103 research outputs found
Models for the Active Site of the [FeFe]-Hydrogenase
In der Natur spielen Enzyme fĂŒr viele Prozesse des Lebens eine entscheidende Rolle. Sie bilden die Basis fĂŒr das Leben auf der Erde. Je nach Wirkungsweise werden Enzyme in unterschiedliche Klassen unterteilt. Eine dieser Klasse bilden die Hydrogenasen, welche das Gleichgewicht zwischen Protonen und Wasserstoff beeinflussen. Besonders in methanogenen (Methan-produzierend), acetogenen (EssigsĂ€ure-produzierend), stickstofffixierenden (Umwandlung von N2 in Ammoniak â biologisches Ăquivalent zum Haber-Bosch-Verfahren), photosynthetischen und sulfatreduzierenden Bakterien sind diese Enzyme anzufinden und verdeutlichen die Rolle des Wasserstoffes als Reduktionsmittel und Energiespeicher. Die Hydrogenasen lassen sich je nach Aufbau ihres aktiven Zentrums in verschiedene Unterarten einteilen: [FeFe]-, [FeNi]- und [Fe]-Hydrogenasen.
Die [FeFe]-Enzyme sind in der Lage das Gleichgewicht (2H+/H2) so zu verschieben, dass Protonen zu Wasserstoff reduziert werden und somit ein effizienter EnergietrĂ€ger zur VerfĂŒgung steht. Obwohl das Enzym isoliert werden konnte, war es bisher, bedingt durch analytische Grenzen, nicht möglich die Struktur des aktiven Zentrums vollstĂ€ndig aufzuklĂ€ren. Aufgrund dieses Tatsache gibt es auch heute noch viele Diskussion ĂŒber den Mechanismus der Wasserstoffkatalyse in diesen Enzymsystemen.
Mit Hilfe verschiedenartiger Modellverbindungen soll diese Frage gelöst und die katalytischen FÀhigkeiten zur Wasserstoffsynthese der industriellen Nutzung zugÀnglich gemacht werden. Besonders Peptid-basierte, Zucker-haltige Komplexe, aber auch Komplexe mit (SCH2)2SiR2 Ligandsystemen standen dabei im Fokus dieser Arbeit. Die elektrochemischen und elektrokatalytischen Eigenschaften dieser Komplexe wurden mit Hilfe der Zyklovoltammetrie untersucht
How Ligand Geometry Affects the Reactivity of Co(II) Cyclam Complexes
Cobalt complexes are extensively studied as bioinspired models for non-heme oxygenases as they facilitate both the stabilization and characterization of metal-oxygen intermediates. As an analog to the well-known Co(cyclam) complex Co{N4} (cyclam=1,4,8,11-tetraazacyclotetradecane), the CoII complex Co{i-N4} with the isomeric isocyclam ligand (isocyclam=1,4,7,11-tetraazacyclotetradecane) was synthesized and characterized. Despite the identical N4 donor set of both complexes, Co{i-N4} enables the 2eâ/2H+ reduction of O2 with a lower overpotential (ηeff of 385â
mV vs. 540â
mV for Co{N4}), albeit with a diminished turnover frequency. Characterization of the intermediates formed upon O2 activation of Co{i-N4} reveals a structurally identified stable ÎŒ-peroxo CoIII dimer as the main product. A superoxo CoIII species is also formed as a minor product, as indicated by EPR spectroscopy. In further reactivity studies, the electrophilicity of these inâ
situ generated CoâO2 species was demonstrated by the oxidation of the OâH bond of TEMPOâH (2,2,6,6-tetramethylpiperidin-1-ol) via a H atom abstraction process. Unlike the known Co(cyclam), Co{i-N4} can be employed in oxygen atom transfer reactions oxidizing triphenylphosphine to the corresponding phosphine oxide highlighting the impact of geometrical modifications of the ligand while preserving the ring size and donor atom set on the reactivity of biomimetic oxygen activating complexes.Deutsche Forschungsgemeinschaft (DFG, German Research Foundation)Fraunhofer Internal ProgramsPeer Reviewe
Insights from 125Te and 57Fe nuclear resonance vibrational spectroscopy: a [4Fe-4Te] cluster from two points of view.
