Structural organization of the Ni and (4Fe-4S) centers in the active form of Desulfovibrio gigas hydrogenase. Analysis of the magnetic interactions by electron paramagnetic resonance spectroscopy

Guigliarelli, Bruno; Moré, C.; Fournel, A.; Asso, Marcel; Hatchikian, E.C.; Williams, Ruth; Cammack, Richard; Bertrand, Patrick

doi:10.1021/bi00014a036

Cited by 77 publications

(91 citation statements)

References 45 publications

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“…This signal accounts for up to 0.3 and 0.25 spin per molecule for the WT and MM variant, and at low temperature, it shows the same spin-spin interaction pattern with the proximal [4Fe4S] cluster ( Fig. 2B) (23). In contrast, the Ni-C signal was much smaller in the case of the FI variant (Ͻ2%), which may result from the doublemutation slightly affecting the redox properties of the active site in this mutant.…”

Section: Resultsmentioning

confidence: 77%

Experimental approaches to kinetics of gas diffusion in hydrogenase

Leroux

Dementin

Burlat

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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152

238

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Hydrogenases, which catalyze H2 to H ؉ conversion as part of the bioenergetic metabolism of many microorganisms, are among the metalloenzymes for which a gas-substrate tunnel has been described by using crystallography and molecular dynamics. However, the correlation between protein structure and gas-diffusion kinetics is unexplored. Here, we introduce two quantitative methods for probing the rates of diffusion within hydrogenases. One uses protein film voltammetry to resolve the kinetics of binding and release of the competitive inhibitor CO; the other is based on interpreting the yield in the isotope exchange assay. We study structurally characterized mutants of a NiFe hydrogenase, and we show that two mutations, which significantly narrow the tunnel near the entrance of the catalytic center, decrease the rates of diffusion of CO and H2 toward and from the active site by up to 2 orders of magnitude. This proves the existence of a functional channel, which matches the hydrophobic cavity found in the crystal. However, the changes in diffusion rates do not fully correlate with the obstruction induced by the mutation and deduced from the x-ray structures. Our results demonstrate the necessity of measuring diffusion rates and emphasize the role of side-chain dynamics in determining these.crystallography ͉ structure/function relationships ͉ substrate tunnel ͉ protein film voltammetry ͉ isotope exchange

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Section: Resultsmentioning

confidence: 77%

Experimental approaches to kinetics of gas diffusion in hydrogenase

Leroux

Dementin

Burlat

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

152

238

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“…S2). This dependence argues for spin-spin interactions between these two species with exchange and dipolar contributions (10,15,16). Such magnetic interactions can be observed in cw EPR spectra between paramagnetic centers localized less than 15 Å apart, suggesting that these centers in Hase I are adjacent to each other (17) (Fig.…”

Section: Resultsmentioning

confidence: 88%

“…1). In addition, the splitting patterns of these EPR spectra show that the exchange interaction lies in an intermediate range ðj2Jj ≈ Δgμ B BÞ (15,16). Furthermore, there could be an additional interaction with the paramagnetic [NiFe] center (S ¼ 1∕2), see below.…”

Section: Resultsmentioning

confidence: 89%

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Characterization of a unique [FeS] cluster in the electron transfer chain of the oxygen tolerant [NiFe] hydrogenase from Aquifex aeolicus

Pandelia

Nitschke

Infossi

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

155

198

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Iron-sulfur clusters are versatile electron transfer cofactors, ubiquitous in metalloenzymes such as hydrogenases. In the oxygentolerant Hydrogenase I from Aquifex aeolicus such electron "wires" form a relay to a diheme cytb, an integral part of a respiration pathway for the reduction of O 2 to water. Amino acid sequence comparison with oxygen-sensitive hydrogenases showed conserved binding motifs for three iron-sulfur clusters, the nature and properties of which were unknown so far. Electron paramagnetic resonance spectra exhibited complex signals that disclose interesting features and spin-coupling patterns; by redox titrations three iron-sulfur clusters were identified in their usual redox states, a ½3Fe4S and two ½4Fe4S , but also a unique high-potential (HP) state was found. On the basis of 57 Fe Mössbauer spectroscopy we attribute this HP form to a superoxidized state of the [4Fe4S] center proximal to the [NiFe] site. The unique environment of this cluster, characterized by a surplus cysteine coordination, is able to tune the redox potentials and make it compliant with the ½4Fe4S 3þ state. It is actually the first example of a biological [4Fe4S] center that physiologically switches between 3þ, 2þ, and 1þ oxidation states within a very small potential range. We suggest that the (1 þ ∕2þ) redox couple serves the classical electron transfer reaction, whereas the superoxidation step is associated with a redox switch against oxidative stress.electrochemistry | EPR | iron-sulfur centers | O2-sensitivity H ydrogenases are metalloproteins occurring in the metabolic pathway of a wide variety of microbial organisms and catalyze the reversible oxidation of dihydrogen: H 2 ⇌ 2H þ þ 2e − (1). The growing interest in alternative sources of energy has focused scientific research on understanding and engineering these enzymes for future applications (2). One of the major limitations of hydrogenases, however, is their sensitivity towards oxygen. Recently, the discovery of hydrogenases that retain catalytic activity in oxygenic environments has potentially opened new applications as "green" vanguard catalysts, in particular as electrocatalysts on electrodes for biofuel cells (3,4).Aquifex aeolicus is a hyperthermophilic Knallgas bacterium with optimum growth temperature of 85°C (5). This microorganism harbors three distinct [NiFe] hydrogenases, among which Hase I is located in the aerobic respiration pathway and attached to the membrane via a diheme cytb (6). Hase I consists of two subunits; the large subunit contains the hetero-bimetallic nickel-iron site and the small subunit the electron transfer cofactors, namely iron-sulfur clusters (6). Based on spectroelectrochemical studies, this enzyme exhibits enhanced thermostability and tolerance for inhibitors (e.g., O 2 and CO) (4, 7).Although the structures of O 2 -sensitive hydrogenases are well characterized (8, 9), such information is still lacking for O 2 -tolerant enzymes. The molecular mechanism and structural determinants for this increased oxygen tolerance remain to ...

