Structural correlations of nitrogenase active sites using nuclear resonance vibrational spectroscopy and QM/MM calculations.

The biological conversion of N2 to NH3 is accomplished by the nitrogenase family, which is collectively comprised of three closely related but unique metalloenzymes. In the present study, we have employed a combination of the synchrotron-based technique of 57Fe nuclear resonance vibrational spectroscopy together with DFT-based quantum mechanics/molecular mechanics (QM/MM) calculations to probe the electronic structure and dynamics of the catalytic components of each of the three unique M N2ase enzymes (M = Mo, V, Fe) in both the presence (holo-) and absence (apo-) of the catalytic FeMco clusters (FeMoco, FeVco and FeFeco). The results described herein provide vibrational mode assignments for important fingerprint regions of the FeMco clusters, and demonstrate the sensitivity of the calculated partial vibrational density of states (PVDOS) to the geometric and electronic structures of these clusters. Furthermore, we discuss the challenges that are faced when employing NRVS to investigate large, multi-component metalloenzymatic systems, and outline the scope and limitations of current state-of-the-art theory in reproducing complex spectra.

Synchrotron Mössbauer source: trade-off between intensity and linewidth.

A synchrotron Mössbauer source (SMS) enables conventional (energy-domain) Mössbauer spectroscopy at synchrotron radiation facilities. In comparison with radioactive sources, SMS provides a beam of several micrometres in size, permitting studies of extremely small samples. The SMS linewidth can be narrowed at the expense of its intensity by varying the angular position and temperature of the key element of the SMS - an iron borate 57FeBO3 crystal. Here, in order to optimize the SMS performance, the angular and temperature dependencies of the SMS parameters have been studied and the optimal angular position and temperature of the crystal have been determined for highest intensity at specified source width. The results show that, when accepting broadening of the source width up to ∼6 natural widths, the intensity of the SMS at the European Synchrotron reaches more than 105 γ-quanta s-1. In the opposite extreme, the width of the source approaches the natural width with intensity decreasing to about 103 γ-quanta s-1. These changes of intensity up to two orders of magnitude take place over a temperature range of about 0.5°C. For all temperature and angular conditions, the instrumental function of the source was derived; we also analyzed the modification of its shape when passing from the `low-width' to `high-intensity' extremes of SMS operation. Finally, we estimated the influence of the temperature instability and mosaicity of the iron borate crystal on the SMS performance.

Design simulations of a horizontally deflecting high-heat-load monochromator.

The performance of a liquid-nitrogen-cooled high-heat-load monochromator with a horizontal scattering plane has been analysed, aiming to preserve the high quality of the X-ray beam in the vertical plane for downstream optics. Using finite-element analysis, height profiles of the crystal surface for various heat loads and the corresponding slope errors in the meridional and sagittal planes were calculated. Then the angular distortions of the reflected beam in both meridional and sagittal planes were calculated analytically and also modelled by ray tracing, revealing a good agreement of the two approaches. The results show that with increasing heat load in the crystal the slope errors of the crystal surface reach their smallest values first in the sagittal and then in the meridional plane. For the considered case of interest at a photon energy of 14.412 keV and the Si(111) reflection with a Bragg angle of 7.88°, the angular distortions of the reflected beam in the sagittal plane are an order of magnitude smaller than in the meridional one. Furthermore, they are smaller than the typical angular size of the beam source at the monochromator position. For a high-heat-load monochromator operating in the horizontal scattering plane, the sagittal angular distortions of the reflected beam appear in the vertical plane. Thus, such an instrument perfectly preserves the high quality of the X-ray beam in the vertical plane for downstream optics. Compared with vertical scattering, the throughput of the monochromatic beam with the horizontal scattering plane is reduced by only 3.3% for the new EBS source, instead of 34.3% for the old ESRF-1 machine. This identifies the horizontal-scattering high-heat-load monochromator as a device essentially free of the heat-load effects in the vertical plane and without significant loss in terms of throughput.

Lattice Dynamics and Structural Phase Transitions in Eu2O3.

Using the density functional theory, we study the structural and lattice dynamical properties of europium sesquioxide (Eu2O3) in the cubic, trigonal, and monoclinic phases. The obtained lattice parameters and energies of the Raman modes show a good agreement with the available experimental data. The Eu-partial phonon density of states calculated for the cubic structure is compared with the nuclear inelastic scattering data obtained from a 20 nm thick Eu2O3 film deposited on a YSZ substrate. A small shift of the experimental spectrum to higher energies results from a compressive strain induced by the substrate. On the basis of lattice and phonon properties, we analyze the mechanisms of structural transitions between different phases of Eu2O3.

