrinsic domain consisting of a large subunit containing the active site, and a small subunit accommodating one to three electron-transferring iron-sulfur clusters. The active sites of [NiFe]-hydrogenases contain a nickel atom coordinated by four cysteine-S ligands, two of which bridge to an iron atom that is further coordinated by three unusual diatomic ligands, two cyanides and one carbonyl (2).Hydrogenases are inactivated by O 2

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Section: Cyclic Voltammetry Experiments On Hyd-1 and Hyd-2-bothsupporting

confidence: 82%

How Escherichia coli Is Equipped to Oxidize Hydrogen under Different Redox Conditions

Lukey

Parkin

Roessler

et al. 2010

Journal of Biological Chemistry

215

271

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Maturation of [NiFe]-hydrogenases in Escherichia coli

2007

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Hydrogenases catalyze the reversible oxidation of dihydrogen. Catalysis occurs at bimetallic active sites that contain either nickel and iron or only iron and the nature of these active sites forms the basis of categorizing the enzymes into three classes, the [NiFe]-hydrogenases, the [FeFe]-hydrogenases and the iron sulfur cluster-free [Fe]-hydrogenases. The [NiFe]-hydrogenases and the [FeFe]-hydrogenases are unrelated at the amino acid sequence level but the active sites share the unusual feature of having diatomic ligands associated with the Fe atoms in the these enzymes. Combined structural and spectroscopic studies of [NiFe]-hydrogenases identified these diatomic ligands as CN- and CO groups. Major advances in our understanding of the biosynthesis of these ligands have been achieved primarily through the study of the membrane-associated [NiFe]-hydrogenases of Escherichia coli. A complex biosynthetic machinery is involved in synthesis and attachment of these ligands to the iron atom, insertion of the Fe(CN)2CO group into the apo-hydrogenase, introduction of the nickel atom into the pre-formed active site and ensuring that the holoenzyme is correctly folded prior to delivery to the membrane. Although much remains to be uncovered regarding each of the individual biochemical steps on the pathway to synthesis of a fully functional enzyme, our understanding of the initial steps in CN- synthesis have revealed that it is generated from carbamoyl phosphate. What is becoming increasingly clear is that the metabolic origins of the carbonyl group may be different.

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Structural differences of oxidized iron–sulfur and nickel–iron cofactors in O 2 -tolerant and O 2 -sensitive hydrogenases studied by X-ray absorption spectroscopy

Sigfridsson

Leidel

Sanganas

et al. 2015

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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The class of [NiFe]-hydrogenases comprises oxygen-sensitive periplasmic (PH) and oxygen-tolerant membrane-bound (MBH) enzymes. For three PHs and four MBHs from six bacterial species, structural features of the nickel-iron active site of hydrogen turnover and of the iron-sulfur clusters functioning in electron transfer were determined using X-ray absorption spectroscopy (XAS). Fe-XAS indicated surplus oxidized iron and a lower number of ~2.7 Å Fe-Fe distances plus additional shorter and longer distances in the oxidized MBHs compared to the oxidized PHs. This supported a double-oxidized and modified proximal FeS cluster in all MBHs with an apparent trimer-plus-monomer arrangement of its four iron atoms, in agreement with crystal data showing a [4Fe3S] cluster instead of a [4Fe4S] cubane as in the PHs. Ni-XAS indicated coordination of the nickel by the thiol group sulfurs of four conserved cysteines and at least one iron-oxygen bond in both MBH and PH proteins. Structural differences of the oxidized inactive [NiFe] cofactor of MBHs in the Ni-B state compared to PHs in the Ni-A state included a ~0.05 Å longer Ni-O bond, a two times larger spread of the Ni-S bond lengths, and a ~0.1 Å shorter Ni-Fe distance. The modified proximal [4Fe3S] cluster, weaker binding of the Ni-Fe bridging oxygen species, and an altered localization of reduced oxygen species at the active site may each contribute to O2 tolerance.

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Electron Paramagnetic Resonance (EPR) Studies on Hydrogenase-1 (HYD1) Purified from a Mutant Strain (AP6) ofEscherichia coliEnhanced in HYD1

Cited by 5 publications

References 0 publications

How Escherichia coli Is Equipped to Oxidize Hydrogen under Different Redox Conditions

How Escherichia coli Is Equipped to Oxidize Hydrogen under Different Redox Conditions

Maturation of [NiFe]-hydrogenases in Escherichia coli

Structural differences of oxidized iron–sulfur and nickel–iron cofactors in O 2 -tolerant and O 2 -sensitive hydrogenases studied by X-ray absorption spectroscopy

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