Search

The extraordinary capability of [NiFe]-hydrogenases to catalyse the reversible interconversion of protons and electrons into dihydrogen (H 2 ) has stimulated numerous experimental and theoretical studies addressing the direct utilization of these enzymes in H 2 production processes. Unfortunately, the introduction of these natural H 2 -catalysts in biotechnological applications is limited by their inhibition under oxidising (aerobic and anaerobic) conditions. With the aim of contributing to overcome this limitation, we studied the oxidative inactivation mechanism of [NiFe]-hydrogenases by performing Density Functional Theory (DFT) calculations on a very large model of their active site in which all the amino acids forming the first and second coordination spheres of the NiFe cluster have been explicitly included. We identified an O 2 molecule and two H 2 O molecules as sources of the two oxygen atoms that are inserted at the active site of the inactive forms of the enzyme (Ni-A and Ni-B) under aerobic and anaerobic conditions, respectively. Furthermore, our results support the experimental evidence that the Ni-A-to-Ni-B ratio strongly depends on the number of reducing equivalents available for the process and on the oxidizing conditions under which the reaction takes place.

[NiFe]-hydrogenases catalyse the relevant H 2  → 2H +  + 2e - reaction. Aerobic oxidation or anaerobic oxidation of this enzyme yields two inactive states called Ni-A and Ni-B. These states differ for the reactivation kinetics which are slower for Ni-A than Ni-B. While there is a general consensus on the structure of Ni-B, the nature of Ni-A is still controversial. Indeed, several crystallographic structures assigned to the Ni-A state have been proposed, which, however, differ for the nature of the bridging ligand and for the presence of modified cysteine residues. The spectroscopic characterization of Ni-A has been of little help due to small differences of calculated spectroscopic parameters, which does not allow to discriminate among the various forms proposed for Ni-A. Here, we report a DFT investigation on the nature of the Ni-A state, based on systematic explorations of conformational and configurational space relying on accurate energy calculations, and on comparisons of theoretical geometries with the X-ray structures currently available. The results presented in this work show that, among all plausible isomers featuring various protonation patterns and oxygenic ligands, the one corresponding to the crystallographic structure recently reported by Volbeda et al. (J Biol Inorg Chem 20:11-22, 19)-featuring a bridging hydroxide ligand and the sulphur atom of Cys64 oxidized to bridging sulfenate-is the most stable. However, isomers with cysteine residues oxidized to terminal sulfenate are very close in energy, and modifications in the network of H-bond with neighbouring residues may alter the stability order of such species.

Proton-coupled electron transfer (PCET) is a fundamental process at the core of oxidation-reduction reactions for energy conversion. The [FeFe]-hydrogenases catalyze the reversible activation of molecular H2 through a unique metallocofactor, the H-cluster, which is finely tuned by the surrounding protein environment to undergo fast PCET transitions. The correlation of electronic and structural transitions at the H-cluster with proton-transfer (PT) steps has not been well-resolved experimentally. Here, we explore how modification of the conserved PT network via a Cys → Ser substitution at position 169 proximal to the H-cluster of Chlamydomonas reinhardtii [FeFe]-hydrogenase (CrHydA1) affects the H-cluster using electron paramagnetic resonance (EPR) and Fourier transform infrared (FTIR) spectroscopy. Despite a substantial decrease in catalytic activity, the EPR and FTIR spectra reveal different H-cluster catalytic states under reducing and oxidizing conditions. Under H2 or sodium dithionite reductive treatments, the EPR spectra show signals that are consistent with a reduced [4Fe-4S]H(+) subcluster. The FTIR spectra showed upshifts of νCO modes to energies that are consistent with an increase in oxidation state of the [2Fe]H subcluster, which was corroborated by DFT analysis. In contrast to the case for wild-type CrHydA1, spectra associated with Hred and Hsred states are less populated in the Cys → Ser variant, demonstrating that the exchange of -SH with -OH alters how the H-cluster equilibrates among different reduced states of the catalytic cycle under steady-state conditions.

Nature is a valuable source of inspiration in the design of catalysts, and various approaches are used to elucidate the mechanism of hydrogenases, the enzymes that oxidize or produce H2. In FeFe hydrogenases, H2 oxidation occurs at the H-cluster, and catalysis involves H2 binding on the vacant coordination site of an iron centre. Here, we show that the reversible oxidative inactivation of this enzyme results from the binding of H2 to coordination positions that are normally blocked by intrinsic CO ligands. This flexibility of the coordination sphere around the reactive iron centre confers on the enzyme the ability to avoid harmful reactions under oxidizing conditions, including exposure to O2. The versatile chemistry of the diiron cluster in the natural system might inspire the design of novel synthetic catalysts for H2 oxidation.

Rotated geometry: The first example of a dinuclear iron(I)-iron(I) complex featuring a fully rotated geometry related to the active site of [Fe-Fe] hydrogenase is reported., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

[FeFe]- and [NiFe]-hydrogenases are dihydrogen-evolving metalloenzymes that share striking structural and functional similarities, despite being phylogenetically unrelated. Most notably, they are able to combine substrate binding and redox functionalities, which has important bearings on their efficiency. Model complexes of [FeFe]-hydrogenases that are able to couple H2 binding with a substrate-dependent intramolecular electron transfer promoting dihydrogen activation were recently shown to reproduce the complex redox chemistry of the all-iron enzyme. Notably, coupling of H2 binding and intramolecular redox events was proposed to have a key role also in [NiFe]-hydrogenases, but this feature is not reproduced in currently available nickel-iron biomimetic compounds. In the present study, we exploit dedicated density functional theory approaches to show that H2 binding and activation on a NiFe core can be favored by the installment of conveniently substituted isocyanoferrocenes, thanks to their ability to undergo intramolecular reduction upon substrate binding. Our results support the concept that a unified view on hydrogenase chemistry is a key element to direct future efforts in the modeling of microbial H2 metabolism.