Phonon Spectroscopy in Antimony and Tellurium Oxides.

α-Sb2O3 (senarmontite), ß-Sb2O3 (valentinite), and α-TeO2 (paratellurite) are compounds with pronounced stereochemically active Sb and Te lone pairs. The vibrational and lattice properties of each have been previously studied but often lead to incomplete or unreliable results due to modes being inactive in infrared or Raman spectroscopy. Here, we present a study of the relationship between bonding and lattice dynamics of these compounds. Mössbauer spectroscopy is used to study the structure of Sb in α-Sb2O3 and ß-Sb2O3, whereas the vibrational modes of Sb and Te for each oxide are investigated using nuclear inelastic scattering, and further information on O vibrational modes is obtained using inelastic neutron scattering. Additionally, vibrational frequencies obtained by density functional theory (DFT) calculations are compared with experimental results in order to assess the validity of the utilized functional. Good agreement was found between DFT-calculated and experimental density of phonon states with a 7% scaling factor. The Sb-O-Sb wagging mode of α-Sb2O3 whose frequency was not clear in most previous studies is experimentally observed for the first time at ∼340 cm-1. Softer lattice vibrational modes occur in orthorhombic ß-Sb2O3 compared to cubic α-Sb2O3, indicating that the antimony bonds are weakened upon transforming from the molecular α phase to the layer-chained ß structure. The resulting vibrational entropy increase of 0.45 ± 0.1 kB/Sb2O3 at 880 K accounts for about half of the α-ß transition entropy. The comparison of experimental and theoretical approaches presented here provides a detailed picture of the lattice dynamics in these oxides beyond the zone center and shows that the accuracy of DFT is sufficient for future calculations of similar material structures.

Diffraction properties of a strongly bent diamond crystal used as a dispersive spectrometer for XFEL pulses.

Self-amplified spontaneous emission (SASE) enables X-ray free-electron lasers (XFELs) to generate hard X-ray pulses of sub-100 fs duration. However, due to the stochastic nature of SASE, the energy spectrum fluctuates from pulse to pulse. Many experiments that employ XFEL radiation require the resolution of the spectrum of each pulse. The work presented here investigates the capacity of a thin strongly bent diamond crystal to resolve the energy spectra of hard X-ray SASE pulses by studying its diffraction properties. Rocking curves of the symmetric C*(440) reflection have been measured for different bending radii. The experimental data match the theoretical modelling based on the Takagi-Taupin equations of dynamical diffraction. A uniform strain gradient has proven to be a valid model of strain deformations in the crystal.

Hard-X-Ray Spectroscopy with a Spectrographic Approach.

Hard-x-ray spectroscopy relies on a suite of modern techniques for studies of vibrational, electronic, and magnetic excitations in condensed matter. At present, the energy resolution of these techniques can be improved only by decreasing the spectral window of the involved optics-monochromators and analyzers-thereby sacrificing the intensity. Here, we demonstrate hard-x-ray spectroscopy with greatly improved energy resolution without narrowing the spectral window by adapting principles of spectrographic imaging to the hard-x-ray regime. Similar to Newton's classical prism, the hard-x-ray spectrograph disperses different "colors"-i.e., energies-of x-ray photons in space. Then, selecting each energy component with a slit ensures high energy resolution, whereas measuring x-ray spectra with all components of a broad spectral window keeps the intensity. We employ the principles of spectrographic imaging for phonon spectroscopy. Here the new approach revealed anomalous soft atomic dynamics in α-iron, a phenomenon which was not previously reported in the literature. We argue that hard-x-ray spectrographic imaging also could be a path to discovering new physics in studies of electronic and magnetic excitations.

Nuclear resonance reflectivity from a [57Fe/Cr]30 multilayer with the Synchrotron Mössbauer Source.

Mössbauer reflectivity spectra and nuclear resonance reflectivity (NRR) curves have been measured using the Synchrotron Mössbauer Source (SMS) for a [57Fe/Cr]30 periodic multilayer, characterized by the antiferromagnetic interlayer coupling between adjacent 57Fe layers. Specific features of the Mössbauer reflectivity spectra measured with π-polarized radiation of the SMS near the critical angle and at the `magnetic' maximum on the NRR curve are analyzed. The variation of the ratio of lines in the Mössbauer reflectivity spectra and the change of the intensity of the `magnetic' maximum under an applied external field has been used to reveal the transformation of the magnetic alignment in the investigated multilayer.

