ENDOR Spectra Research Articles

EPR and ENDOR studies of self( α)-irradiation effects in Pu(VI) oxalate: evidence for participation of 5f electrons of 239Pu in chemical bonding with CO 2−

EPR and ENDOR investigations have been carried out on polycrystalline samples of self( α)-irradiated PuO 2C 2O 4·3H 2O (cumulative dose = 1 MGy). The 239Pu ( I=1/2) hyperfine structure on the EPR signal of the CO 2 − radical ion has given clear evidence for participation of 5f electrons of 239Pu in chemical bonding. The ENDOR spectra were used to identify unambiguously the hyperfine interaction between the unpaired electron on the CO 2 − radical ion and the nuclear spin of 239Pu. In addition to the CO 2 − radical ion, EPR evidence was also obtained for the formation of O 2 − during the autoradiolysis of PuO 2C 2O 4·3H 2O.

Diiron Complexes of 1,8-Naphthyridine-Based Dinucleating Ligands as Models for Hemerythrin

The hydroxo-bridged diiron(II) compounds [Fe2(BPEAN)(μ-OH)(OTf)](OTf)2 (1) and [Fe2(BEPEAN)(μ-OH)](OTf)3 (2) were prepared by using 1,8-naphthyridine-based dinucleating ligands BPEAN and BEPEAN, where BPEAN = 2,7-bis{bis[2-(2-pyridyl)ethyl]aminomethyl}-1,8-naphthyridine and BEPEAN = 2,7-bis{bis[2-(2-(5-ethyl)pyridyl)ethyl]aminomethyl}-1,8-naphthyridine. When compound 2 was treated with excess 30% aqueous H2O2 in acetonitrile at −40 °C, a red-brown species (3) was produced. The UV−vis spectrum of 3 exhibited an absorption maximum at 505 nm (ε = 1500 M-1 cm-1), close to that observed for oxyHr. Resonance Raman experiments revealed an isotope-sensitive O−O stretching band at 868 cm-1. When a mixture of 1:1 H2O2/D2O2 (25% in 1:1 H2O/D2O) was used to generate 3, a broader Raman band centered at 870 cm-1 appeared, indicating the peroxide group to be protonated. The 1H ENDOR spectrum of 3, cryoreduced to the diiron(II,III) state, showed a signal with A ≈ 12 MHz that disappeared when D2O2 in D2O was used to generate 3, providing further evidence for the presence of a hydroperoxide ligand bound to iron. The EPR spectrum of the cryoreduced sample revealed that 3 has a (μ-oxo)diiron(III) core, a conclusion supported by Mössbauer spectroscopy. The Mössbauer spectrum exhibited the unusual quadrupole splitting values that are characteristic of the diiron(III) center of oxyHr. Thus, all spectroscopic properties of 3 are consistent with it being a hydroperoxo-bound (μ-oxo)diiron(III) complex. The hydroperoxide ligand is more resistant to deprotonation than in mononuclear iron(III) analogues, which may reflect the presence of a hydrogen bond between the hydroperoxide and bridging oxide groups. At room temperature, acetonitrile/water solutions of 3 decayed to iron(II) species, releasing the iron-bound hydroperoxide group to form O2.

Hyperfine Shifts in Low-Spin Iron(III) Hemes: A Ligand Field Analysis

The hyperfine shifts of heme nuclei in low-spin iron(III) porphyrins of known geometry are predicted from: (i) a ligand field analysis, and (ii) the Kurland and McGarvey theory (R. J. Kurland, B. R. McGarvey, J. Magn. Reson.1970, 2, 286−301). The ligand field parameters are optimized to reproduce the experimentally available g values and magnetic susceptibility anisotropy, the latter being obtained from the analysis of pseudo-contact shifts. Simple known geometric relationships (I. Bertini, C. Luchinat, G. Parigi, F. A. Walker, J. Biol. Inorg. Chem.1999, 4, 515−519; N. V. Shokhirev, F.A. Walker, J. Biol. Inorg. Chem.1998, 3, 581−594), are used to relate the contact shifts to the various heme positions. The systems investigated are: metMyoglobin cyanide, cytochrome b5 and cytochrome c. It is shown that the contact hyperfine coupling constants recalculated from this approach are different to those obtained through the simple and generally used McConnell equation, and that the deviation can be more or less severe, depending on the position on the heme and on the orientation of the axial ligands. The contact shift tensor is highly anisotropic and therefore the shift is dependent on the orientation of the molecule in a magnetic field. This theoretical analysis is necessary to interpret ENDOR spectra and NMR spectra in partially oriented systems.