Iron-sulfur clusters are common building blocks for electron transport and active sites of metalloproteins. Their comprehensive investigation is crucial for understanding these enzymes, which play important roles in modern biomimetic catalysis and biotechnology applications. We address this issue by utilizing (Et4N)3[Fe4Te4(SPh)4], a tellurium modified version of a conventional reduced [4Fe-4S]+ cluster, and performed both 57Fe- and 125Te-NRVS to reveal its characteristic vibrational features. Our analysis exposed major differences in the resulting 57Fe spectrum profile as compared to that of the respective [4Fe-4S] cluster, and between the 57Fe and 125Te profiles. DFT calculations are applied to rationalize structural, electronic, vibrational, and redox-dependent properties of the [4Fe-4Te]+ core. We herein highlight the potential of sulfur/tellurium exchange as a method to isolate the iron-only motion in enzymatic systems
The Geometry of the Catalytic Active Site in [FeFe]-hydrogenases is Determined by Hydrogen Bonding and Proton Transfer
[FeFe]-hydrogenases are efficient metalloenzymes that catalyze the oxidation and evolution of molecular hydrogen, H2. They serve as a blueprint for the design of synthetic H2-forming catalysts. [FeFe]-hydrogenases harbor a six-iron cofactor that comprises a [4Fe-4S] cluster and a unique diiron site with cyanide, carbonyl, and hydride ligands. To address the ligand dynamics in catalytic turnover and upon carbon monoxide (CO) inhibition, we replaced the native aminodithiolate group of the diiron site by synthetic dithiolates, inserted into wild-type and amino acid variants of the [FeFe]-hydrogenase HYDA1 from Chlamydomonas reinhardtii. The reactivity with H2 and CO was characterized using in situ and transient infrared spectroscopy, protein crystallography, quantum chemical calculations, and kinetic simulations. All cofactor variants adopted characteristic populations of reduced species in the presence of H2 and showed significant changes in CO inhibition and reactivation kinetics. Differences were attributed to varying interactions between polar ligands and the dithiolate head group and/or the environment of the cofactor (i.e., amino acid residues and water molecules). The presented results show how catalytically relevant intermediates are stabilized by inner-sphere hydrogen bonding suggesting that the role of the aminodithiolate group must not be restricted to proton transfer. These concepts may inspire the design of improved enzymes and biomimetic H2-forming catalysts
Stepwise isotope editing of [FeFe]-hydrogenases exposes cofactor dynamics
The six-iron cofactor of [FeFe]-hydrogenases (H-cluster) is the most efficient
H2-forming catalyst in nature. It comprises a diiron active site with three
carbon monoxide (CO) and two cyanide (CNâ) ligands in the active oxidized
state (Hox) and one additional CO ligand in the inhibited state (Hox-CO). The
diatomic ligands are sensitive reporter groups for structural changes of the
cofactor. Their vibrational dynamics were monitored by real-time attenuated
total reflection Fourier-transform infrared spectroscopy. Combination of 13CO
gas exposure, blue or red light irradiation, and controlled hydration of three
different [FeFe]-hydrogenase proteins produced 8 Hox and 16 Hox-CO species
with all possible isotopic exchange patterns. Extensive density functional
theory calculations revealed the vibrational mode couplings of the carbonyl
ligands and uniquely assigned each infrared spectrum to a specific labeling
pattern. For Hox-CO, agreement between experimental and calculated infrared
frequencies improved by up to one order of magnitude for an apical CNâ at the
distal iron ion of the cofactor as opposed to an apical CO. For Hox, two
equally probable isomers with partially rotated ligands were suggested.