[FeFe] hydrogenases are H2-evolving enzymes that feature a diiron cluster in their active site (the [2Fe]H cluster). One of the iron atoms has a vacant coordination site that directly interacts with H2, thus favoring its splitting in cooperation with the secondary amine group of a neighboring, flexible azadithiolate ligand. The vacant site is also the primary target of the inhibitor O2. The [2Fe]H cluster can span various redox states. The active-ready form (Hox) attains the Fe(II)Fe(I) state. States more oxidized than Hox were shown to be inactive and/or resistant to O2. In this work, we used density functional theory to evaluate whether azadithiolate-to-iron coordination is involved in oxidative inhibition and protection against O2, a hypothesis supported by recent results on biomimetic compounds. Our study shows that Fe-N(azadithiolate) bond formation is favored for an Fe(II)Fe(II) active-site model which disregards explicit treatment of the surrounding protein matrix, in line with the case of the corresponding Fe(II)Fe(II) synthetic system. However, the study of density functional theory models with explicit inclusion of the amino acid environment around the [2Fe]H cluster indicates that the protein matrix prevents the formation of such a bond. Our results suggest that mechanisms other than the binding of the azadithiolate nitrogen protect the active site from oxygen in the so-called H ox (inact) state.

[FeFe]-hydrogenases are dihydrogen-evolving metalloenzymes that are able to combine substrate binding and redox functionalities, a feature that has important bearing on their efficiency. New-generation bioinspired systems such as Fe(2)[(SCH(2))(2)NBn](CO)(3)(Cp*Fe(C(5)Me(4)CH(2)PEt(2)))(dppv) were shown to mimic H(2) oxidation and splitting processes performed by the [FeFe]-hydrogenase/ferredoxin system, and key mechanistic aspects of such reaction are theoretically investigated in the present contribution. We found that H(2) binding and heterolytic cleavage take place concomitantly on DFT models of the synthetic catalyst, due to a substrate-dependent intramolecular redox process that promotes dihydrogen activation. Therefore, formation of an iron-dihydrogen complex as a reaction intermediate is excluded in the biomimetic system, at variance with the case of the enzyme. H(2) uptake at the synthetic system also requires an energetically disfavored isomerization of the amine group acting as a base during splitting. A possible strategy to stabilize the conformation competent for H(2) binding is proposed, along with an analysis of the reactivity of a triiron complex in which di(thiomethyl)amine--the chelating group naturally occurring in [FeFe]-hydrogenases--substitutes the benzyl-containing dithiolate ligand.

A QM/MM investigation of the active-ready (H(ox)) form of [FeFe]-hydrogenase from D. desulfuricans, in which the electronic properties of all Fe-S clusters (H, F and F') have been simultaneously described using DFT, was carried out with the aim of disclosing a possible interplay between the H-cluster and the accessory iron-sulfur clusters in the initial steps of the catalytic process leading to H(2) formation. It turned out that one-electron addition to the active-ready form leads to reduction of the F'-cluster and not of the H-cluster. Protonation of the H-cluster in H(ox) is unlikely, and in any case it would not trigger electron transfer from the accessory Fe(4)S(4) clusters to the active site. Instead, one-electron reduction and protonation of the active-ready form trigger electron transfer within the protein, a key event in the catalytic cycle. In particular, protonation of the H-cluster after one-electron reduction of the enzyme lowers the energy of the lowest unoccupied molecular orbitals localized on the H-cluster to such an extent that a long-range electron transfer from the F'-cluster towards the H-cluster itself is allowed., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Key stereoelectronic properties of Desulfovibrio desulfuricans [FeFe]-hydrogenase (DdH) were investigated by quantum mechanical description of its complete inorganic core, which includes a Fe(6)S(6) active site (the H-cluster), as well as two ancillary Fe(4)S(4) assemblies (the F and F' clusters). The partially oxidized, active-ready form of DdH is able to efficiently bind dihydrogen, thus starting H(2) oxidation catalysis. The calculations allow us to unambiguously assign a mixed Fe(II)Fe(I) state to the catalytic core of the active-ready enzyme and show that H(2) uptake exerts subtle, yet crucial influences on the redox properties of DdH. In fact, H(2) binding can promote electron transfer from the H-cluster to the solvent-exposed F'-cluster, thanks to a 50% decrease of the energy gap between the HOMO (that is localized on the H-cluster) and the LUMO (which is centered on the F'-cluster). Our results also indicate that the binding of the redox partners of DdH in proximity of its F'-cluster can trigger one-electron oxidation of the H(2)-bound enzyme, a process that is expected to have an important role in H(2) activation. Our findings are analyzed not only from a mechanistic perspective, but also in consideration of the physiological role of DdH. In fact, this enzyme is known to be able to catalyze both the oxidation and the evolution of H(2), depending on the cellular metabolic requirements. Hints for the design of targeted mutations that could lead to the enhancement of the oxidizing properties of DdH are proposed and discussed.

Recent advances aimed at modeling the chemistry of the active site of [FeFe]-hydrogenases (the H-cluster, composed by a catalytic Fe(2)S(2) subcluster and an Fe(4)S(4) portion) have led to the synthesis of binuclear coordination compounds containing a noninnocent organophosphine ligand [2,3-bis(diphenylphosphino)maleic anhydride, bma] that is able to undergo monoelectron reduction, analogously to the tetranuclear Fe(4)S(4) subcluster portion of the H-cluster. However, such a synthetic model was shown to feature negligible electronic communication between the noninnocent ligand and the remaining portion of the cluster, at variance with the enzyme active site. Here, we report a theoretical investigation that shows why the electron transfer observed in the enzyme upon protonation of the catalytic Fe(2)S(2) subsite cannot take place in the bma-containing cluster. In addition, we show that targeted modifications of the bma ligand are sufficient to restore the electronic communication within the model, such that electron density can be more easily withdrawn from the noninnocent ligand, as a result of protonation of the iron centers. Similar results were also obtained with a ligand derived from cobaltocene. The relevance of our findings is discussed from the perspective of biomimetic reproduction of proton reduction to yield molecular hydrogen., (© 2011 American Chemical Society)