Two-dimensional pulsed TRIPLE at 95 GHz

The one-dimensional (1D) pulsed TRIPLE resonance experiment, introduced by Mehring et al. (M. Mehring, P. Hofer, and A. Grupp, Ber. Bunseges. Phys. Chem. 91, 1132-1137 (1987)) is a modification of the standard Davies ENDOR experiment where an additional RF pi-pulse is applied during the mixing time. While the first RF pulse is set to one of the ENDOR transitions, the frequency of the second RF pulse is scanned to generate the TRIPLE spectrum. The difference between this spectrum and the ENDOR spectrum yields the difference TRIPLE spectrum, which exhibits only ENDOR lines that belong to the same M(S) manifold as the one selected by the first RF pulse. We have extended this experiment in two dimensions (2D) by sweeping the frequencies of both RF pulses. This experiment is particularly useful when the spectrum is congested and consists of signals originating from different paramagnetic centers. The connectivities between the peaks in the 2D spectrum enable a straightforward assignment of the signals to their respective centers and M(S) manifolds, thus providing the relative signs of hyperfine couplings. Carrying out the experiment at high fields has the additional advantage that nuclei with different nuclear gyromagnetic ratios are well separated. This is particularly true for protons which appear at significantly higher frequencies than other nuclei. The feasibility and effectiveness of the experiment is demonstrated at W-band (94.9 GHz) on a crystal of Cu(2+)-doped l-histidine. Homonuclear (1)H-(1)H, (14)N/(35)Cl-(14)N/(35)Cl and heteronuclear (1)H-(14)N/(35)Cl 2D TRIPLE spectra were measured and from the various connectivities in the 2D map the (1)H, (14)N, and (35)Cl signals that belong to two different Cu(2+) centers were identified and grouped according to their M(S) manifolds. Copyright 2000 Academic Press.

ENDOR spectroscopic studies of stable semiquinone radicals bound to the Escherichia coli cytochrome bo3 quinol oxidase.

The putative oxidation of ubiquinol by the cytochrome bo3 terminal oxidase of Escherichia coli in sequential one-electron steps requires stabilization of the semiquinone. ENDOR spectroscopy has recently been used to study the native ubisemiquinone radical formed in the cytochrome bo3 quinone-binding site [Veselov, A.V., Osborne, J.P., Gennis, R.B. & Scholes, C.P. (2000) Biochemistry 39, 3169-3175]. Comparison of these spectra with those from the decyl-ubisemiquinone radical in vitro indicated that the protein induced large changes in the electronic structure of the ubisemiquinone radical. We have used quinone-substitution experiments to obtain ENDOR spectra of ubisemiquinone, phyllosemiquinone and plastosemiquinone anion radicals bound at the cytochrome bo3 quinone-binding site. Large changes in the electronic structures of these semiquinone anion radicals are induced on binding to the cytochrome bo3 oxidase. The changes in electronic structure are, however, independent of the electronic structures of these semiquinones in vitro. Thus it is shown to be the structure of this binding site in the protein, not the covalent structure of the bound quinone, that determines the electronic structure of the protein-bound semiquinone.