Interconversion between these structures implies dynamic ligand reorientation
at the H-cluster. Our experimental protocol for site-selective 13CO isotope
editing combined with computational species assignment opens new perspectives
for characterization of functional intermediates in the catalytic cycle
A dithiacyclam-coordinated silver(i) polymer with anti-cancer stem cell activity
A cancer stem cell (CSC) active, solution stable, silver(i) polymeric complex bearing a dithiacyclam ligand is reported. The complex displays similar potency towards CSCs to salinomycin in monolayer and three-dimensional cultures. Mechanistic studies suggest CSC death results from cytosol entry, an increase in intracellular reactive oxygen species, and caspase-dependent apoptosis
Bridging hydride at reduced H-cluster species in [FeFe]-hydrogenases revealed by infrared spectroscopy, isotope editing, and quantum chemistry
[FeFe]-Hydrogenases contain a H2-converting cofactor (H-cluster) in which a canonical [4Feâ4S] cluster is linked to a unique diiron site with three carbon monoxide (CO) and two cyanide (CNâ) ligands (e.g., in the oxidized state, Hox). There has been much debate whether reduction and hydrogen binding may result in alternative rotamer structures of the diiron site in a single (Hred) or double (Hsred) reduced H-cluster species. We employed infrared spectro-electrochemistry and site-selective isotope editing to monitor the CO/CNâ stretching vibrations in [FeFe]-hydrogenase HYDA1 from Chlamydomonas reinhardtii. Density functional theory calculations yielded vibrational modes of the diatomic ligands for conceivable H-cluster structures. Correlation analysis of experimental and computational IR spectra has facilitated an assignment of Hred and Hsred to structures with a bridging hydride at the diiron site. Pronounced ligand rotation during ÎŒH binding seems to exclude Hred and Hsred as catalytic intermediates. Only states with a conservative H-cluster geometry featuring a ÎŒCO ligand are likely involved in rapid H2 turnover
Spectroscopical Investigations on the Redox Chemistry of [FeFe]-Hydrogenases in the Presence of Carbon Monoxide
[FeFe]-hydrogenases efficiently catalyzes hydrogen conversion at a unique [4Feâ4S]-[FeFe] cofactor, the so-called H-cluster. The catalytic reaction occurs at the diiron site, while the [4Feâ4S] cluster functions as a redox shuttle. In the oxidized resting state (Hox), the iron ions of the diiron site bind one cyanide (CNâ) and carbon monoxide (CO) ligand each and a third carbonyl can be found in the FeâFe bridging position (”CO). In the presence of exogenous CO, A fourth CO ligand binds at the diiron site to form the oxidized, CO-inhibited H-cluster (Hox-CO). We investigated the reduced, CO-inhibited H-cluster (HredÂŽ-CO) in this work. The stretching vibrations of the diatomic ligands were monitored by attenuated total reflection Fourier-transform infrared spectroscopy (ATR FTIR). Density functional theory (DFT) at the TPSSh/TZVP level was employed to analyze the cofactor geometry, as well as the redox and protonation state of the H-cluster. Selective 13CO isotope editing, spectro-electrochemistry, and correlation analysis of IR data identified a one-electron reduced, protonated [4Feâ4S] cluster and an apical CNâ ligand at the diiron site in HredÂŽ-CO. The reduced, CO-inhibited H-cluster forms independently of the sequence of CO binding and cofactor reduction, which implies that the ligand rearrangement at the diiron site upon CO inhibition is independent of the redox and protonation state of the [4Feâ4S] cluster. The relation of coordination dynamics to cofactor redox and protonation changes in hydrogen conversion catalysis and inhibition is discussed
Accumulating the hydride state in the catalytic cycle of [FeFe]-hydrogenases
H2 turnover at the [FeFe]-hydrogenase cofactor (H-cluster) is assumed to
follow a reversible heterolytic mechanism, first yielding a proton and a
hydrido-species which again is double-oxidized to release another proton.
Three of the four presumed catalytic intermediates (Hox, Hred/Hred and Hsred)
were characterized, using various spectroscopic techniques. However, in
catalytically active enzyme, the state containing the hydrido-species, which
is eponymous for the proposed heterolytic mechanism, has yet only been
speculated about. We use different strategies to trap and spectroscopically
characterize this transient hydride state (Hhyd) for three wild-type
[FeFe]-hydrogenases. Applying a novel set-up for real-time attenuated total-
reflection Fourier-transform infrared spectroscopy, we monitor compositional
changes in the state-specific infrared signatures of [FeFe]-hydrogenases,
varying buffer pH and gas composition. We selectively enrich the equilibrium
concentration of Hhyd, applying Le Chatelierâs principle by simultaneously
increasing substrate and product concentrations (H2/H+). Site-directed
manipulation, targeting either the proton-transfer pathway or the adt ligand,
significantly enhances Hhyd accumulation independent of pH
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