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This study comprises analyses of contents of mycotoxins, such as deoxynivalenol and zearalenone, as well as the level of oxidative stress in ears of a susceptible wheat cultivar Hanseat and cv. Arina, resistant to a pathogenic fungus Fusarium graminearum. Starting from 48 h after inoculation, a marked increase was observed in the contents of these mycotoxins in ears of wheat; however, the greatest accumulation was recorded in the late period after inoculation, i.e., during development of disease. Up to 120 h after inoculation, in ears of both wheat cultivars, the level of deoxynivalenol was higher than that of zearalenone. The susceptible cultivar was characterized by a much greater accumulation of deoxynivalenol than the resistant cultivar. At the same time, in this cultivar, in the time from 0 to 72 h after inoculation, a marked post-infection increase was observed in the generation of the superoxide radical (O2•-). Additionally, its level, at all the time points after inoculation, was higher than in the control. In wheat cv. Arina, a markedly higher level of O2- generation in relation to the control was found up to two hours after inoculation and, next, at a later time after inoculation. In turn, the level of semiquinone radicals detected by electron paramagnetic resonance (EPR) increased at later culture times, both in cv. Hanseat and Arina; however, in infested ears of wheat, it was generally lower than in the control. Analysis of disease symptoms revealed the presence of more extensive lesions in ears of a susceptible wheat cv. Hanseat than resistant cv. Arina. Additionally, ergosterol level as a fungal growth indicator was higher in ears of susceptible wheat than in the resistant cultivar. [ABSTRACT FROM AUTHOR]

Examinationof electron transfer and proton transfer reactionsof lumiflavin and proton transfer reactions of the lumiflavin radicalanion by Fourier transform ion cyclotron resonance mass spectrometryis described. From the equilibrium constant determined for electrontransfer between 1,4-naphthoquinone and lumiflavin the electron affinityof lumiflavin is deduced to be 1.86 ± 0.1 eV. Measurements ofthe rate constants and efficiencies for proton transfer reactionsindicate that the proton affinity of the lumiflavin radical anionis between that of difluoroacetate (331.0 kcal/mol) and p-formyl-phenoxide (333.0 kcal/mol). Combining the electron affinityof lumiflavin with the proton affinity of the lumiflavin radical aniongives a lumiflavin hydrogen atom affinity of 59.7 ± 2.2 kcal/mol.The ΔG298deduced from these resultsfor adding an H atom to gas phase lumiflavin, 52.1 ± 2.2 kcal/mol,is in good agreement with ΔG298foradding an H atom to aqueous lumiflavin from electrochemical measurementsin the literature, 51.0 kcal/mol, and that from M06-L density functionalcalculations in the literature, 51.2 kcal/mol, suggesting little,if any, solvent effect on the H atom addition. The proton affinityof lumiflavin deduced from the equilibrium constant for the protontransfer reaction between lumiflavin and 2-picoline is 227.3 ±2.0 kcal mol–1. Density functional theory calculationson isomers of protonated lumiflavin provide a basis for assigningthe most probable site of protonation as position 1 on the isoalloxazinering and for estimating the ionization potentials of lumiflavin neutralradicals. [ABSTRACT FROM AUTHOR]

Abstract: 1. Recent results suggest that the major flux is carried by a monomeric function, not by an intermonomer electron flow. 2. The bifurcated reaction at the Qo-site involves sequential partial processes, — a rate limiting first electron transfer generating a semiquinone (SQ) intermediate, and a rapid second electron transfer in which the SQ is oxidized by the low potential chain. 3. The rate constant for the first step in a strongly endergonic, proton-first-then-electron mechanism, is given by a Marcus–Brønsted treatment in which a rapid electron transfer is convoluted with a weak occupancy of the proton configuration needed for electron transfer. 4. A rapid second electron transfer pulls the overall reaction over. Mutation of Glu-295 of cyt b shows it to be a key player. 5. In more crippled mutants, electron transfer is severely inhibited and the bell-shaped pH dependence of wildtype is replaced by a dependence on a single pK at ~8.5 favoring electron transfer. Loss of a pK ~6.5 is explained by a change in the rate limiting step from the first to the second electron transfer; the pK ~8.5 may reflect dissociation of QH . 6. A rate constant (<103 s−1) for oxidation of SQ in the distal domain by heme b L has been determined, which precludes mechanisms for normal flux in which SQ is constrained there. 7. Glu-295 catalyzes proton exit through H+ transfer from QH , and rotational displacement to deliver the H+ to exit channel(s). This opens a volume into which Q − can move closer to the heme to speed electron transfer. 8. A kinetic model accounts well for the observations, but leaves open the question of gating mechanisms. For the first step we suggest a molecular “escapement”; for the second a molecular ballet choreographed through coulombic interactions. This article is part of a Special Issue entitled: Respiratory complex III and related bc complexes. [Copyright &y& Elsevier]