Conformational relaxation following reduction of the photoactive bacteriopheophytin in reaction centers from Blastochloris viridis. : Influence of mutations at position M208

The photochemically trapped bacteriopheophytin (BPh) b radical anion in the active branch (Φ A − ) of reaction centers (RCs) from Blastochloris (formerly called Rhodopseudomonas) viridis is characterized by 1H-ENDOR as well as optical absorption spectroscopy. The two site-directed mutants YF(M208) and YL(M208), in which tyrosine at position M208 is replaced by phenylalanine and leucine, respectively, are investigated and compared with the wild type. The residue at M208 is in close proximity to the primary electron donor, P, the monomeric bacteriochlorophyll (BChl), B A, and the BPh, Φ A, that are involved in the transmembrane electron transfer to the quinone, Q A, in the RC. The analysis of the ENDOR spectra of Φ A − at 160 K indicates that two distinct states of Φ A − are present in the wild type and the mutant YF(M208). Based on a comparison with Φ A − in RCs of Rhodobacter sphaeroides the two states are interpreted as torsional isomers of the 3-acetyl group of Φ A. Only one Φ A − state occurs in the mutant YL(M208). This effect of the leucine residue at position M208 is explained by steric hindrance that locks the acetyl group in one specific position. On the basis of these results, an interpretation of the optical absorption difference spectrum of the state Φ A − Q A − is attempted. This state can be accumulated at 100 K and undergoes an irreversible change between 100 and 200 K [Tiede et al., Biochim. Biophys. Acta 892 (1987) 294–302]. The corresponding absorbance changes in the BChl Q x and Q y regions observed in the wild type also occur in the YF(M208) mutant but not in YL(M208). The observed changes in the wild type and YF(M208) are assigned to RCs in which the 3-acetyl group of Φ A changes its orientation. It is concluded that this distinct structural relaxation of Φ A can significantly affect the optical properties of B A and contribute to the light-induced absorption difference spectra.

Effects of temperature on the ESR and ENDOR spectra of the DMPO–OBut spin adduct; the problems of identifying the trapped radical

The ESR and ENDOR spectra and hyperfine coupling constants (hfi) of the tertiary butoxyl spin adduct of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) change markedly, but reversibly, with increase in temperature from 140 to 280 K, and the spectra can be cycled repeatedly between these temperature limits. The variation with temperature of the hfi of nitrogen, β and γ protons arises through a change in equilibrium concentrations of the planar and puckered conformers of the adduct pyrroline ring which are interconverting rapidly on the ESR time-scale (107 MHz). A simple mathematical model has been derived which successfully simulates the experimental temperature variation of the coupling constants, gives their values in the two conformers and estimates of 4.3 ± 0.5 kJ mol−1 for the energy difference between the conformers in three solvents. Accurate hfi measurements by ENDOR spectroscopy across a narrow temperature range (140–160 K) corroborated ESR experiments and afforded the assignment of all magnetically inequivalent protons that have previously eluded observation. By using the di-tert-butyl tetroxide equilibrium, tROOOORt ⇌ 2tROO· as a clean source of tert-peroxyl radicals, it was shown that these radicals are not trapped singly by DMPO to give a peroxyl spin adduct but give an ESR-silent complex which decomposes at higher temperatures to the DMPO–OBut spin adduct. These studies highlight the problems of identifying the trapped radical from ESR spectra of spin adducts especially when the spectra are observed at only one temperature. Copyright © 2000 John Wiley & Sons, Ltd.

An ENDOR study of H atoms in HD solid at 4.2 K

An ENDOR spectrum of H atoms in the γ-ray irradiated HD solid was measured at 4.2 K for elucidating the local environment of HD molecules surrounding H atoms. The analysis of the ENDOR spectrum indicates that the H atoms are not trapped in substitutional but in pseudo octahedral interstitial sites of HD solid. It is postulated that an H atom, produced by the γ-irradiation, initially enters the octahedral interstitial site, and then the HD molecules at the first nearest site from the H atom are pushed back by zero-point vibration of the H atom.