In this work, we employ density functional theory to assign vibrational signatures of [FeFe]-hydrogenase intermediates to molecular structures. For this purpose, we perform an exhaustive analysis of structures and harmonic vibrations of a series of CN and CO containing model clusters of the [FeFe]-hydrogenase enzyme active site considering also different charges, counterions, and solvents. The pure density functional BP86 in combination with a triple-ζ polarized basis set produce reliable molecular structures as well as harmonic vibrations. Calculated CN and CO stretching vibrations are analyzed separately. Scaled vibrational frequencies are then applied to assign intermediates in [FeFe]-hydrogenase's reaction cycle. The results nicely complement the previous studies of Darensbourg and Hall, and Zilberman et al. The infrared spectrum of the H(ox) form is in very good agreement with the calculated spectrum of the Fe(I)Fe(II) model complex featuring a free coordination site at the distal Fe atom, as well as, with the calculated spectra of the complexes in which H(2) or H(2)O are coordinated at this site. The spectrum of H(red) measured from Desulfovibrio desulfuricans is compatible with a mixture of a Fe(I)Fe(I) species with all terminal COs, and a Fe(I)Fe(I) species with protonated dtma ligand, while the spectrum of H(red) recently measured from Chlamydomonas reinhardtii is compatible with a mixture of a Fe(I)Fe(I) species with a bridged CO, and a Fe(II)Fe(II) species with a terminal hydride bound to the Fe atom., (© 2011 American Chemical Society)

Carbon monoxide is often described as a competitive inhibitor of FeFe hydrogenases, and it is used for probing H(2) binding to synthetic or in silico models of the active site H-cluster. Yet it does not always behave as a simple inhibitor. Using an original approach which combines accurate electrochemical measurements and theoretical calculations, we elucidate the mechanism by which, under certain conditions, CO binding can cause permanent damage to the H-cluster. Like in the case of oxygen inhibition, the reaction with CO engages the entire H-cluster, rather than only the Fe(2) subsite.

The presence of Fe-bound cyanide ligands in the active site of the proton-reducing enzymes [FeFe]-hydrogenases has led to the hypothesis that such Brønsted-Lowry bases could be protonated during the catalytic cycle, thus implying that hydrogen isocyanide (HNC) might have a relevant role in such crucial microbial metabolic paths. We present a hybrid quantum mechanical/molecular mechanical (QM/MM) study of the energetics of CN(-) protonation in the enzyme, and of the effects that cyanide protonation can have on [FeFe]-hydrogenase active sites. A detailed analysis of the electronic properties of the models and of the energy profile associated with H(2) evolution clearly shows that such protonation is dysfunctional for the catalytic process. However, the inclusion of the protein matrix surrounding the active site in our QM/MM models allowed us to demonstrate that the amino acid environment was finely selected through evolution, specifically to lower the Brønsted-Lowry basicity of the cyanide ligands. In fact, the conserved hydrogen-bonding network formed by these ligands and the neighboring amino acid residues is able to impede CN(-) protonation, as shown by the fact that the isocyanide forms of [FeFe]-hydrogenases do not correspond to stationary points on the enzyme QM/MM potential-energy surface., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A DFT study of protonation thermodynamics in H(2)-evolving biomimetic catalysts related to [FeFe]-hydrogenases active site is presented here. Taking as a reference system the electrocatalytic dihydrogen evolution mechanism recently proposed for the synthetic assembly [Fe(2)(CO)(4)(kappa(2)-Ph(2)PCH(2)CH(2)PPh(2))(mu-S(CH(2))(3)S)] (a, which is able to release H(2) after having undergone monoelectron reduction steps and three sequential protonation reactions), we show how the reduction of model complexes to oxidation states lower than those observed in [FeFe]-hydrogenases cofactor leads to a protonation regiochemistry that has no counterpart in the enzymatic mechanism of H(2) production. In particular, double protonation of the metal centers turned out to be disfavored in a by up to 12.5 kcal mol(-1) with respect to alternative protonation paths; as for the regiochemistry of triple protonation, the formation of eta(2)-H(2) adducts is disfavored by at least approximately 25 kcal mol(-1). Structural analysis of the theoretical models also revealed that over-reduction of synthetic complexes, though necessary for observing H(2) evolution from the currently available biomimetic electrocatalysts, can generally impair their structural integrity. Possible approaches for the modulation of protonation regiochemistry are then proposed; in particular, it turned out that a targeted use of sigma-donating ligands showing low basicity can favor double protonation of iron centers.

[FeFe]-hydrogenases are highly efficient H(2)-evolving metalloenzymes that include cyanides and carbonyls in the active site. The latter is an Fe(6)S(6) cluster (the so-called H-cluster) that can be subdivided into a binuclear portion carrying the CO and CN(-) groups and a tetranuclear subcluster. The fundamental role of cyanide ligands in increasing the basicity of the H-cluster has been highlighted previously. Here a more subtle but crucial role played by the two CN(-) ligands in the active site of [FeFe]-hydrogenases is disclosed. In fact, QM/MM calculations on all-atom models of the enzyme from Desulfovibrio desulfuricans show that the cyanide groups fine-tune the electronic and redox properties of the active site, affecting both the protonation regiochemistry and electron transfer between the two subclusters of the H-cluster. Despite the crucial role of cyanides in the protein active site, the currently available bioinspired electrocatalysts generally lack CN(-) groups in order to avoid competition between the latter and the catalytic metal centers for proton binding. In this respect, we show that a targeted inclusion of phosphine ligands in hexanuclear biomimetic clusters may restore the electronic and redox features of the wild-type H-cluster.

The active site of the [FeFe] hydrogenases contains two Fe ions bound to one Cys ligand, three CO molecules, two CN(-) ions, and a dithiolate ligand. The nature of the last of these has been much discussed, and it has been suggested that it contains C, N, or O as the bridgehead atom. Most experimental studies indicate a N atom, whereas a recent density functional theory (DFT) study of a crystal structure indicated an O atom. Here, we performed quantum refinement on the same crystal structure with five different models of the dithiolate ligand X(CH(2)S(-))(2), with X = CH(2), NH(2)(+), NH (two conformations), or O; we found that structures with a N bridgehead atom actually provide the best fit to the raw crystallographic data. Quantum refinement is standard crystallographic refinement in which the molecular mechanics force field normally used to supplement the experimental raw data to give a more chemical structure is replaced by more accurate DFT calculations for the active site. Thereby, we obtain structures that are an ideal compromise between DFT and crystallography.