Kinetic and thermodynamic parameters of redox reactions and electron transfer between o-benzosemiquinone radicals and iron ions have been determined. The rate constant of the anion-radicals oxidation with Cu 2+ ions has been calculated. [ABSTRACT FROM AUTHOR]

The mechanism of molecular oxygen activation is the subject of controversy in the copper amine oxidase family. At their active sites, copper amine oxidases contain both a mononuclear copper ion and a protein-derived quinone cofactor. Proposals have been made for the activation of molecular oxygen via both a Cu(II)-ami-noquinol catalytic intermediate and a Cu(I)-semiquinone intermediate. Using protein crystallographic freeze-trapping methods under low oxygen conditions combined with single-crystal microspectrophotometry, we have determined structures corresponding to the iminoquinone and semiquinone forms of the enzyme. Methylamine reduction at acidic or neutral pH has revealed protonated and deprotonated forms of the iminoquinone that are accompanied by a bound oxygen species that is likely hydrogen peroxide. However, methylamine reduction at pH 8.5 has revealed a copper-ligated cofactor proposed to be the semiquinone form. A copper-ligated orientation, be it the sole identity of the semiquinone or not, blocks the oxygen-binding site, suggesting that accessibility of Cu(I) may be the basis of partitioning O2 activation between the aminoquinol and Cu(I). [ABSTRACT FROM AUTHOR]

Efficient energy conversion often requires stabilization of one-electron intermediates within catalytic sites of redox enzymes. While quinol oxidoreductases are known to stabilize semiquinones, one of the famous exceptions includes the quinol oxidation site of cytochrome bc1 (Qo), for which detection of any intermediate states is extremely difficult. Here we discover a semiquinone at the Qo site (SQo) that is coupled to the reduced Rieske cluster (FeS) via spin-spin exchange interaction. This interaction creates a new electron paramagnetic resonance (EPR) transitions with the most prominent g = 1.94 signal shifting to 1.96 with an increase in the EPR frequency from X- to Q-band. The estimated value of isotropic spin-spin exchange interaction (∣J0∣ = 3500 MHz) indicates that at a lower magnetic field (typical of X-band) the SQo-FeS coupled centers can be described as a triplet state. Concomitantly with the appearance of the SQo-FeS triplet state, we detected a g = 2.0045 radical signal that corresponded to the population of unusually fast-relaxing SQo for which spin-spin exchange does not exist or is too small to be resolved. The g = 1.94 and g = 2.0045 signals reached up to 20% of cytochrome bc1 monomers under aerobic conditions, challenging the paradigm of the high reactivity of SQo toward molecular oxygen. Recognition of stable SQo reflected in g = 1.94 and g = 2.0045 signals offers a new perspective on understanding the mechanism of Qo site catalysis. The frequency-dependent EPR transitions of the SQo-FeS coupled system establish a new spectroscopic approach for the detection of SQo in mitochondria and other bioenergetic systems. [ABSTRACT FROM AUTHOR]

Density functional theory methods were employed to elucidate the interactions between calcium ions and various o-semiquinone radicals mimicking the interactions occurring in biochemical systems. Predicted changes in the molecular and electronic structures of the radicals on Ca coordination were correlated with the changes of g tensor and compared with those exerted by Mg ions (reported by us previously). In order to broaden the insight into the differences between the Mg and Ca complexes, their relative stability was estimated on the basis of theoretically predicted Gibbs energies for the process of the complex formation. [ABSTRACT FROM AUTHOR]

As a rule, o-semiquinones decay through disproportionation leading to equimolar amounts of catechol and o-quinone products. However, the o-semiquinone 1S generated by pulse radiolysis oxidation of the eumelanin precursor 5,6-dihydroxyindole (1) decays with second-order kinetics to generate broad visible chromophores that are incompatible with the predicted absorption of 5,6-indolequinone (1Q). Using an integrated chemical, pulse radiolytic and computational approach as well as deuterium labeling, we show herein that 1S and related 5,6-dihydroxyindole semiquinones decay mainly by a free radical coupling mechanism. This conclusion was supported by the inverse kinetic isotope effect observed with deuterated 1S, the identification of unprecedented dihydrobiindole products by one-electron oxidation of 1, the good matching of simulated absorption profiles of free radical coupling products of 1S with experimental spectra, and a detailed computational analysis of the kinetics and thermodynamics of the disproportionation equilibrium and free radical coupling of 1S versus 1-1Q coupling. These results disclose, to the best of our knowledge, the first example of free radical dimerization of o-semiquinones outcompeting the classic disproportionation-driven catechol-quinone coupling and suggest that this hitherto unrecognized process may be of broader relevance than previously believed. [ABSTRACT FROM AUTHOR]