W-Band ENDOR Investigation of the Manganese-Binding Site of Concanavalin A: Determination of Proton Hyperfine Couplings and Their Signs

W-band (95 GHz) pulsed EPR and electron−nuclear double resonance spectroscopic techniques were used to investigate the manganese S1 binding site of the protein concanavalin A in a frozen solution and in a single crystal. 1H ENDOR spectra were collected by using the Davies ENDOR sequence, whereas Mims ENDOR was applied to record 2H ENDOR spectra. Different EPR transitions were selected by performing the ENDOR measurements at different magnetic fields within the EPR spectrum. This selection, combined with the large thermal polarization achieved at magnetic fields of ∼3.4 T at low temperatures, allowed the assignment of ENDOR signals to their respective MS manifolds, thus providing the sign of the hyperfine couplings. The exchangeable and nonexchangeable protons were differentiated by comparing 1H and 2H ENDOR spectra of a solution of the protein prepared in D2O and of crystals soaked in D2O. The two imidazole protons, located on the carbons flanking the Mn-bound nitrogen, are magnetically equivalent, situated 3.56 Å from the Mn2+ and their hyperfine coupling is purely dipolar. The four protons of the two water ligands are all inequivalent, and four values of A∥ and A⊥ were determined. All possible combinations of these values yield distances in the range of 2.67 to 3.24 Å and aiso between −1.13 and 1.37 MHz. By limiting aiso to positive values, only four distances remain, 2.67, 2.76, 2.99, and 3.24 Å, corresponding to the four Mn−Hwater distances.

Photophysics of AgCl doped with [Cl5Ir(N-methylpyrazinium)]-: II. ENDOR studies of 13C enriched and 2D exchanged complexes

Additional information has been obtained on the structure of donor complexes produced by photoelectron trapping at [Cl5Ir(NMP)]- (NMP = N-methylpyrazinium cation) centres in AgCl microcrystals. Several different [Cl5Ir(NMP)]- complexes were prepared by full or partial deuteration of the NMP ligand protons, and by isotopically enriching the methyl group carbon with 13C. The analysis of the corresponding changes in the [Cl5Ir(NMP)]2- powder ENDOR spectra made it possible to assign SHF lines to specific proton/deuteron sites. In addition, an SHF interaction from the methyl group carbon was resolved in the 13C-enriched compounds. These observations, combined with preliminary results for the related donor [{(CN)}5Fe(NMP)]3- and consideration of the g matrix, support the previous contention that [Cl5Ir(NMP)]- is incorporated intact into AgCl dispersions during precipitation.

Effects of temperature on the ESR and ENDOR spectra of the DMPO-OBut spin adduct; the problems of identifying the trapped radical

The ESR and ENDOR spectra and hyperfine coupling constants (hfi) of the tertiary butoxyl spin adduct of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) change markedly, but reversibly, with increase in temperature from 140 to 280 K, and the spectra can be cycled repeatedly between these temperature limits. The variation with temperature of the hfi of nitrogen, β and γ protons arises through a change in equilibrium concentrations of the planar and puckered conformers of the adduct pyrroline ring which are interconverting rapidly on the ESR time-scale (107 MHz). A simple mathematical model has been derived which successfully simulates the experimental temperature variation of the coupling constants, gives their values in the two conformers and estimates of 4.3 ± 0.5 kJ mol−1 for the energy difference between the conformers in three solvents. Accurate hfi measurements by ENDOR spectroscopy across a narrow temperature range (140–160 K) corroborated ESR experiments and afforded the assignment of all magnetically inequivalent protons that have previously eluded observation. By using the di-tert-butyl tetroxide equilibrium, tROOOORt ⇌ 2tROO· as a clean source of tert-peroxyl radicals, it was shown that these radicals are not trapped singly by DMPO to give a peroxyl spin adduct but give an ESR-silent complex which decomposes at higher temperatures to the DMPO–OBut spin adduct. These studies highlight the problems of identifying the trapped radical from ESR spectra of spin adducts especially when the spectra are observed at only one temperature. Copyright © 2000 John Wiley & Sons, Ltd.