Fe(2)(S(2)C(3)H(6))(CO)(6) (a) is a simple model of the [FeFe] hydrogenase catalytic site. The topology of the potential energy surface (PES) of this complex, of its cationic and anionic species (a(+) and a(-)), and of its lowest triplet state was studied using density functional theory (DFT) with BP86 and B3LYP functionals, while selected low- and high-lying singlet excited states were studied with the time-dependent density functional theory (TDDFT). The global minima of a and a(-) PESs are characterized by an all-terminal CO ligand arrangement, while the two rotated forms are transition states (TS). On the contrary, for the a(+) and lowest triplet state PES, the three forms considered are local minima, and the syn rotated form is the global minimum. The relative stability of the rotated forms and the all-terminal CO form on the a, a(+), and a(-) PESs is discussed in light of the Quantum Theory of Atoms in Molecules (QTAIM) analysis of the electron density. By comparing the Fe-Fe bond features of the three forms for each PES, we found that the global minimum structure is characterized by the shortest Fe-Fe bond distance and highest electron density at the Fe-Fe critical point. This approach gave evidence that in the a rotated forms, the weak Fe-C(mu) interaction between the Fe atom of the unrotated Fe(CO)(3) and the C atom of the semibridged CO is formed to the detriment of the Fe-Fe bond interaction. These results suggest that the stabilization of the rotated forms on the cationic PES might be due to the formation of the weak Fe-C(mu) interaction minimizing the weakening of the Fe-Fe bond. The low-lying and lowest triplet excited-state PES investigated are characterized by the stabilization of the rotated forms over the all-terminal CO ligand arrangement. On the first singlet 1(1)A'' excited-state PES, an Fe(CO)(3) semirotated structure is the lowest-energy stationary point, while the exploration of the 1(1)A' and 2(1)A'' singlet excited PESs evidences the stabilization of the rotated over the all-terminal CO forms. Singlet excited-state optimized geometry results are compared with excited-state nuclear distortions recently obtained from resonance Raman excitation profiles. Finally, the results of the exploration of the 6(1)A' and 9(1)A' high-lying excited PESs are discussed in light of the recent ultraviolet photolysis experiments on a.

Nature's recipe: A theoretical study analyzes how the environment of the [FeFe] hydrogenase's catalytic cofactor affects its chemical properties, particularly the relative stability of complexes with bridging and terminal hydride ligands (see picture; Fe teal, S yellow, C green, N blue, O red, H gray). The results help to elucidate key rules for the design of bioinspired synthetic catalysts for H(2) production.

The molecular and electronic structure of the Fe 6S 6 H-cluster of [FeFe] hydrogenase in relevant redox and protonation states have been investigated by DFT. The calculations have been carried out according to the broken symmetry approach and considering different environmental conditions. The large negative charge of the H-cluster leads, in a vacuum, to structures different from those observed experimentally in the protein. A better agreement with experimental data is observed for solvated complexes, suggesting that the protein environment could buffer the large negative charge of the H-cluster. The comparison of Fe 6S 6 and Fe 2S 2 DFT models shows that the presence of the Fe 4S 4 moiety does not affect appreciably the geometry of the [2Fe] H cluster. In particular, the Fe 4S 4 cluster alone cannot be invoked to explain the stabilization of the mu-CO forms observed in the enzyme (relative to all-terminal CO species). As for protonation of the hydrogen cluster, it turned out that mu-H species are always more stable than terminal hydride isomers, leading to the conclusion that specific interactions of the H-cluster with the environment, not considered in our calculations, would be necessary to reverse the stability order of mu-H and terminal hydrides. Otherwise, protonation of the metal center and H 2 evolution in the enzyme are predicted to be kinetically controlled processes. Finally, subtle modifications in the H-cluster environment can change the relative stability of key frontier orbitals, triggering electron transfer between the Fe 4S 4 and the Fe 2S 2 moieties forming the H-cluster.

[FeFe]-hydrogenases harbor a {2Fe3S} assembly bearing two CO and two CN- groups, a mu-CO ligand, and a vacant coordination site trans to the mu-CO group. Recent theoretical results obtained studying the isolated {2Fe3S} subsite indicated that one of the CN- ligands can easily move from the crystallographic position to the coordination site trans to the mu-CO group; such an isomerization would have a major impact on substrates and inhibitors binding regiochemistry and, consequently, on the catalytic mechanism. To shed light on this crucial issue, we have carried out hybrid QM/MM and free energy perturbation calculations on the whole enzyme, which demonstrate that the protein environment plays a crucial role and maintains the CN- group fixed in the position observed in the crystal structure; these results strongly support the hypothesis that the vacant coordination site trans to the mu-CO group has a crucial functional relevance both in the context of CO-mediated inhibition of the enzyme and in dihydrogen oxidation/evolution catalysis.

Fe-only hydrogenases are enzymes that catalyze dihydrogen production or oxidation, due to the presence of an unusual Fe(6)S(6) cluster (the so-called H-cluster) in their active site, which is composed of a Fe(2)S(2) subsite, directly involved in catalysis, and a classical Fe(4)S(4) cubane cluster. Here, we present a hybrid quantum mechanical and molecular mechanical (QM/MM) investigation of the Fe-only hydrogenase from Desulfovibrio desulfuricans, in order to unravel key issues regarding the activation of the enzyme from its completely oxidized inactive state (Hoxinact) and the influence of the protein environment on the structural and catalytic properties of the H-cluster. Our results show that the Fe(2)S(2) subcluster in the Fe(II)Fe(II) redox state - which is experimentally observed for the completely oxidized form of the enzyme - binds a water molecule to one of its metal centers. The computed QM/MM energy values for water binding to the diferrous subsite are in fact over 70 kJ mol(-1); however, the affinity toward water decreases by 1 order of magnitude after a one-electron reduction of H(ox)(inact), thus leading to the release of coordinated water from the H-cluster. The investigation of a catalytic cycle of the Fe-only hydrogenase that implies formation of a terminal hydride ion and a di(thiomethyl)amine (DTMA) molecule acting as an acid/base catalyst indicates that all steps have reasonable reaction energies and that the influence of the protein on the thermodynamic profile of H(2) production catalysis is not negligible. QM/MM results show that the interactions between the Fe(2)S(2) subsite and the protein environment could give place to structural rearrangements of the H-cluster functional for catalysis, provided that the bidentate ligand that bridges the iron atoms in the binuclear subsite is actually a DTMA residue.