On the Structure and Reaction Mechanism of Human Acireductone Dioxygenase.

Acireductone dioxygenase (ARD) is an intriguing enzyme from the methionine salvage pathway that is capable of catalysing two different oxidation reactions with the same substrate depending on the type of the metal ion in the active site. To date, the structural information regarding the ARD-acireductone complex is limited and possible reaction mechanisms are still under debate. The results of joint experimental and computational studies undertaken to advance knowledge about ARD are reported. The crystal structure of an ARD from Homo sapiens was determined with selenomethionine. EPR spectroscopy suggested that binding acireductone triggers one protein residue to dissociate from Fe2+ , which allows NO (and presumably O2 ) to bind directly to the metal. Mössbauer spectroscopic data (interpreted with the aid of DFT calculations) was consistent with bidentate binding of acireductone to Fe2+ and concomitant dissociation of His88 from the metal. Major features of Fe vibrational spectra obtained for the native enzyme and upon addition of acireductone were reproduced by QM/MM calculations for the proposed models. A computational (QM/MM) study of the reaction mechanisms suggests that Fe2+ promotes O-O bond homolysis, which elicits cleavage of the C1-C2 bond of the substrate. Higher M3+ /M2+ redox potentials of other divalent metals do not support this pathway, and instead the reaction proceeds similarly to the key reaction step in the quercetin 2,3-dioxygenase mechanism.

A high spatial resolution synchrotron Mössbauer study of the Tazewell IIICD and Esquel pallasite meteorites.

Metallic phases in the Tazewell IIICD iron and Esquel pallasite meteorites were examined using 57Fe synchrotron Mössbauer spectroscopy. Spatial resolution of ~10-20 µm was achieved, together with high throughput, enabling individual spectra to be recorded in less than 1 h. Spectra were recorded every 5-10 µm, allowing phase fractions and hyperfine parameters to be traced along transects of key microstructural features. The main focus of the study was the transitional region between kamacite and plessite, known as the "cloudy zone." Results confirm the presence of tetrataenite and antitaenite in the cloudy zone as its only components. However, both phases were also found in plessite, indicating that antitaenite is not restricted exclusively to the cloudy zone, as previously thought. The confirmation of paramagnetic antitaenite as the matrix phase of the cloudy zone contrasts with recent observations of a ferromagnetic matrix phase using X-ray photoemission electron spectroscopy. Possible explanations for the different results seen using these techniques are proposed.

Isoprenoid Biosynthesis in Pathogenic Bacteria: Nuclear Resonance Vibrational Spectroscopy Provides Insight into the Unusual [4Fe-4S] Cluster of the E.†coli LytB/IspH Protein.

The LytB/IspH protein catalyzes the last step of the methylerythritol phosphate (MEP) pathway which is used for the biosynthesis of essential terpenoids in most pathogenic bacteria. Therefore, the MEP pathway is a target for the development of new antimicrobial agents as it is essential for microorganisms, yet absent in humans. Substrate-free LytB has a special [4Fe-4S](2+) cluster with a yet unsolved structure. This motivated us to use synchrotron-based nuclear resonance vibrational spectroscopy (NRVS) in combination with quantum chemical-molecular mechanical (QM/MM) calculations to gain more insight into the structure of substrate-free LytB. The apical iron atom of the [4Fe-4S](2+) is clearly linked to three water molecules. We additionally present NRVS data of LytB bound to its natural substrate, (E)-4-hydroxy-3-methylbut-2-en-1-yl diphosphate (HMBPP) and to the inhibitors (E)-4-amino-3-methylbut-2-en-1-yl diphosphate and (E)-4-mercapto-3-methylbut-2-en-1-yl diphosphate.

Performance of a silicon monochromator under high heat load.

The performance of a cryogenically cooled double-crystal silicon monochromator was studied under high-heat-load conditions with total absorbed powers and power densities ranging from 8 to 780 W and from 8 to 240 W mm(-2), respectively. When the temperature of the first crystal is maintained close to the temperature of zero thermal expansion of silicon, the monochromator shows nearly ideal performance with a thermal slope error of 0.6 µrad. By tuning the size of the first slit, the regime of the ideal performance can be maintained over a wide range of heat loads, i.e. from power densities of 110 W mm(-2) (at total absorbed power of 510 W) to 240 W mm(-2) (at total absorbed power of 240 W).

Nuclear quantum memory for hard x-ray photon wave packets.