A detailed kinetic model has been developed to describe the oxidation of Cu(I) by O2 and the reduction of Cu(II) by 1,4-hydroquinone (H2Q) in the presence of O2 in 0.7 M NaCl solution over a pH range of 6.5-8.0. The reaction between Cu(I) and O2 is shown to be the most important pathway in the overall oxidation of Cu(I), with the rate constant for this oxidation process increasing with an increasing pH. In 0.7 M NaCl solutions, Cu(II) is capable of catalyzing the oxidation of H2Q in the presence of O2 with the monoanion, HQ-, the kinetically active hydroquinone form, reducing Cu(II) with an intrinsic rate constant of (5.0 ± 0.4) × 107 M-1 s-1. Acting as a chain-propagating species, the deprotonated semiquinone radical (SQ•-) generated from both the one-electron oxidation of H2Q and the one-electron reduction of 1,4-benzoquinone (BQ) also reacts rapidly with Cu(II) and Cu(I), with the same rate constant of (2.0 ± 0.5) × 107 M-1 s-1. In addition to its role in reformation of Cu(II) via continuous oxidation of Cu(I), O2 rapidly removes SQ•-, resulting in the generation of O2•-. Agreement between half-cell reduction potentials of different redox couples provides confirmation of the veracity of the proposed model describing the interactions of copper and quinone species in circumneutral pH saline solutions. [ABSTRACT FROM AUTHOR]

For the first time, an expansive study into the concentration and extended decay behavior of environmentally persistent free radicals in PM2.5 was performed. Results from this study revealed three types of radical decay—a fast decay, slow decay, and no decay—following one of four decay patterns: a relatively fast decay exhibiting a 1/e lifetime of 1-21 days accompanied by a slow decay with a 1/e lifetime of 21-5028 days (47% of samples); a single slow decay including a 1/e lifetime of 4-2083 days (24% of samples); no decay (18% of samples); and a relatively fast decay displaying an average 1/e lifetime of 0.25-21 days followed by no decay (11% of samples). Phenol correlated well with the initial radical concentration and fast decay rate. Other correlations for common atmospheric pollutants (ozone, NOx, SO2, etc.) as well as meteorological conditions suggested photochemical processes impact the initial radical concentration and fast decay rate. The radical signal in PM2.5 was remarkably similar to semiquinones in cigarette smoke. Accordingly, radicals inhaled from PM2.5 were related to the radicals inhaled from smoking cigarettes, expressed as the number of equivalent cigarettes smoked. This calculated to 0.4-0.9 cigarettes per day for nonextreme air quality in the United States. [ABSTRACT FROM AUTHOR]

The objective of this study was to analyse selected brands of fermented (red and black) and unfermented (white and green) teas for free radical content with the use of electron paramagnetic resonance (EPR) spectroscopy and for contents of flavan-3-ols by means of high-performance liquid chromatography. Analyses were also conducted for the polyphenolic profile of infusions of the analysed teas (with the Folin-Ciocalteu's method) and their antioxidant activity (in reaction with a DPPH radical) at three brewing times (5, 10 and 15 min). The obtained results showed the possibility of using rapid spectroscopic method EPR to evaluate the oxidative changes in tea leaves caused by enzymatic fermentation. The number of free radicals in teas was negatively correlated with contents of flavan-3-ols, (−)-EGCG in particular. The main signals observed in EPR spectra of teas were attributed to semiquinone radicals; however, also signals attributed to carbohydrate radicals were detected. Regarding unfermented teas, it was ascertained that teas with the highest content of flavan-3-ols, (−)-EGCG in particular, were characterised by the lowest content of semiquinone radicals and a high content of carbohydrate radicals. The group of fermented teas demonstrated to contain mainly semiquinone radicals. The total phenolic content and antioxidant activity of the tea infusions were strongly diversified depending on the kind and brand of tea as well as on the extraction time. The predominating flavan-3-ol in the analysed teas was epigallocatechin gallate (−)-EGCG, the content of which was additionally highly correlated with the antioxidant activity of the tea infusions. [ABSTRACT FROM AUTHOR]

