Two-dimensional Hyperfine Sublevel Correlation Spectroscopy Research Articles

Back-clocking of Fe2+/Fe1+ spin states in a H2-producing catalyst by advanced EPR

A mononuclear Fe-(P(PPh2)3) ((P(PPh2)3) = tris[2-diphenylphospino)ethyl]phosphine) catalyst was studied in situ under catalytic conditions using advanced electron paramagnetic resonance (EPR) techniques. Fe-(P(PPh2)3) efficiently catalyses H2 production using HCOOH as substrate. Dual-mode continuous-wave (CW) EPR, used to study the initial Fe2+(S = 2) state, shows that the complex is characterised by a – rather small – zero field splitting parameter Δ = 0.45 cm−1 and geff = 8.0. In the presence of HCOOH substrate the complex evolves and a unique Fe1+(S = 1/2) state is trapped. The Fe1+ atom is coordinated by four 31P nuclei in a pseudo-C3 symmetry. Only a small fraction of the Fe1+ spin density is delocalised onto the 31P atoms. Four-pulse electron spin echo envelope modulation (ESEEM) and two-dimensional hyperfine sublevel correlation spectroscopy (2D-HYSCORE) data reveal the existence of two types of 1H couplings. One corresponds to weak, purely dipolar coupling, tentatively assigned to phenyl protons. The most important is a – rather unusual – 1H coupling with negative Aiso (−2.75 MHz) and strong dipolar part (T = 5.5 MHz). This 1H is located on the pseudo-C3 symmetry axis of the Fe1+-(P(PPh2)3-HCOO− complex where one substrate molecule, formate anion, is coordinated on the Fe1+ atom.

Caught in the act

Femtobiology freeze‐frames crucial split seconds of chemical reactions to investigate how enzymes function. The potential prize from this knowledge could be new avenues for drug development or ways to produce clean energy.

The structure and activation of substrate water molecules in the S2 state of photosystem II studied by hyperfine sublevel correlation spectroscopy

The water-splitting protein, photosystem II, catalyzes the light-driven oxidation of water to dioxygen. The solar water oxidation reaction takes place at the catalytic center, referred to as the oxygen-evolving complex, of photosystem II. During the catalytic cycle, the oxygen-evolving complex cycles through five distinct intermediate states, S0–S4. In this study, we trap the oxygen-evolving complex in the S2 intermediate state by low temperature illumination of photosystem II isolated from three different species, Thermosynechococcus vulcanus, the PsbB variant of Synechocystis PCC 6803 and spinach. We apply two-dimensional hyperfine sublevel correlation spectroscopy to detect weak magnetic interactions between the paramagnetic tetra-nuclear manganese cluster of the S2 state of the OEC and the surrounding protons. We identify five groups of protons that are interacting with the tetra-nuclear manganese cluster. From the values of hyperfine interactions and using the recently reported 1.9 A resolution X-ray structure of the OEC in the S1 state [Umena et al., Nature, 2011, 473, 55], we discuss the assignments of the five groups of protons and draw important conclusions on the structure of the oxygen-evolving complex in the S2 state. In addition, we conclude that the structure of OEC is nearly identical in photosystem II from Thermosynechococcus vulcanus, the PsbB variant of Synechocystis PCC 6803 and spinach.

High-Resolution Two-Dimensional1H and14N Hyperfine Sublevel Correlation Spectroscopy of the Primary Quinone of Photosystem II

Quinones are naturally occurring isoprenoids that are widely exploited by photosynthetic reaction centers. Protein interactions modify the properties of quinones such that similar quinone species can perform diverse functions in reaction centers. Both type I and type II (oxygenic and nonoxygenic, respectively) reaction centers contain quinone cofactors that serve very different functions as the redox potential of similar quinones can operate at up to 800 mV lower reduction potential when present in type I reaction centers. However, the factors that determine quinone function in energy transduction remain unclear. It is thought that the location of the quinone cofactor, the geometry of its binding site, and the "smart" matrix effects from the surrounding protein environment greatly influence the functional properties of quinones. Photosystem II offers a unique system for the investigation of the factors that influence quinone function in energy transduction. It contains identical plastoquinones in the primary and secondary quinone acceptor sites, Q(A) and Q(B), which exhibit very different functional properties. This study is focused on elucidating the tuning and control of the primary semiquinone state, Q(A)(-), of photosystem II. We utilize high-resolution two-dimensional hyperfine sublevel correlation spectroscopy to directly probe the strength and orientation of the hydrogen bonds of the Q(A)(-) state with the surrounding protein environment of photosystem II. We observe two asymmetric hydrogen bonding interactions of reduced Q(A)(-) in which the strength of each hydrogen bond is affected by the relative nonplanarity of the bond. This study confirms the importance of hydrogen bonds in the redox tuning of the primary semiquinone state of photosystem II.