The di-iron complex Fe2(S2C3H6)(CO)6 (a), one of the simplest functional models of the Fe-hydrogenases active site, is able to electrocatalyze proton reduction. In the present study, the H2 evolving path catalyzed by a has been characterized using density functional theory. It is showed that, in the early stages of the catalytic cycle, a neutral mu-H adduct is formed; monoelectron reduction and subsequent protonation can give rise to a diprotonated neutral species (a-muH-SH), which is characterized by a mu-H group, a protonated sulfur atom, and a CO group bridging the two iron centers, in agreement with experimental IR data indicating the formation of a long-lived mu7-CO species. H2 release from a-muH-SH, and its less stable isomer a-H2 is kinetically unfavorable, while the corresponding monoanionic compounds (a-muH-SH- and a-H2-) are more reactive in terms of dihydrogen evolution, in agreement with experimental data. The key species involved in electrocatalysis have structural features different from the hypothetical intermediates recently proposed to be involved in the enzymatic process, an observation that is possibly correlated with the reduced catalytic efficiency of the biomimetic di-iron assembly.

The MoCu CO dehydrogenase enzyme not only transforms CO into CO2 but it can also oxidise H2. Even if its hydrogenase activity has been known for decades, a debate is ongoing on the most plausible mode for the binding of H2 to the enzyme active site and the hydrogen oxidation mechanism. In the present work, we provide a new perspective on the MoCu-CODH hydrogenase activity by improving the in silico description of the enzyme. Energy refinement—by means of the BigQM approach—was performed on the intermediates involved in the dihydrogen oxidation catalysis reported in our previously published work (Rovaletti, et al. "Theoretical Insights into the Aerobic Hydrogenase Activity of Molybdenum–Copper CO Dehydrogenase." Inorganics 7 (2019) 135). A suboptimal description of the H2–HN(backbone) interaction was observed when the van der Waals parameters described in previous literature for H2 were employed. Therefore, a new set of van der Waals parameters is developed here in order to better describe the hydrogen–backbone interaction. They give rise to improved binding modes of H2 in the active site of MoCu CO dehydrogenase. Implications of the resulting outcomes for a better understanding of hydrogen oxidation catalysis mechanisms are proposed and discussed. [ABSTRACT FROM AUTHOR]

Catalytic H2 oxidation has been dissected by means of DFT into the key steps common to the Fe2 unit of both the [FeFe]‐hydrogenase cofactor and selected biomimics. The aim was to elucidate the molecular details underlying the very different performances of the two systems. We found that the better enzyme performance is based on a single iron atom that is maintained electron‐poor, favoring H2 binding, although embedded within a highly electron‐rich cofactor, ensuring a facile oxidation of the Fe2–H2 adduct. This is due to 1) CN− coordinating to both iron atoms, due to their amphipathic Lewis acid/base properties, and 2) the 4Fe4S subunit further withdrawing electrons from the Fe2 core. Preserving a moderate electron deficiency at a single iron also helps the cofactor preserve hydride affinity, which favors H2 cleavage. Such valuable characteristics allow the biocatalyst to turnover close to equilibrium conditions. All previous biomimicry has shown, in contrast, the impossibility to properly balance the two apparently contrasting aforementioned requisites, although evident progress has been made by the H2‐ase community. Disclosure of the differences identified could inspire the design of novel biomimics, for instance, reconsidering the use of CN− in the catalyst architecture. Indeed, in the presence of bases normally employed in oxidative catalysis, undesired stable protonation at coordinated CN−, which affects the opposite process (proton reduction), could be overcome. Nature versus biomimicry: Dissecting H2 oxidation by the [FeFe]‐H2ase core in comparison with a series of Fe2S2 models has revealed that CN− ligands and the 4Fe4S subunit are the key to the higher catalytic performance of the enzyme compared with the synthetic compounds. Indeed, cyanides and cubane provide a single electron‐poor iron (which assists H2 binding) embedded within an electron‐rich environment, which in turn ensures a facile subsequent oxidation. [ABSTRACT FROM AUTHOR]

The present paper reports a study on the energetics of protonation of a hydrogenase biomimetic complex, [Fe2(μ-adt)(CO)4(PMe3)2] (adt = N-benzylazadithiolate), and of its homologue featuring triphenylphosphine ligands in place of trimethylphosphines. Formation of a terminal hydride on one of the Fe centres was considered first, given the key relevance of terminal hydride species in the enzymatic mechanism. Theoretical calculations highlight that, in a vacuum, terminal protonation of the selected Fe ion in the PPh3-bearing organometallic complex is highly favoured when compared to the analogous reaction involving the PMe3-containing species, but the trend is inverted in the case of models optimized in a continuum polarizable. An unexpected parallel is thus established between relative basicities of PPh3 and PMe3 in vacuum or in solution phase [C. A. Tolman, J. Am. Chem. Soc. 1970, 92, 2953; G. M. Bancroft, Inorg. Chem. 1986, 25, 3675], and the energetics of terminal hydride formation upon protonation of [FeFe]-hydrogenase biomimetic complexes bearing such organophosphorous ligands. Bridging hydride formation was also considered in the present study: calculations showed that protonation of the PMe3-bearing organometallic complex is again strongly favoured in vacuo, as compared to the case of the PPh3-containing model. However, protonation energies become significantly smaller when solvent effects are taken into account. Such differences between protonation reactions modelled in vacuo and in the polarizable continuum are rationalized in light of the different electrostatic properties of the diiron complexes here considered. Implications for the design and modelling of biomimetic catalysts are briefly discussed in light of recent literature. [ABSTRACT FROM AUTHOR]