Abstract: Electron flow within the neuronal nitric oxide synthase reductase domain (nNOSrd) includes hydride transfer from NADPH to FAD followed by two one-electron transfer reactions from FAD to FMN. We have used stopped flow spectrometry to closely monitor these electron transfer steps for both the wild type and the ΔG810 mutant of nNOSrd using a protocol involving both global analyses of the photodiode array spectral scans and curve fittings of single wavelength kinetic traces. The charge transfer complex and interflavin electron transfer events recorded at 750nm and 600nm, respectively, show the kinetics in different time frames. All electron transfer events are slow enough at 4°C to enable measurements of rate constants even for the fast charge transfer event. To our knowledge this is the first time the rate constants for the charge transfer between NADP+ and FADH2 have been determined for NOS. These procedures allow us to conclude that (1) binding of the second NADPH is necessary to drive the full reduction of FMN and; (2) charge transfer and the subsequent interflavin electron transfer have distinct spectral features that can be monitored separately with stopped flow spectroscopy. These studies also enable us to conclude that interflavin electron transfer reported at 600nm is not limiting in NOS catalysis. [Copyright &y& Elsevier]

Abstract: Humic acids (HAs) and humin (HU) complexes with Hg(II) ions were studied using EPR, FTIR and CP MAS 13C NMR spectroscopies. The analysis of the EPR spectra showed, especially for HA, a significant decrease in the g-factor value for the resulting Hg(II) radical species, as compared with that observed for the parent radicals. The concentration of the formed radical complexes was very low, precluding their detailed characterization. For this reason hydroxybenzenes and their benzoic acids were used as models of the humic substances’ radical centres able to form radical ligands towards Hg(II) ions. The resulting radicals, characterized by a g-factor lower than that of the free electron (2.0006–2.00017), were efficiently produced upon Hg(II) complexation only for the hydroxybenzenes with their OH groups in the ortho position and, what is more important, the presence of carboxylic substituents was not necessary. It is most likely that formation of the radicals characterised by low g-factor proceeded in two steps. The first is oxidation process of the hydroxybenzenes and their derivatives resulting in a typical phenolic/semiquinone type radical while in the second step the radical is complexed by the excess of Hg(II), as the result spin density of the radical is shifted towards aromatic ring leading to the observed lowering of the g parameter value. The redox character of the process starting from Hg(II) was confirmed by the formation of elemental mercury. The g-factor of the semiquinone radical obtained from 2,4,6-trihydroxybenzoic acid was not changed by Hg(II) ions coordination, as indicated by g =2.0034. [Copyright &y& Elsevier]

NADH:ubiquinone oxidoreductase (complex I) pumps protons across the membrane using downhill redox energy. The Escherichia coli complex I consists of 13 different subunits named NuoA-N coded by the nuo operon. Due to the low abundance of the protein and some difficulty with the genetic manipulation of its large ∼15-kb operon, purification of E. coli complex I has been technically challenging. Here, we generated a new strain in which a polyhistidine sequence was inserted upstream of nuoE in the operon. This allowed us to prepare large amounts of highly pure and active complex I by efficient affinity purification. The purified complex I contained 0.94±0.1 mol of FMN, 29.0 ± 0.37 mol of iron, and 1.99 ± 0.07 mol of ubiquinone/ 1 mol of complex I. The extinction coefficient of isolated complex I was 495mM -1cm-1at 274nm and 50.3mM -1cm-1 at 410 nm. NADH: ferricyanide activity was 219 ± 9.7 μmol/ min/mg by using HEPES-Bis-Tris propane, pH 7.5. Detailed EPR analyses revealed two additional iron-sulfur cluster signals, N6a and N6b, in addition to previously assigned signals. Furthermore, we found small but significant semiquinone signal(s), which have been reported only for bovine complex I. The line width was ∼12 G, indicating its neutral semiquinone form. More than 90% of the semiquinone signal originated from the single entity with P½ (half-saturation power level) = 1.85 milliwatts. The semiquinone signal(s) decreased by 60% when with asimicin, a potent complex I inhibitor. The functional role of semiquinone and the EPR assignment of clusters N6a/N6b are discussed. [ABSTRACT FROM AUTHOR]

