Spontaneous activation of [FeFe]-hydrogenases by an inorganic [2Fe] active site mimic

Esselborn, Julian; Lambertz, Camilla; Adamska-Venkatesh, Agnieszka; Simmons, Trevor R.; Berggren, Gustav; Noth, Jens; Siebel, Judith F.; Hemschemeier, Anja; Artero, Vincent; Reijerse, Edward J.; Fontecave, Marc; Lubitz, Wolfgang; Happe, Thomas

doi:10.1038/nchembio.1311

Cited by 332 publications

(510 citation statements)

References 35 publications

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“…CpI and CrHydA1 were prepared as described previously, using the semisynthetic procedure (37). All electrochemical experiments were carried out using a Ivium CompactStat E800 workstation equipped with a frequency response analyzer.…”

Section: Methodsmentioning

confidence: 99%

Frequency and potential dependence of reversible electrocatalytic hydrogen interconversion by [FeFe]-hydrogenases

Pandey

Islam

Happe

et al. 2017

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

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The kinetics of hydrogen oxidation and evolution by [FeFe]hydrogenases have been investigated by electrochemical impedance spectroscopy-resolving factors that determine the exceptional activity of these enzymes, and introducing an unusual and powerful way of analyzing their catalytic electron transport properties. Attached to an electrode, hydrogenases display reversible electrocatalytic behavior close to the 2H + /H 2 potential, making them paradigms for efficiency: the electrocatalytic "exchange" rate (measured around zero driving force) is therefore an unusual parameter with theoretical and practical significance. Experiments were carried out on two [FeFe]-hydrogenases, CrHydA1 from the green alga Chlamydomonas reinhardtii, which contains only the active-site "H cluster," and CpI from the fermentative anaerobe Clostridium pasteurianum, which contains four low-potential FeS clusters that serve as an electron relay in addition to the H cluster. Data analysis yields catalytic exchange rates (at the formal 2H + /H 2 potential, at 0°C) of 157 electrons (78 molecules H 2 ) per second for CpI and 25 electrons (12 molecules H 2 ) per second for CrHydA1. The experiments show how the potential dependence of catalytic electron flow comprises frequency-dependent and frequency-independent terms that reflect the proficiencies of the catalytic site and the electron transfer pathway in each enzyme. The results highlight the "wire-like" behavior of the Fe-S electron relay in CpI and a low reorganization energy for electron transfer on/off the H cluster.hydrogen | electrocatalysis | impedance spectroscopy | hydrogenase | electron transfer

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Section: Methodsmentioning

confidence: 99%

Frequency and potential dependence of reversible electrocatalytic hydrogen interconversion by [FeFe]-hydrogenases

Pandey

Islam

Happe

et al. 2017

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

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“…However, intricate molecular constructions with exogeneous redox active centres attached to the diiron unit that mimic the structure of the full H-cluster suffer from instability and low activity 8 . Full catalytic activity for proton reduction approaching the efficiency of the enzyme has only been achieved in hybrid constructs where the synthetic model (m-(SCH 2 ) 2 NH)[Fe(CO) 2 (CN)] 2 2 À has been inserted into the apo-[FeFe]-H 2 ase protein 9,10 . If the substantive goal of creating robust electrocatalysts in simple to assemble, earth-abundant materials is to be realized, we posit that a broader search for metal/ligand frameworks is needed.…”

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confidence: 99%

Redox active iron nitrosyl units in proton reduction electrocatalysis

et al. 2014

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Base metal, molecular catalysts for the fundamental process of conversion of protons and electrons to dihydrogen, remain a substantial synthetic goal related to a sustainable energy future. Here we report a diiron complex with bridging thiolates in the butterfly shape of the 2Fe2S core of the [FeFe]-hydrogenase active site but with nitrosyl rather than carbonyl or cyanide ligands. This binuclear [(NO)Fe(N 2 S 2 )Fe(NO) 2 ] þ complex maintains structural integrity in two redox levels; it consists of a (N 2 S 2 )Fe(NO) complex (N 2 S 2 ¼ N,N 0 -bis(2-mercaptoethyl)-1,4-diazacycloheptane) that serves as redox active metallodithiolato bidentate ligand to a redox active dinitrosyl iron unit, Fe(NO) 2 . Experimental and theoretical methods demonstrate the accommodation of redox levels in both components of the complex, each involving electronically versatile nitrosyl ligands. An interplay of orbital mixing between the Fe(NO) and Fe(NO) 2 sites and within the iron nitrosyl bonds in each moiety is revealed, accounting for the interactions that facilitate electron uptake, storage and proton reduction.

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“…Consequently, we assign the signal to the formation of [2Fe] pdt ‐HydA1, residing in the paramagnetic H ox ‐like state previously described as [4Fe‐4S] 2+ ‐[Fe I Fe II ] pdt 14, 16. To further support the assignment of the observed EPR signal we generated purified [2Fe] pdt ‐HydA1 ([2Fe] pdt ‐HydA1 pur ) through a modified literature protocol 8, 13, 27. The purified semi‐synthetic protein was oxidized by thionine to generate the H ox ‐like state, and the EPR spectrum of [2Fe] pdt ‐HydA1 pur showed the expected rhombic signal (Figure 2, spectrum d) 14.…”

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confidence: 80%

“…Spin quantification shows a 3–3.5‐fold increase in spin concentration after a 3 h incubation, thus reaching 70–85 % of the intensity expected from the in vitro catalytic assays. The requirement for incubation on an hour time‐scale clearly indicates that the rate of cofactor insertion is significantly slower than that observed under in vitro conditions, in which it occurs within minutes 13, 28. In summary, the EPR data shows that the artificial maturation method allows spontaneous formation of a synthetically modified H‐cluster with good yields, at intracellular concentrations readily observable by X‐band EPR spectroscopy.…”

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confidence: 86%

In Vivo EPR Characterization of Semi‐Synthetic [FeFe] Hydrogenases

et al. 2018

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EPR spectroscopy reveals the formation of two different semi‐synthetic hydrogenases in vivo. [FeFe] hydrogenases are metalloenzymes that catalyze the interconversion of molecular hydrogen and protons. The reaction is catalyzed by the H‐cluster, consisting of a canonical iron–sulfur cluster and an organometallic [2Fe] subsite. It was recently shown that the enzyme can be reconstituted with synthetic cofactors mimicking the composition of the [2Fe] subsite, resulting in semi‐synthetic hydrogenases. Herein, we employ EPR spectroscopy to monitor the formation of two such semi‐synthetic enzymes in whole cells. The study provides the first spectroscopic characterization of semi‐synthetic hydrogenases in vivo, and the observation of two different oxidized states of the H‐cluster under intracellular conditions. Moreover, these findings underscore how synthetic chemistry can be a powerful tool for manipulation and examination of the hydrogenase enzyme under in vivo conditions.

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Spontaneous activation of [FeFe]-hydrogenases by an inorganic [2Fe] active site mimic

Cited by 332 publications

References 35 publications

Frequency and potential dependence of reversible electrocatalytic hydrogen interconversion by [FeFe]-hydrogenases

Frequency and potential dependence of reversible electrocatalytic hydrogen interconversion by [FeFe]-hydrogenases

Redox active iron nitrosyl units in proton reduction electrocatalysis

In Vivo EPR Characterization of Semi‐Synthetic [FeFe] Hydrogenases

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