EPR and ENDOR studies on CO2- radicals in γ-irradiated B-type carbonated apatites

A gamma-irradiated synthetic B-type carbonated apatite calcined at 300°C was studied by EPR and ENDOR. The room temperature EPR spectrum is composed of an axially symmetric species (g⊥=2.0025 and g‖=1.9971) attributed to rotating CO2− radicals. At low temperatures (T<10 K), the CO2−1H and 31P ENDOR spectra were obtained for different magnetic field settings. Computer simulations of these ENDOR spectra, based on the ‘‘molecular orientation–selection’’ principle, enabled one to allocate the CO2− radicals in the carbonated apatite lattice. It was established that the CO2− radicals are located on PO43− sites with a vacancy on the nearest OH− site. In addition, discussions related to the substitution mechanisms and the nature of the precursors of the CO2− radicals are presented.

Structure of N2H4•+formed in X-irradiated Li(N2H5)SO4 single crystals

X-irradiated Li(N2H5)SO4 single crystals were investigated using ESR and ENDOR spectroscopy at several temperatures. The hydrazine radical cation N2H4•+ was selectively produced by irradiation at room temperature. From the analysis of the orientation dependent ENDOR spectra, the 1H-hfc tensors of the cation radical were precisely obtained and the radical structure was supported by theoretical calculations. It is suggested that the cation radical has a planar π* structure D2h (2B2g) in the crystal down to 230 K. By using the evaluated 1H-hf tensor the powder ESR line shape was successfully simulated. Concomitant with the radical formation, the N–N bond of N2H4•+ is suggested to reorient so as to optimize hydrogen bond interactions. 1H-ENDOR line splitting for the N2H4•+ radical was observed at temperatures below 230 K. Apparently this splitting is due to a reversible structural change where one of the NH2 moieties in N2H4•+ becomes slightly bent out of the molecular plane, whereas the other one remains planar. This deformation evidently arises from interactions between the cation radical, adjacent H2N–NH3+ molecules and the SO4–LiO4 framework. Interacting N2H5+···N2H4•+···N2H5+ molecules along the c-axis are proposed to explain the deformation mechanism.

Recognition of chirality in nitroxides using EPR and ENDOR spectroscopy

The question regarding chirality in nitroxides was rigorously investigated, namely, (a) are multiple EPR and ENDOR spectra expected from nitroxides with chiral centers, (b) which structures provide multiple structures and (c) under what conditions (temperature and solvent) are they obtained. It is shown that nitroxides with one chiral center produce only one EPR/ENDOR spectrum in toluene or ethanol (293–193 K); the nitroxyl function is not a chiral center under these conditions. Nitroxides with two chiral centers (α- and β-position) give two EPR/ENDOR spectra consistent with a pair of diastereomeric configurations (see series II), but this observation was not apparent in every case (see series III). This anomaly was attributed to a stereoselective production of certain nitroxides which apparently do not produce diastereomeric mixtures in alcohols as solvent. Further examples of multiple EPR/ENDOR spectra were found when the position of chirality is in the α- and β-position to a remote spin center such as the nitroxide function (see series IV and V). Finally, EPR/ENDOR spectra were obtained from nitroxide derivatives wherein the absolute configuration at the α-carbon atom was known. Distinctly different EPR/ENDOR spectra were indeed obtained for every diastereomer, thus establishing this point unequivocally. Copyright © 1999 John Wiley & Sons, Ltd.

Recognition of chirality in nitroxides using EPR and ENDOR spectroscopy

The question regarding chirality in nitroxides was rigorously investigated, namely, (a) are multiple EPR and ENDOR spectra expected from nitroxides with chiral centers, (b) which structures provide multiple structures and (c) under what conditions (temperature and solvent) are they obtained. It is shown that nitroxides with one chiral center produce only one EPR/ENDOR spectrum in toluene or ethanol (293–193 K); the nitroxyl function is not a chiral center under these conditions. Nitroxides with two chiral centers (α- and β-position) give two EPR/ENDOR spectra consistent with a pair of diastereomeric configurations (see series II), but this observation was not apparent in every case (see series III). This anomaly was attributed to a stereoselective production of certain nitroxides which apparently do not produce diastereomeric mixtures in alcohols as solvent. Further examples of multiple EPR/ENDOR spectra were found when the position of chirality is in the α- and β-position to a remote spin center such as the nitroxide function (see series IV and V). Finally, EPR/ENDOR spectra were obtained from nitroxide derivatives wherein the absolute configuration at the α-carbon atom was known. Distinctly different EPR/ENDOR spectra were indeed obtained for every diastereomer, thus establishing this point unequivocally. Copyright © 1999 John Wiley & Sons, Ltd.