The Mo/Cu-dependent CO dehydrogenase from Oligotropha carboxidovorans is an enzyme that is able to catalyze CO oxidation to CO2; moreover, it can also oxidize H2, thus eliciting a characteristic EPR signal. Interestingly, the Ag-substituted enzyme form proved unable to catalyze H2 oxidation. In the present contribution, we characterized the reactivity of the enzyme with H2 by quantumchemical calculations. It was found that dihydrogen binding to the wild-type enzyme requires significant structural rearrangements of the active site Theoretical EPR spectra for plausible H2-bound models of the partially reduced, paramagnetic active site are also presented and compared with the experimental counterpart. Finally, density functional theory modeling shows that Ag substitution impairs H2 binding at the active site. [ABSTRACT FROM AUTHOR]

Upon irradiation with ultraviolet wavelengths, Fe2(S2C3H6)(CO)6, a simple model of the [FeFe]-hydrogenase active site, undergoes CO dissociation to form the unsaturated Fe2(S2C3H6)(CO)5 species and successively a solvent adduct at the vacant coordination site. In the present work, the CO-photolysis of Fe2(S2C3H6)(CO)6 was investigated by density functional theory (DFT) and time-dependent DFT (TDDFT). Trans Fe2(S2C3H6)(CO)5 form and the corresponding trans heptane or acetonitrile solvent adducts are the lowest energy ground state forms. CO dissociation barriers computed for the lowest triplet state are roughly halved with respect to those for the ground state suggesting that some low-lying excited potential energy surface (PES) could be loosely bound with respect to FeC bond cleavage. The TDDFT excited state PESs and geometry optimizations for the excited states likely involved in the CO-photolysis suggest that the FeS bond elongation and the partial isomerization toward the rotated form could take place simultaneously, favoring the trans CO photodissociation. © 2014 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

[FeFe]- and [NiFe]-hydrogenases are dihydrogen-evolving metalloenzymes that share striking structural and functional similarities, despite being phylogenetically unrelated. Most notably, they are able to combine substrate binding and redox functionalities, which has important bearings on their efficiency. Model complexes of [FeFe]-hydrogenases that are able to couple H2 binding with a substrate-dependent intramolecular electron transfer promoting dihydrogen activation were recently shown to reproduce the complex redox chemistry of the all-iron enzyme. Notably, coupling of H2 binding and intramolecular redox events was proposed to have a key role also in [NiFe]-hydrogenases, but this feature is not reproduced in currently available nickel–iron biomimetic compounds. In the present study, we exploit dedicated density functional theory approaches to show that H2 binding and activation on a NiFe core can be favored by the installment of conveniently substituted isocyanoferrocenes, thanks to their ability to undergo intramolecular reduction upon substrate binding. Our results support the concept that a unified view on hydrogenase chemistry is a key element to direct future efforts in the modeling of microbial H2 metabolism. [ABSTRACT FROM AUTHOR]

[FeFe]-hydrogenases are dihydrogen-evolving met-alloenzymes that are able to combine substrate binding and redox functionalities, a feature that has important bearing on their efficiency. New-generation bioinspired systems such as Fe2[(SCH2)2NBn](CO)3(Cp*Fe(C5Me4CH2PEt2))(dppv) were shown to mimic H2 oxidation and splitting processes performed by the [FeFe]-hydrogenase/ferredoxin system, and key mechanistic aspects of such reaction are theoretically investigated in the present contribution. We found that H2 binding and heterolytic cleavage take place concomitantly on DFT models of the synthetic catalyst, due to a substrate-dependent intramolecular redox process that promotes dihydrogen activation. Therefore, formation of an iron-- dihydrogen complex as a reaction intermediate is excluded in the biomimetic system, at variance with the case of the enzyme. H2 uptake at the synthetic system also requires an energetically disfavored isomerization of the amine group acting as a base during splitting. A possible strategy to stabilize the conformation competent for H2 binding is proposed, along with an analysis of the reactivity of a triiron complex in which di(thiomethyl)amine--the chelating group naturally occurring in [FeFe]-hydrogenases--substitutes the benzyl-containing dithiolate ligand. [ABSTRACT FROM AUTHOR]

In this work, we employ density functional theory to assign vibrational signatures of [FeFe]-hydrogenase intermediates to molecular structures. For this purpose, we perform an exhaustive analysis of structures and harmonic vibrations of a series of CN and CO containing model clusters of the [FeFe]-hydrogenase enzyme active site considering also different charges, counterions, and solvents. The pure density functional BP86 in combination with a triple-ζ polarized basis set produce reliable molecular structures as well as harmonic vibrations. Calculated CN and CO stretching vibrations are analyzed separately. Scaled vibrational frequencies are then applied to assign intermediates in [FeFe]-hydrogenase's reaction cycle. The results nicely complement the previous studies of Darensbourg and Hall,1,2 and Zilberman et al. 3-5 The infrared spectrum of the Hox form is in very good agreement with the calculated spectrum of the FeIFeII model complex featuring a free coordination site at the distal Fe atom, as well as, with the calculated spectra of the complexes in which H2 or H2O are coordinated at this site. The spectrum of Hred measured from Desulfovibrio desulfuricans is compatible with a mixture of a FeIFeI species with all terminal COs, and a FeIFeI species with protonated dtma ligand, while the spectrum of Hred recently measured from Chlamydomonas reinhardtii is compatible with a mixture of a FeIFeI species with a bridged CO, and a FeIIFeII species with a terminal hydride bound to the Fe atom. [ABSTRACT FROM AUTHOR]