Optical quantum memories are key elements in modern quantum technologies to reliably store and retrieve quantum information. At present, they are conceptually limited to the optical wavelength regime. Recent advancements in x-ray quantum optics render an extension of optical quantum memory protocols to ultrashort wavelengths possible, thereby establishing quantum photonics at x-ray energies. Here, we introduce an x-ray quantum memory protocol that utilizes mechanically driven nuclear resonant 57Fe absorbers to form a comb structure in the nuclear absorption spectrum by using the Doppler effect. This room-temperature nuclear frequency comb enables us to control the waveform of x-ray photon wave packets to a high level of accuracy and fidelity using solely mechanical motions. This tunable, robust, and highly flexible system offers a versatile platform for a compact solid-state quantum memory at room temperature for hard x-rays.

Thermal deformation of cryogenically cooled silicon crystals under intense X-ray beams: measurement and finite-element predictions of the surface shape.

X-ray crystal monochromators exposed to white-beam X-rays in third-generation synchrotron light sources are subject to thermal deformations that must be minimized using an adequate cooling system. A new approach was used to measure the crystal shape profile and slope of several cryogenically cooled (liquid nitrogen) silicon monochromators as a function of beam power in situ and under heat load. The method utilizes multiple angular scans across the Bragg peak (rocking curve) at various vertical positions of a narrow-gap slit downstream from the monochromator. When increasing the beam power, the surface of the liquid-nitrogen-cooled silicon crystal deforms from a concave shape at low heat load to a convex shape at high heat load, passing through an approximately flat shape at intermediate heat load. Finite-element analysis is used to calculate the crystal thermal deformations. The simulated crystal profiles and slopes are in excellent agreement with experiments. The parameters used in simulations, such as material properties, absorbed power distribution on the crystal and cooling boundary conditions, are described in detail as they are fundamental for obtaining accurate results.

Heme protonation affects iron-NO binding in the NO transport protein nitrophorin.

The nitrophorins (NPs) comprise an unusual group of heme proteins with stable ferric heme iron nitric oxide (Fe-NO) complexes. They are found in the salivary glands of the blood-sucking kissing bug Rhodnius prolixus, which uses the NPs to transport the highly reactive signaling molecule NO. Nuclear resonance vibrational spectroscopy (NRVS) of both isoform NP2 and a mutant NP2(Leu132Val) show, after addition of NO, a strong structured vibrational band at around 600 cm-1, which is due to modes with significant Fe-NO bending and stretching contribution. Based on a hybrid calculation method, which uses density functional theory and molecular mechanics, it is demonstrated that protonation of the heme carboxyl groups does influence both the vibrational properties of the Fe-NO entity and its electronic ground state. Moreover, heme protonation causes a significant increase of the gap between the highest occupied and lowest unoccupied molecular orbital by almost one order of magnitude leading to a stabilization of the Fe-NO bond.

Nuclear inelastic scattering and Mössbauer spectroscopy as local probes for ligand binding modes and electronic properties in proteins: vibrational behavior of a ferriheme center inside a ß-barrel protein.

In this work, we present a study of the influence of the protein matrix on its ability to tune the binding of small ligands such as NO, cyanide (CN(-)), and histamine to the ferric heme iron center in the NO-storage and -transport protein Nitrophorin 2 (NP2) from the salivary glands of the blood-sucking insect Rhodnius prolixus. Conventional Mössbauer spectroscopy shows a diamagnetic ground state of the NP2-NO complex and Type I and II electronic ground states of the NP2-CN(-) and NP2-histamine complex, respectively. The change in the vibrational signature of the protein upon ligand binding has been monitored by Nuclear Inelastic Scattering (NIS), also called Nuclear Resonant Vibrational Spectroscopy (NRVS). The NIS data thus obtained have also been calculated by quantum mechanical (QM) density functional theory (DFT) coupled with molecular mechanics (MM) methods. The calculations presented here show that the heme ruffling in NP2 is a consequence of the interaction with the protein matrix. Structure optimizations of the heme and its ligands with DFT retain the characteristic saddling and ruffling only if the protein matrix is taken into account. Furthermore, simulations of the NIS data by QM/MM calculations suggest that the pH dependence of the binding of NO, but not of CN(-) and histamine, might be a consequence of the protonation state of the heme carboxyls.

The 57Fe Synchrotron Mössbauer Source at the ESRF.