Pulsed Electron-Nuclear Double Resonance (ENDOR) at 140 GHz

We describe a spectrometer for pulsed ENDOR at 140 GHz, which is based on microwave IMPATT diode amplifiers and a probe consisting of a TE011 cavity with a high-quality resonance circuit for variable radiofrequency irradiation. For pulsed EPR we obtain an absolute sensitivity of 3 × 109 spins/Gauss at 20 K. The performance of the spectrometer is demonstrated with pulsed ENDOR spectra of a standard bis-diphenylene-phenyl-allyl (BDPA) doped into polystyrene and of the tyrosyl radical from E. coli ribonucleotide reductase (RNR). The EPR spectrum of the RNR tyrosyl radical displays substantial g-anisotropy at 5 T and is used to demonstrate orientation-selective Davies-ENDOR.

ENDOR and ESR Studies of Radical Cations of Methyl-Substituted Benzene in Halocarbon Matrices

The radical cations of methyl-substituted benzene toluene, p-xylene, o-xylene, m-xylene, and their deuterated analogues generated in CFCl3 and CF3CCl3 matrices by X-ray irradiation at 77 K were investigated by ESR and high-resolution ENDOR spectroscopy. The ESR and ENDOR spectra in these radical cations are dominated by large axially symmetric hyperfine splitting due to methyl-group protons. The anisotropic hyperfine coupling constants (hfcc) of methyl protons and ring protons in radical cations of toluene and p-xylene were measured accurately by ENDOR spectroscopy. In the case of deuterated p-xylene and o-xylene radical cations in CF3CCl3 matrix, the hyperfine coupling constants are scaled by their magnetogyric ratio (γH/γD = 6.51). Theoretical calculations of the isotropic and dipolar hyperfine coupling constants in these radical cations are found to be in good agreement with experimental results. The experimental and theoretical calculations supported the stabilization of the 2B2g type of state for the...

The radical ions of acenaphtho[1,2-b][1,4]dithiine derivatives and acenaphtho[1,2-b][1,4]oxathiine: Solution EPR and ENDOR studies. The X-ray crystal structures of 8,9-bis(methylsulfanyl)acenaphtho[1,2-b][1,4]dithiine and its complexes with 7,7,8,8-tetracyano-p-quinodimethane (TCNQ), 2,5-dibromo-TCNQ and iodine

Syntheses of acenaphtho[1,2-b][1,4]oxathiine 4 and 8,9-bis(methylsulfanyl)acenaphtho[1,2-b][1,4]dithiine 5 are reported. The fairly persistent radical ions of acenaphtho[1,2-b][1,4]dithiine 1, acenaphtho[1,2-b][1,4]benzodithiine 2, acenaphtho[1,2-b]thieno[3,4-e][1,4]dithiine 3, and 4, i.e. 1˙+–4˙+ and 1˙––4˙–, were generated from their neutral precursors and their EPR and ENDOR spectra are reported. Oxidation to the radical cations implies an electron removal from the heteroatoms of the donor dithiine or oxathiine moiety which houses the bulk of the spin population. Accordingly, in the EPR spectra of 1˙+–4˙+, the 1H-hyperfine pattern is only 0.25–0.70 mT wide, while the 33S satellites extend over 2–3 mT. Upon reduction of 1–4 to their radical anions, the extra electron resides in the acenaphthylene π-system. Thus, the EPR spectra of 1˙––4˙– have a total width of ca. 2.2 mT with no observable 33S satellites. The g factors of the radical cations comply with the values for oxidised S-donors, whereas those of the corresponding anions are typical of reduced π-systems without heteroatoms. The crystal structure of 5 is characterised by pseudo-dimers of molecules folded by 54° along the S· · ·S vector of the 1,4-dithiine ring. The structural motif of both 5∶TCNQ and 5∶Br2TCNQ comprises mixed stacks of donor and acceptor moieties. The bond lengths in 5∶TCNQ and 5∶Br2TCNQ show minor perturbation compared with pure 5, which can be attributed to a small degree of charge transfer. The asymmetric unit of 5∶(I7–) comprises one radical cation 5˙+, two I2 molecules and one I3– anion. The radical cation, in contrast with the structures of 5, 5∶TCNQ and 5∶Br2TCNQ, shows only a minor boat-like distortion, and the radical cations form a stepwise stack, parallel to the b axis. Iodine species form an infinite zig-zag chain, running in the general direction of the c axis, i.e. normal to the stacks. The conductivity values of these complexes are low (σrt = ≤ 10–8 S cm–1).