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“…Using pulsed electron-spin resonance (ESR) [16], spin-spin relaxation or coherence times (denoted T 2 ) were determined in several molecular magnets and metalloproteins with S = 1 2 ground states. In iron-sulfur clusters, for instance, T 2 is several hundreds of nanoseconds [17,18,19], while in the Cr 7 Ni and Cr 7 Mn antiferromagnetic rings, T 2 ≈ 400 ns at 4.5 K, increasing to 3.8 µs for deuterated Cr 7 Ni at 1.8 K [20]; and T 2 = 2.6 µs for an antiferromagnetic iron(III) trimer [21]. Recently, quan-tum oscillations were observed in the molecular magnet V 15 [22].…”

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confidence: 99%

Direct Observation of Quantum Coherence in Single-Molecule Magnets

et al. 2008

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Direct evidence of quantum coherence in a single-molecule magnet in frozen solution is reported with coherence times as long as T2 = 630 ± 30 ns. We can strongly increase the coherence time by modifying the matrix in which the single-molecule magnets are embedded. The electron spins are coupled to the proton nuclear spins of both the molecule itself and interestingly, also to those of the solvent. The clear observation of Rabi oscillations indicates that we can manipulate the spin coherently, an essential prerequisite for performing quantum computations.PACS numbers: 75.30.Gw, 75.50.Xx, The key concept in quantum information processing is that a quantum bit (qubit) may be not just 0 or 1, as in ordinary computer bits, but an arbitrary superposition of 0 and 1. This means that any two-level system, that can be put into a superposition state, is a qubit candidate [1]. The required superposition state is created by electromagnetic radiation pulses with a frequency corresponding to the energy splitting between the two levels ( Fig. 1c). The contribution of each of the two levels to the superposition state has a cyclic dependence on the pulse length, leading to so-called Rabi oscillations [1]. The observation of such oscillations is a proof-of-principle for the viability of performing quantum computations with a particular system. Quantum computers will probably not be realized from single atoms but will most likely utilize solid state devices, such as superconducting junctions, semiconductor structures, or molecular magnets [1,2]. For these large systems the quantum coherence decays fast, which drastically shortens the time available for quantum computation. Molecular magnets have been considered as qubits because they can be easily organized into large-scale ordered arrays by surface self-assembly [? ], and because they possess excited electronic-spin states required for two-qubit gate operations [5,6]. Single-molecule magnets (SMMs) are exchange-coupled clusters with highspin ground states [3]. The Ising-type anisotropy creates an energy barrier toward magnetization relaxation [ Fig. 1(b)], and many fascinating quantum phenomena have been observed in these systems, such as quantum tunnelling of the magnetization and quantum phase interference [3]. The large splitting of the two lowest states of SMMs in zero field (in principle) allows performing coherent spin-manipulations without external magnetic field, which simplifies any practical implementation. SMMs have also been proposed for the implementation of Grover's algorithm [2] allowing numbers between 0 and 2 2S−2 to be stored in a single molecule.The long coherence time is a crucial first step towards successful implementation of SMMs as qubits [1]. Therefore, recent years have seen a great deal of activity in trying to determine the quantum coherence times in SMMs, which was estimated to be of the order of 10 ns [7,8,9,10]. In several cases, energy gaps between superposition states have been reported that are larger than the expected decoherence energy scale [...

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Structural organization of the Ni and (4Fe-4S) centers in the active form of Desulfovibrio gigas hydrogenase. Analysis of the magnetic interactions by electron paramagnetic resonance spectroscopy

Cited by 77 publications

References 45 publications

Experimental approaches to kinetics of gas diffusion in hydrogenase

Experimental approaches to kinetics of gas diffusion in hydrogenase

Characterization of a unique [FeS] cluster in the electron transfer chain of the oxygen tolerant [NiFe] hydrogenase from Aquifex aeolicus

Direct Observation of Quantum Coherence in Single-Molecule Magnets

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