Extracellular redox-active compounds, flavins and other quinones, have been hypothesized to play a major role in the delivery of electrons from cellular metabolic systems to extracellular insoluble substrates by a diffusion-based shuttling two-electron-transfer mechanism. Here we show that flavin molecules secreted by Shewanella oneidensis MR-1 enhance the ability of its outer-membrane c-type cytochromes (OM c-Cyts) to transport electrons as redox cofactors, but not free-form flavins. Whole-cell differential pulse voltammetry revealed that the redox potential of flavin was reversibly shifted more than 100 mV in a positive direction, in good agreement with increasing microbial current generation. Importantly, this flavin/OM c-Cyts interaction was found to facilitate a one-electron redox reaction via a semiquinone, resulting in a 103- to 105-fold faster reaction rate than that of free flavin. These results are not consistent with previously proposed redox-shuttling mechanisms but suggest that the flavin/OM c-Cyts interaction regulates the extent of extracellular electron transport coupled with intracellular metabolic activity. [ABSTRACT FROM AUTHOR]

Abstract: In photosystem II membrane fragments with oxidized cytochrome (Cyt) b559 reduction of Cyt b559 by plastoquinol formed in the membrane pool under illumination and by exogenous decylplastoquinol added in the dark was studied. Reduction of oxidized Cyt b559 by plastoquinols proceeds biphasically comprising a fast component with a rate constant higher than (10s)−1, named phase I, followed by a slower dark reaction with a rate constant of (2.7min)−1 at pH6.5, termed phase II. The extents of both components of Cyt b559 reduction increased with increasing concentrations of the quinols, with that, maximally a half of oxidized Cyt b559 can be photoreduced or chemically reduced in phase I at pH6.5. The photosystem II herbicide dinoseb but not 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) competed with the quinol reductant in phase I. The results reveal that the two components of the Cyt b559 redox reaction reflect two redox equilibria attaining in different time domains. One-electron redox equilibrium between oxidized Cyt b559 and the photosystem II-bound plastoquinol is established in phase I of Cyt b559 reduction. Phase II is attributed to equilibration of Cyt b559 redox forms with the quinone pool. The quinone site involved in phase I of Cyt b559 reduction is considered to be the site regulating the redox potential of Cyt b559 which can accommodate quinone, semiquinone and quinol forms. The properties of this site designated here as QD clearly suggest that it is distinct from the site QC found in the photosystem II crystal structure. [Copyright &y& Elsevier]

Abstract: We re-examine the pH dependence of partial processes of ubihydroquinone (QH2) turnover in Glu-295 mutants in Rhodobacter sphaeroides to clarify the mechanistic role. In more crippled mutants, the bell-shaped pH profile of wildtype was replaced by dependence on a single pK at ~8.5 favoring electron transfer. Loss of the pK at 6.5 reflects a change in the rate-limiting step from the first to the second electron transfer. Over the range of pH 6–8, no major pH dependence of formation of the initial reaction complex was seen, and the rates of bypass reactions were similar to the wildtype. Occupancy of the Qo-site by semiquinone (SQ) was similar in the wildtype and the Glu→Trp mutant. Since heme b L is initially oxidized in the latter, the bifurcated reaction can still occur, allowing estimation of an empirical rate constant <103 s−1 for reduction of heme b L by SQ from the domain distal from heme b L, a value 1000-fold smaller than that expected from distance. If the pK ~8.5 in mutant strains is due to deprotonation of the neutral semiquinone, with Q•− as electron donor to heme b L, then in wildtype this low value would preclude mechanisms for normal flux in which semiquinone is constrained to this domain. A kinetic model in which Glu-295 catalyzes H+ transfer from QH•, and delivery of the H+ to exit channel(s) by rotational displacement, and facilitates rapid electron transfer from SQ to heme b L by allowing Q•− to move closer to the heme, accounts well for the observations. [Copyright &y& Elsevier]

When the quinone, vitamin K1(VK1), is electrochemicallyreduced in aqueous-acetonitrile solutions (CH3CN with 7.22M H2O), it undergoes a two-electron reduction to form thedianion that is hydrogen-bonded with water [VK1(H2O)y2–]. EPR and voltammetryexperiments have shown that the persistent existence of the semiquinoneanion radical (also hydrogen-bonded with water) [VK1(H2O)x–•] in aqueous or organic–aqueous solutions is a result of VK1(H2O)y2–undergoing a net homogeneous electron transfer reaction (comproportionation)with VK1, and not via direct one-electron reduction ofVK1. When 1 mM solutions of VK1were electrochemicallyreduced by two electrons in aqueous-acetonitrile solutions, quantitativeEPR experiments indicated that the amount of VK1(H2O)x–•produced was up to approximately 35% of all the reduced species. In situelectrochemical ATR-FTIR experiments on sequentiallyone- and two-electron bulk reduced solutions of VK1(showingstrong absorbances at 1664, 1598, and 1298 cm–1)in CH3CN containing <0.05 M H2O led to thedetection of VK1–•with strongabsorbances at 1710, 1703, 1593, 1559, 1492, and 1466 cm–1and VK1(H2O)y2–with strong absorbances at 1372 and 1342 cm–1. [ABSTRACT FROM AUTHOR]