Abstract: In this work DFT has been used to characterize model complexes structurally related to the CO-inhibited form (Hox-CO) of [FeFe]-hydrogenases. The investigation of a recently synthesized diiron complex ([Fe2{MeSCH2C(Me)(CH2S)2}(CN)2(CO)4]−, [M. Razavet, S.J. Borg, S.J. George, S.P. Best, S.A. Fairhurst, C.J. Pickett, Chem. Commun. 2002, 700–701]) that closely reproduces most features of the inhibited enzyme cofactor, led to the conclusion that the computation of DFT energy differences, as well as the comparison between computed and experimental IR and EPR spectra, does not allow to confidently distinguish among isomers differing for the position of CO and CN ligands, an issue which is relevant not only to fully understand the mechanism of CO-mediated inhibition of the enzyme, but more generally to further understand the factors affecting substrates coordination to the enzyme active site. The latter observation prompted us to probe the effect of the electronic properties of ligands on the structural features of a series of [Fe2(SCH2 XCH2S)(CN)2(CO)3(L)] n − complexes related to the Hox-CO form of the enzyme but differing for the nature of L (CO, (CH3)2S, CH3S−, CH3O− and F−) and X (CH2, NH and O). Results revealed that the electronic properties of ligands, as well as the nature of the chelating group bridging the two iron atoms, can affect the coordination geometry of the distal metal center. In particular, it turned out that the inclusion of hard ligands in the Fe coordination sphere could be a viable strategy to selectively favour isomers featuring two CO groups trans to each other. On the other hand, the substitution of propanedithiolate with a di(thiomethyl)amine residue led to the selective stabilization of structures featuring a CN ligand in trans to the μ-CO group, thanks to the formation of an intramolecular hydrogen bond. The relevance of these DFT results for the design of novel biomimetic models of the CO-inhibited [FeFe]-hydrogenases active site is discussed. [Copyright &y& Elsevier]

Abstract: In the present contribution, a density functional theory (DFT) investigation is described regarding a recently synthesized Fe6S6 complex – see C. Tard, X. Liu, S.K. Ibrahim, M. Bruschi, L. De Gioia, S.C. Davies, X. Yang, L.-S. Wang, G. Sawers, C.J. Pickett, Nature 433 (2005) 610 – that is structurally and functionally related to the [FeFe]-hydrogenases active site (the so-called H-cluster, which includes a binuclear subsite directly involved in catalysis and an Fe4S4 cubane). The analysis of relative stabilities and atomic charges of different isomers evidenced that the structural and redox properties of the synthetic assembly are significantly different from those of the enzyme active site. A comparison between the hexanuclear cluster and simpler synthetic diiron models is also described; the results of such a comparison indicated that the cubane moiety can favour the stabilization of the cluster in a structure closely resembling the H-cluster geometry when the synthetic Fe6S6 complex is in its dianionic state. However, the opposite effect is observed when the synthetic cluster is in its monoanionic form. [Copyright &y& Elsevier]

The di-iron complex Fe2(S2C3H6)(CO)6 (a), one of the simplest functional models of the Fe-hydrogenases active site, is able to electrocatalyze proton reduction. In the present study, the H2 evolving path catalyzed by a has been characterized using density functional theory. It is showed that, in the early stages of the catalytic cycle, a neutral μ-H adduct is formed; monoelectron reduction and subsequent protonation can give rise to a diprotonated neutral species (a-μH-SH), which is characterized by a μ-H group, a protonated sulfur atom, and a CO group bridging the two iron centers, in agreement with experimental IR data indicating the formation of a long-lived μ-CO species. H2 release from a-μH-SH, and its less stable isomer a-H2 is kinetically unfavorable, while the corresponding monoanionic compounds (a-μH-SH- and a-H2-) are more reactive in terms of dihydrogen evolution, in agreement with experimental data. The key species involved in electrocatalysis have structural features different from the hypothetical intermediates recently proposed to be involved in the enzymatic process, an observation that is possibly correlated with the reduced catalytic efficiency of the biomimetic di-iron assembly. [ABSTRACT FROM AUTHOR]

Density functional theory was used to compare reaction pathways for H2 formation and H+ reduction catalyzed by models of the binuclear cluster found in the active site of [Fe] hydrogenases. Terminal W binding to an FeI-FI form, followed by monoelectron reduction and protonation of the di(thiomethyl)amine ligand, can conveniently lead to H2 formation and release, suggesting that this mechanism could be operative within the enzyme active site. However, a pathway that implies the initial formation of FeII-FeII μ-H species and release of H2 from an FeII-FeI form is characterized by only slightly less favored energy profiles. In both cases, H2 formation becomes less favored when taking into account the competition between CN and amine groups for H+ binding, an observation that can be relevant for the design of novel synthetic catalysts. H2 cleavage can take place on FeII-FeII redox species, in agreement with previous proposals [Fan, H.-J.; Hall, M. B. J. Am. Chem. Soc. 2001, 123, 3828] and, in complexes characterized by terminal CO groups, does not need the involvement of an external base. The step in H2 oxidation characterized by larger energy barriers corresponds to the second W extraction from the cluster, both considering FeII-FeII and FeII-FeIII species. A comparison of the different reaction pathways reveals that H2 formation could involve only FI1-FI, FeII-FeI, and FeII-FeII species, whereas FeIII-FeII species might be relevant in H2 cleavage. [ABSTRACT FROM AUTHOR]

The Mo/Cu-dependent CO dehydrogenase from O. carboxidovorans is an enzyme that is able to catalyse CO oxidation to CO 2 ; moreover, it also expresses hydrogenase activity, as it is able to oxidize H 2 . Here, we have studied the dihydrogen oxidation catalysis by this enzyme using QM/MM calculations. Our results indicate that the equatorial oxo ligand of Mo is the best suited base for catalysis. Moreover, extraction of the first proton from H 2 by means of this basic centre leads to the formation of a Mo–OH–Cu I H hydride that allows for the stabilization of the copper hydride, otherwise known to be very unstable. In light of our results, two mechanisms for the hydrogenase activity of the enzyme are proposed. The first reactive channel depends on protonation of the sulphur atom of a Cu-bound cysteine residues, which appears to favour the binding and activation of the substrate. The second reactive channel involves a frustrated Lewis pair, formed by the equatorial oxo group bound to Mo and by the copper centre. In this case, no binding of the hydrogen molecule to the Cu center is observed but once H 2 enters into the active site, it can be split following a low-energy path. [ABSTRACT FROM AUTHOR]

The article focuses on [FeFe]-hydrogenases, hydrogen-evolving enzymes that are able to catalyze H2 production at very low overpotentials under mild conditions. The components of the active site of [FeFe]-hydrogenases include an Fe6S6 complex, known as the H-cluster, which is further composed of a classical Fe4S4 cubane cluster. Three CO and two CN- ligands and a bidentate chelating ligand coordinate the Fe atoms of the binuclear subcluster.