Ligand Environment of the S2State of Photosystem II: A Study of the Hyperfine Interactions of the Tetranuclear Manganese Cluster by 2D14N HYSCORE Spectroscopy

The solar water-splitting protein complex, photosystem II, catalyzes the light-driven oxidation of water to dioxygen in Nature. The four-electron oxidation reaction of water occurs at the tetranuclear manganese-calcium-oxo catalytic cluster that is present in the oxygen-evolving complex of photosystem II. The mechanism of light-driven water oxidation has been a subject of intense interest, and the oxygen-evolving complex of photosystem II has been studied extensively by structural and biochemical methods. While the recent X-ray crystal structures and single-crystal EXAFS investigations provide a model for the geometry of the tetranuclear manganese-calcium-oxo catalytic cluster, there is limited knowledge of the protein environment that surrounds the catalytic cluster. In this study, we demonstrate the application of two-dimensional hyperfine sublevel correlation spectroscopy to determine the magnetic couplings of the catalytic cluster with the (14)N atoms of surrounding amino acid residues in the S(2) state of the oxygen-evolving complex of photosystem II. We utilize two-dimensional difference spectroscopy to facilitate unambiguous assignments of the spectral features and identify at least three separate (14)N atoms that are interacting with the catalytic cluster in the S(2) state. The results presented here, for the first time, identify previously unknown ligands to the catalytic cluster of photosystem II and provide avenues for the assignment of residues by site-directed mutagenesis and the refinement of computational and mechanistic models of photosystem II.

1D- and 2D-ESEEM Study of the Semiquinone Radical QA- of Photosystem II

The semiquinone radical QA- has been studied by Electron Spin−Echo Envelope Modulation (ESEEM) spectroscopy in Photosystem II membranes at various pH values. The observed nuclear modulations have been assigned by the use of two-dimensional Hyperfine Sublevel Correlation Spectroscopy (HYSCORE) and numerical simulations. Two protein 14N nuclei (NI and NII) were found to be magnetically coupled with the QA- spin, and on the basis of 14N-NQR and 14N-ESEEM data from the literature, NI is assigned to an amide nitrogen from the protein backbone while NII is assigned to the amino nitrogen, Nδ, of an imidazole. A physical explanation for such couplings is suggested where the coupling occurs through H-bonds from the protein to the carbonyls of the semiquinone. In PSII membranes treated with CN-, only the NI coupling is present above pH 8.5 while both NI and NII couplings are present at lower pH values. In samples treated at high pH to remove the iron, both NI and NII couplings are present throughout the pH range st...

Hyperfine sublevel correlation spectroscopy in lithium silicate glasses

The structure of γ-irradiated (Li2O)0.1(SiO2)0.9 glass has been studied with two-dimensional hyperfine sublevel correlation spectroscopy (HYSCORE). The theoretical evaluation of the HYSCORE experimental data revealed the existence of weak couplings between the electron spin and nearby Li. The anisotropic hyperfine coupling T is equal to 4.0 MHz suggesting a nonbridging oxygen—Li distance of 2.0 Å and an angle of ±60° away from the gz axis. The Aiso value is equal to 0.5 MHz, corresponding to 0.135% contribution of the Li 2s orbital to the ground state. Thus, an estimation can be made of the covalent character of the bond between the nonbridging oxygen and the alkali.

Electronic structure ofB2O3glass studied by one- and two-dimensional electron-spin-echo envelope modulation spectroscopy