The design of a (57)Fe Synchrotron Mössbauer Source (SMS) for energy-domain Mössbauer spectroscopy using synchrotron radiation at the Nuclear Resonance beamline (ID18) at the European Synchrotron Radiation Facility is described. The SMS is based on a nuclear resonant monochromator employing pure nuclear reflections of an iron borate ((57)FeBO(3)) crystal. The source provides (57)Fe resonant radiation at 14.4 keV within a bandwidth of 15 neV which is tunable in energy over a range of about ±0.6 µeV. In contrast to radioactive sources, the beam of γ-radiation emitted by the SMS is almost fully resonant and fully polarized, has high brilliance and can be focused to a 10 µm × 5 µm spot size. Applications include, among others, the study of very small samples under extreme conditions, for example at ultrahigh pressure or combined high pressure and high temperature, and thin films under ultrahigh vacuum. The small cross section of the beam and its high intensity allow for rapid collection of Mössbauer data. For example, the measuring time of a spectrum for a sample in a diamond anvil cell at ∼100 GPa is around 10 min, whereas such an experiment with a radioactive point source would take more than one week and the data quality would be considerably less. The SMS is optimized for highest intensity and best energy resolution, which is achieved by collimation of the incident synchrotron radiation beam and thus illumination of the high-quality iron borate crystal within a narrow angular range around an optimal position of the rocking curve. The SMS is permanently located in an optics hutch and is operational immediately after moving it into the incident beam. The SMS is an in-line monochromator, i.e. the beam emitted by the SMS is directed almost exactly along the incident synchrotron radiation beam. Thus, the SMS can be easily utilized with all existing sample environments in the experimental hutches of the beamline. Owing to a very strong suppression of electronic scattering for pure nuclear reflections (∼10(-9)), SMS operation does not required any gating of the prompt electronic scattering. Thus, the SMS can be utilized in any mode of storage ring operation.

Interpretation of nuclear resonant vibrational spectra of rubredoxin using a combined quantum mechanics and molecular mechanics approach.

Nuclear resonant vibrational spectra of the reduced and oxidized form of a mutant of rubredoxin from Pyrococcus abyssii were measured and are compared with simulated spectra that were calculated by a combined quantum mechanics (QM) and molecular mechanics (MM) method. Density functional theory was used for the QM level. Calculations were performed for different models of rubredoxin. Realistic spectra were simulated with reduced models that include at least the iron center, the four cysteins coordinating it, and the residues connected to the cysteins together with a QM layer that comprises the first two coordination shells of the iron center. Larger QM layers did not lead to significant changes of the simulated spectra.

Phonon confinement and interface lattice dynamics of ultrathin high-k rare earth sesquioxide films: the case of Eu2O3 on YSZ(001).

The spatial confinement of atoms at surfaces and interfaces significantly alters the lattice dynamics of thin films, heterostructures and multilayers. Ultrathin films with high dielectric constants (high-k) are of paramount interest for applications as gate layers in current and future integrated circuits. Here we report a lattice dynamics study of high-k Eu2O3 films with thicknesses of 21.3, 2.2, 1.3, and 0.8 nm deposited on YSZ(001). The Eu-partial phonon density of states (PDOS), obtained from nuclear inelastic scattering, exhibits broadening of the phonon peaks accompanied by up to a four-fold enhancement of the number of low-energy states compared to the ab initio calculated PDOS of a perfect Eu2O3 crystal. Our analysis demonstrates that while the former effect reflects the reduced phonon lifetimes observed in thin films due to scattering from lattice defects, the latter phenomenon arises from an ultrathin EuO layer formed between the thin Eu2O3 film and the YSZ(001) substrate. Thus, our work uncovers another potential source of vibrational anomalies in thin films and multilayers, which has to be cautiously considered.

Space Charge-Limited Current Transport Mechanism in Crossbar Junction Embedding Molecular Spin Crossovers.

Spin crossover complexes are among the most studied classes of molecular switches and have attracted considerable attention for their potential technological use as active units in multifunctional devices. A fundamental step toward their practical implementation is the integration in macroscopic devices adopting hybrid vertical architectures. First, the physical properties of technological interest shown by these materials in the bulk phase have to be retained once they are deposited on a solid surface. Herein, we describe the study of a hybrid molecular inorganic junction embedding the spin crossover complex [Fe(qnal)2] (qnal = quinoline-naphthaldehyde) as an active switchable thin film sandwiched within energy-optimized metallic electrodes. In these junctions, developed and characterized with the support of state of the art techniques including synchrotron Mössbauer source (SMS) spectroscopy and focused-ion beam scanning transmission electron microscopy, we observed that the spin state conversion of the Fe(II)-based spin crossover film is associated with a transition from a space charge-limited current (SCLC) transport mechanism with shallow traps to a SCLC mechanism characterized by the presence of an exponential distribution of traps concomitant with the spin transition temperature.