FeFe hydrogenases catalyze H2 oxidation and formation at an inorganic active site (the "H-cluster"), which consists of a [Fe2(CO)3(CN)2(dithiomethylamine)] subcluster covalently attached to a Fe4S4 subcluster. This active site is photosensitive: visible light has been shown to induce the release of exogenous CO (a reversible inhibitor of the enzyme), shuffle the intrinsic CO ligands, and even destroy the H-cluster. These reactions must be understood because they may negatively impact the use of hydrogenase for the photoproduction of H2. Here, we explore in great detail the reactivity of the excited states of the H-cluster under catalytic conditions by examining, both experimentally and using TDDFT calculations, the simplest photochemical reaction: the binding and release of exogenous CO. A simple dyad model can be used to predict which excitations are active. This strategy could be used for probing other aspects of the photoreactivity of the H-cluster. [ABSTRACT FROM AUTHOR]

FeFe hydrogenases catalyze H2 oxidation and production using an "H-cluster", where two Fe ions are bound by an aza-dithiolate (adt) ligand. Various hypotheses have been proposed (by us and others) to explain that the enzyme reversibly inactivates under oxidizing, anaerobic conditions: intramolecular binding of the N atom of adt, formation of the so-called "Hox/inact" state or nonproductive binding of H2 to isomers of the H-cluster. Here, we show that none of the above explains the new finding that the anaerobic, oxidative, H2-dependent reversible inactivation is strictly dependent on the presence of Cl- or Br-. We provide experimental evidence that chloride uncompetitively inhibits the enzyme: it reversibly binds to catalytic intermediates of H2 oxidation (but not to the resting "Hox" state), after which oxidation locks the active site into a stable, saturated, inactive form, the structure of which is proposed here based on DFT calculations. The halides interact with the amine group of the H-cluster but do not directly bind to iron. It should be possible to stabilize the inhibited state in amounts compatible with spectroscopic investigations to explore further this unexpected reactivity of the H-cluster of hydrogenase. [ABSTRACT FROM AUTHOR]

A novelreaction pathway to synthesize the linear trinuclear clusters[Fe3(CO)5(κ2-diphosphine)(μ-dithiolate)2] via the direct reaction of the dinuclear hexacarbonyl precursor[Fe2(CO)6(μ-dithiolate)] with the mononuclearspecies [Fe(CO)2(κ2-diphosphine)(κ2-dithiolate)] has been developed with diphosphine (dppe) anddithiolate (pdt = propanedithiolate) (1) or adtBn({SCH2}2NBn = azadithiolate) (2). A crystallographic study was carried out on 2andMössbauer spectroscopy, and DFT calculations have been usedto describe the electronic and structural properties of 1. The electrochemical properties of 1in the absenceand in the presence of a weak acid have been the subject of a preliminaryinvestigation. [ABSTRACT FROM AUTHOR]

[FeFe]-hydrogenases are the most efficient biological catalysts available for the H 2 evolution reaction. Their active site – the H-cluster – features a diiron subsite which has the peculiar characteristic of bearing cyanide groups hydrogen-bonded to the apoprotein as well as carbonyl ligands. Notably, one of the CO ligands is disposed in bridging position between the metal centers. This allows one of the Fe ions to retain a square pyramidal coordination – which determines the assumption of the so-called “rotated structure” – with a vacant coordination site in trans to the μ-CO group, ready to bind protons when the active site is in the Fe I Fe I state. Many Fe I Fe I biomimetic models have been synthesized and characterized so far, but most of them fail to reproduce the orientation of the diatomic ligands that is observed in the enzyme active site. In the present contribution we carried out a density functional theory investigation, with the aim of evaluating whether the establishment of hydrogen bonding at the level of cyanides is sufficient to favor rotation of ligands around one of the Fe centers, in analogy with the reduced H-cluster. To this end, we carried out an investigation of the potential energy surface of an isolated Fe 2 S 2 model bearing CN and CO groups, as well as of the supramolecular complex formed by the diiron model and a porphyrin derivative hydrogen-bonded to the former. As far as the isolated Fe 2 S 2 species are concerned, the sole μ-CO models individuated in the course of our potential energy surface scans are the ones in which both cyanides are bound to same iron center, while no CO-bridged minima could be found in the case of models having one CN group bound to each of the metal ions. The latter represents a major difference with respect to the coordination geometry of the reduced diiron subcluster in the enzyme. However, perturbation of the diiron model by a porphyrin ring designed to donate hydrogen bonds to the cyanide groups – thus, at least partially, reproducing the network of H-bond between the H-cluster and the apoprotein in [FeFe]-hydrogenases – significantly changes the picture in this regard. Therefore, possible strategies to modulate the disposition of ligands around the metal centers of biomimetic [FeFe]-hydrogenases models are discussed in light of computed geometries and relative stabilities of the supramolecular complexes. [ABSTRACT FROM AUTHOR]

[Display omitted] • Coordination models of bioinorganic compounds. • Density Functional Theory approaches in bioinorganic chemistry. • Co, Fe and Cu-mediated small molecules activation. • The cultural and research legacy of professor Renato Ugo. A few bio-inorganic systems have been studied by means of modern quantum chemistry (QM) methods, such as the Density Functional Theory (DFT) in connection with a large set of basis functions. At this level, DFT has proven to give reliable (semi-quantitative) description of several properties of transition metals complexes which are either relevant part of a natural enzyme or ad hoc built to mimic the behaviour of enzymes. The first example considered is the N,N'-ethylenebis(acetylacetonatiminato)Co(II) (Coacacen) , a very famous synthetic oxygen carrier. We report results of a DFT investigation of Coacacen in its free form as well as in penta-coordinated form with a suitable axial ligand and, finally, in hexa -coordinated form bound to dioxygen. The problems related to the electronic structure of Coacacen derivatives are very old, and have been investigated (mainly experimentally) for several years. Coacacen has been the first bio-inorganic system investigated in the group of Renato Ugo in the 1970s. In addition, a short review will be presented concerning the "modern" problems of the bio-inorganics, as they are presently investigated in our research group based at the University of Milano-Bicocca (Italy): the catalytic cycle characteristic of Fe-Fe hydrogenases and the model compounds describing the interaction between copper and amyloid β-peptides. [ABSTRACT FROM AUTHOR]