Despite the extensive study of the biosynthesis of the complex molecule B12 (cobalamin), the mechanism by which the lower ligand 5,6-dimethylbenzimidazole (DMB) is formed has remained something of a mystery. However, recent work has identified and characterized a DMB-synthase (BluB) responsible for the oxygen-dependent, single enzyme conversion of FMN to DMB. In this work, we have identified BluB homologs from the aerobic purple, nonsulfur, photosynthetic bacterium Rhodobacter capsulatus and the aerobic soil bacterium Bacillus megaterium and have demonstrated DMB synthesis by the use of a novel complementation assay in which a B12 deficient strain, substituted with the precursor cobinamide is recovered either by the addition of DMB or by the recombinant expression of a bluB gene. The DMB-synthetic activity of the purified recombinant BluB enzymes was further confirmed in vitro by providing the enzyme with FMNH2 and oxygen and observing the formation of DMB by HPLC. The formation of a 4a-peroxyflavin intermediate, the first step in the oxygen dependent mechanism of DMB biosynthesis, is reported here and is the first intermediate in the enzyme catalysed reaction to be demonstrated experimentally to date. The identification and characterization of an FMN-binding protein found on the cobI operon of B. megaterium, CbiY, is also detailed, revealing an FMN-containing enzyme which is able to stabilize a blue flavin semiquinone upon reduction with a 1-electron donor. [ABSTRACT FROM AUTHOR]

Photosystem II uses light to drive water oxidation and plastoquinone (PQ) reduction. PQ reduction involves two PQ cofactors, QA and QB, working in series. QA is a one-electron carrier, whereas QB undergoes sequential reduction and protonation to form QBH2. QBH2 exchanges with PQ from the pool in the membrane. Based on the atomic coordinates of the Photosystem II crystal structure, we analyzed the proton transfer (PT) energetics adopting a quantum mechanical/molecular mechanical approach. The potential-energy profile suggests that the initial PT to QB•- occurs from the protonated, D1-His252 to QB•- via D1-Ser264. The second PT is likely to occur from D1-His215 to QBH- via an H-bond with an energy profile with a single well, resulting in the formation of QBH2 and the D1-His215 anion. The pathway for reprotonation of D1-His215- may involve bicarbonate, D1-Tyr246 and water in the QB site. Formate ligation to Fe2+ did not significantly affect the protonation of reduced QB, suggesting that formate inhibits QBH2 release rather than its formation. The presence of carbonate rather than bicarbonate seems unlikely because the calculations showed that this greatly perturbed the potential of the nonheme iron, stabilizing the Fe3+ state in the presence of QB•-, a situation not encountered experimentally. H-bonding from D1-Tyr246 and D2-Tyr244 to the bicarbonate ligand of the nonheme iron contributes to the stability of the semiquinones. A detailed mechanistic model for QB reduction is presented. [ABSTRACT FROM AUTHOR]

Abstract: The nature of the electronic coupling in transition metal Donor–Acceptor and Donor–Bridge–Acceptor systems has become a subject of interest in the last decade due to the importance of electronic coupling in molecular electronics. Here we review the synthesis, structure, magnetism, and spectroscopy of donor–acceptor biradicals as they pertain to understanding the electronic origins and magnitude of the electronic coupling matrix element that figures prominently in electron transfer (transport). Stable D–A systems may be thought of as ground state analogues of charge separated states generated in photoinduced electron transfer processes and as model systems for understanding single molecule mediated electron transport between biased nanoelectrodes. The strong coupling (adiabatic) regime will be covered in this review as well as the methodology for probing the magnitude of the electronic coupling in this limit. D–A biradicals can facilitate long-range ferromagnetic exchange between localized spins mediated by delocalized electrons, and this is a new phenomenon in molecular systems. The use of the VBCI approach to develop a complete electronic structure description of strong electronic coupling in D–A and D–B–A biradicals, the origin of long-range biradical ferromagnetic exchange, and the relationship of these biradicals to molecular electronic materials will form the basis of the review. [Copyright &y& Elsevier]