${\mathrm{B}}_{2}{\mathrm{O}}_{3}$ in glass and crystalline states have been subjected to \ensuremath{\gamma} irradiation at room temperature and subsequently studied by continuous-wave electron paramagnetic resonance and electron-spin-echo envelope modulation (ESEEM) spectroscopy at liquid-helium temperature. The ESEEM study of ${\mathrm{B}}_{2}{\mathrm{O}}_{3}$ revealed the existence of weak magnetic couplings between the irradiation-induced paramagnetic centers and nearby ${}^{10}\mathrm{B},$ ${}^{11}\mathrm{B}$ nuclei. The assignment of the observed couplings was achieved by the use of the two-dimensional hyperfine sublevel correlation spectroscopy. A detailed theoretical analysis of the ESEEM spectra and a $S=\frac{1}{2},$ $I=\frac{3}{2}$ system in both time and frequency domain is also presented. Computer simulation of the spectra, Hartree-Fock self-consistent field, and modified neglect of differential overlap calculations revealed that the unpaired electron in the ${\mathrm{B}}_{2}{\mathrm{O}}_{3}$ glass is associated with a dangling bond of oxygen attached to a boron of a boroxol ring. The paramagnetic centers of the ${\mathrm{B}}_{2}{\mathrm{O}}_{3}$ crystal are associated with oxygen dangling bonds in boron trigonal units. Pertinent structural models for the glass ${\mathrm{B}}_{2}{\mathrm{O}}_{3}$ are examined at a microscopic level.

ESEEM Investigations of the High pH and Low pH Forms of Chicken Liver Sulfite Oxidase

Two-pulse and four-pulse electron spin−echo envelope modulation spectroscopy (ESEEMS) at two operational frequencies and two-dimensional hyperfine sublevel correlation spectroscopy (HYSCORES), have been used to probe the MoV coordination environment of sulfite oxidase in H2O and D2O solutions, buffered at pH 9.5 and 7.0 with ∼100 mM Tris-type buffers. At pH 9.5 the ESEEM and HYSCORE results definitively reveal the presence of one solvent-exchangeable D(H) near the MoV center, probably in the form of a Mo−OH(D) moiety. The orientation of this group is not fixed (although it is substantially restricted) and thus gives rise to a distribution of hyperfine interaction (hfi) parameters. The resulting loss of amplitude makes direct observation of a proton-related line using ESEEM impossible. However, such a line is observable in ESEEM spectra of the comparable deuterated enzyme because the narrower distribution of hfi parameters leads to less line broadening of the ESEEM spectra. ESEEM and HYSCORE spectra of sul...

Coordination Geometry of the Copper−Pyridine Complex in Frozen Solution As Studied by Proton and Deuterium Two-Dimensional Hyperfine Sublevel Correlation Electron Spin Resonance Spectroscopy

The coordination geometry of the tetrapyridine−copper(II) complex in frozen solution is investigated by proton and deuterium two-dimensional hyperfine sublevel correlation spectroscopy (HYSCORE) electron spin resonance experiments. In particular, the deuterium experiment demonstrates the potential of this method, which lies in the superior spectral resolution of the two-dimensional spectra. This allows us to resolve deuterium nuclear quadrupole splittings of the cross peak ridges even in orientationally disordered systems, which in turn yield structural information about the overall complex symmetry. Proton and deuterium experiments show pronounced cross peak ridges from protons and deuteriums at the C2 and C6 carbon atoms of the pyridine molecule. The coordination geometry within the complexes could be deduced from orientation-selective deuterium spectra. Severe deviations from the D4h complex symmetry were found in such a way that the pyridine molecules are arranged with their molecular mirror plane per...

A Practical Strategy for Determination of Proton Hyperfine Interaction Parameters in Paramagnetic Transition Metal Ion Complexes by Two-Dimensional HYSCORE Electron Spin Resonance Spectroscopy in Disordered Systems

A practical strategy is outlined for the determination of proton hyperfine parameters in paramagnetic transition metal ion complexes in disordered systems from a single two-dimensional hyperfine sublevel correlation spectroscopy (2D HYSCORE) electron spin resonance experiment. Both dipolar and isotropic hyperfine interaction parameters can directly be determined from the cross peak ridges in the HYSCORE spectrum in the limit of the point dipole approximation. This approach is justified by spectral simulations for isotropic and axially symmetric g tensors. If the HYSCORE spectrum is measured at the g⊥ spectral region of the electron spin resonance powder spectrum, the orientation of the hyperfine interaction tensor with respect to the g tensor frame can also be deduced from the shape of the cross peak ridges in many cases. Two experimental examples are presented. Using this approach, the hyperfine interaction parameters for protons in [Cu(H2O)6]2+ and in [Cu(C5H5N)4]2+ complexes, both incorporated into mesoporous (L)Cu−MCM-41 silica tube material, are determined from a single 2D HYSCORE spectrum. The parameters are in good agreement with independent measurements by electron spin echo modulation spectroscopy.