On the electronic structure of [Pt(4,4′-X2-bipy)Cl2]0/−/2−: an electrochemical and spectroscopic (UV/Vis, EPR, ENDOR) study

A series of complexes of general formula [Pt(4,4′-X2-bipy)Cl2] (bipy = 2,2′-bipyridine; X = NH2, OEt, Me, H, Ph, Cl or CO2Me) has been prepared and their redox chemistry and UV/Vis spectroscopy examined. The half-wave potential of the first reduction process from cyclic voltammetry varies linearly with the Hammett parameter of X, and also with the MLCT maximum from UV/Vis spectroscopy. The first reduction processes of the complexes with X = OEt, Me, H, Ph, Cl or CO2Me are reversible and these complexes undergo a second quasi-reversible or irreversible reduction at potentials 580–760 mV more negative. The reduced, 17 e– species are characterised by EPR spectroscopy and the total platinum contribution (5d and 6p) to the SOMOs is calculated to be only ca. 7–12%. The bulk of the unpaired electron density is carried in the bipy ligand π* system and orientation-selective 1H ENDOR spectra of [Pt(4,4′-X2-bipy)Cl2]– (X = H or CO2Me) show that C5 and C5′ carry the greatest spin density among the four ring C(H) positions. The second reduction product of [Pt(4,4′-(CO2Me)2-bipy)Cl2] is EPR silent indicating spin-pairing of the two reduction electrons in the same orbital in this particular complex.

Modification of electron transfer from the quinone electron carrier, A(1), of Photosystem 1 in a site directed mutant D576 double right arrow L within the Fe-S-x binding site of PsaA and in second site suppressors of the mutation in Chlamydomonas reinhardti

A site directed mutant of the Photosystem I reaction center of Chlamydomonas reinhardtii has been described previously. [Hallahan et al. (1995) Photosynth Res 46: 257–264]. The mutation, PsaA: D576L, changes the conserved aspartate residue adjacent to one of the cysteine ligands binding the Fe-SX center to PsaA. The mutation, which prevents photosynthetic growth, was observed to change the EPR spectrum of the Fe-SA/B centers bound to the PsaC subunit. We suggested that changes in binding of PsaC to the PsaA/PsaB reaction center prevented efficient electron transfer. Second site suppressors of the mutation have now been isolated which have recovered the ability to grow photosynthetically. DNA analysis of four suppressor strains showed the original D576L mutation is intact, and that no mutations are present elsewhere within the Fe-SX binding region of either PsaA or PsaB, nor within PsaC or PsaJ. Subsequent genetic analysis has indicated that the suppressor mutation(s) is nuclear encoded. The suppressors retain the altered binding of PsaC, indicating that this change is not the cause of failure to grow photosynthetically. Further analysis showed that the rate of electron transfer from the quinone electron carrier A1 to Fe-SX is slowed in the mutant (by a factor of approximately two) and restored to wild type rates in the suppressors. ENDOR spectra of A1·– in wild-type and mutant preparations are identical, indicating that the electronic structure of the phyllosemiquinone is not changed. The results suggest that the quinone to Fe-SX center electron transfer is sensitive to the structure of the iron-sulfur center, and may be a critical step in the energy conversion process. They also indicate that the structure of the reaction center may be modified as a result of changes in proteins outside the core of the reaction center.