Abstract: New bis-o-benzosemiquinonato tin(IV) complexes (3,6-SQ)2SnCl2, (3,6-SQ)2SnBr2 and (3,6-SQ)2SnPh2 (3,6-SQ is radical-anion 3,6-di-tert-butyl-o-benzosemiquinone) were synthesized by different methods. The reaction of tin(IV) catecholates (3,6-Cat)SnX2·2THF (Х=Cl, Br) with nitrogen(IV) oxide leads to EPR-active six-coordinated mono-o-benzosemiquinonato complexes of (3,6-SQ)SnX2(NO2)·THF type which undergo symmetrization to the corresponding diradical bis-o-benzosemiquinonato species (3,6-SQ)2SnX2. Diphenyltin(IV) catecholate (3,6-Cat)SnPh2·THF reacts with Br2, I2 and NO2 to give five-coordinated species (3,6-SQ)SnPh2X (X=Br, I, NO2) which transform to bis-(o-benzosemiquinonato)diphenyltin(IV) (3,6-SQ)2SnPh2. The molecular structures of (3,6-SQ)2SnBr2 and (3,6-SQ)2SnPh2 were determined by X-ray analysis. [Copyright &y& Elsevier]

A series of stable paramagnetic organomercury compounds containing the 4,6-di- tert-butyl- N-(2,6-diisopropylphenyl)- o-iminobenzosemiquinone ligand (imSQ) with the general formula imSQHgR (R = Ph, Et, Pr n, Bu n, Pr i, Hex c, CH 2SiMe 3, p-FPh, Fc, p-Me 2NPh) were synthesized. The molecular and electronic structures of the synthesized compounds were investigated by EPR spectroscopy and X-ray diffraction. The o-iminosemiquinone ligand shows the unusual amidophenoxy-type coordination mode. In the EPR spectra of the compounds imSQHgR, the hyperfine coupling constant of magnetic mercury isotopes ( 199,201Hg) depends on the nature of the substituent R, the temperature, and the nature of the solvent. Voltammetric investigations showed that the complex imSQHgFc undergoes three reversible one-electron redox steps related to the redox transformations of both the o-iminoquinone ligand and the metallocene moiety of the molecule. [ABSTRACT FROM AUTHOR]

Abstract: The interactions of lanthanide ions, REE(III), with semiquinone radicals naturally occurring in humic (HA) and hymatomelanic (HY) acids of different origin were studied mainly by EPR spectroscopy. Quantitative EPR analysis proved that only the semiquinone free radical concentration in both, HA and HY, was affected by interaction with some of the lanthanide ions, whereas the EPR g parameters of the formed radical–REE(III) complexes remained unchanged. The radical concentration was practically unaffected by the REE(III) ions with 4f1–4f6 electron configurations and in some degree increased by REE(III) ions with 4f8–4f14 configuration. The lanthanide playing an exceptional role was Gd(III) (4f7), which diminished strongly the free radicals concentration. The quenching ability of Gd(III) was stronger towards the radicals provided by HA than by HY, what was assigned to higher content in HA of both, oxygen-containing functional groups and conjugated aromatic rings, leading to more efficient Gd(III)–radical interaction. The semiquinone free radicals of HA and HY originated from the compost were more reactive than those from peat, as the humic acids from mature compost with less bulk density have more accessible functional groups. [Copyright &y& Elsevier]

Abstract: Grains of five genotypes of wheat (four Polish and one Finnish), differing in their tolerance to drought stress were chosen for this investigation. Electron paramagnetic resonance spectroscopy allowed observation of transition metal ions (Mn, Fe, Cu) and different types of stable radicals, including semiquinone centers, present in seed coats, as well as several types of carbohydrate radicals found mainly in the inner parts of grains. The content of paramagnetic metal centers was higher in sensitive genotypes (Radunia, Raweta) than in tolerant ones (Parabola, Nawra), whereas the Finnish genotype (Manu) exhibited intermediate amounts. Similarly, the concentrations of both types of radicals, carbohydrates and semiquinone were significantly higher in the grains originating from more sensitive wheat genotypes. The nature of carbohydrate radicals and their concentrations were confronted with the kinds and amounts of sugars found by the biochemical analyses and microscopy observations. It is suggested that some long lived radicals (semiquinone and starch radicals) occurring in grains could be indicators of stress resistance of wheat plants. [Copyright &y& Elsevier]

Quinones are essential cofactors in many physiologicalprocesses,among them proton-coupled electron transfer (PCET) in photosynthesisand respiration. A key intermediate in PCET is the monoprotonatedsemiquinone radical. In this work we produced the monoprotonated benzosemiquinone(BQH•) by UV illumination of BQ dissolved in 2-propanolat cryogenic temperatures and investigated the electronic and geometricstructures of BQH•in the solid state (80 K) usingEPR and ENDOR techniques at 34 GHz. The g-tensorof BQH•was found to be similar to that of the anionicsemiquinone species (BQ•–) in frozen solution.The peaks present in the ENDOR spectrum of BQH•wereidentified and assigned by 1H/2H substitutions.The experiments reconfirmed that the hydroxyl proton (O–H)on BQH•, which is abstracted from a solvent molecule,mainly originates from the central CH group of 2-propanol. They alsoshowed that the protonation has a strong impact on the electron spindistribution over the quinone. This is reflected in the hyperfinecouplings (hfc’s) of the ring protons, which dramatically changedwith respect to those typically observed for BQ•–. The hfc tensor of the O–H proton was determined by a detailedorientation-selection ENDOR study and found to be rhombic, resemblingthose of protons covalently bound to carbon atoms in a π-system(i.e., α-protons). It was found that the O–H bond liesin the quinone plane and is oriented along the direction of the quinoneoxygen lone pair orbital. DFT calculations were performed on differentstructures of BQH•coordinated by four, three, orzero 2-propanol molecules. The O–H bond length was found tobe around 1.0 Å, typical for a single covalent O–H bond.Good agreement between experimental and DFT results were found. Thisstudy provides a detailed picture of the electronic and geometricstructures of BQH•and should be applicable to othernaturally occurring quinones. [ABSTRACT FROM AUTHOR]

Abstract: The catecholic derivative 2,3-dihydroxybenzoic acid (2,3-DHBA or pyrocatechuic acid) represents a diversity of actions in enzymatic processes. In the present study DFT calculations (at the B3LYP/TZVP level of theory) have been performed for neutral 2,3DHBA and its dimer (models 1–1a), several semiquinone forms of 2,3-DHBA, namely the neutral (models 2–4), the monoionized (models 5–7), the di-ionized (models 8) and the dimer 7a. The more stable species in each case are those with the carboxyl group protonated. Oxygenated adducts were also constructed (models 10–15) in which the dioxygen is either H-bonded to the catecholic or carboxylic hydrogen or it is concerned to act on the intradiol or extradiol carbon atoms. The side-on placement of O2 on C(2) facilitates the intradiol C–C cleavage. Protonation of the oxygenated on C(1) adduct leads to decarboxylation of 2,3-DHBA. Isolation in the solid state and characterization by ESMS, IR, NMR, electronic spectra of the blue-green oxidized product of 2,3-DHBA (solid 1) supports the possibility of the existence of the semiquinone form or the hydroperoxide–adduct. Experimental spectroscopic data are correlated to the calculated spectroscopic parameters. In the ESMS the decarboxylation and degradation products as well a peroxo-adduct have been detected. Oxygenated species may also account for the plethora of redox signals in the cyclic voltammograms of solid 1 (in DMSO solutions). [Copyright &y& Elsevier]

Na+-translocating NADH:quinone oxidoreductase (Na+-NQR) is a component of respiratory electron-transport chain of various bacteria generating redox-driven transmembrane electrochemical Na+ potential. We found that the change in Na+ concentration in the reaction - medium has no effect on the thermodynamic properties of prosthetic groups of Na+-NQR from Vibrio harveyi, as was revealed by the anaerobic equilibrium redox titration of the enzymes EPR spectra. On the other hand, the change in Na+ concentration strongly alters the EPR spectral properties of the radical pair formed by the two anionic semiquinones of FMN residues bound to the NqrB and NqrC subunits (FMNNqrB and FMNNqrC). Using data obtained by pulse X- and Q-band EPR as well as by pulse ENDOR and ELDOR spectroscopy, the interspin distance between FMNNqrB and FMNNqrC was found to be 15.3 Å in the absence and 20.4 Å in the presence of Na+, respectively. Thus, the distance between the covalently bound FMN residues can vary by about S Å upon changes in Na+ concentration. Using these results, we propose a scheme of the sodium potential generation by Na+-NQR based on the redox- and sodium-dependent conformational changes in the enzyme. [ABSTRACT FROM AUTHOR]

The blue-light photo-reduction (conversion of oxidized flavin quinone via flavin semiquinone to fully reduced flavin hydroquinone) and dark re-oxidation of the flavins riboflavin and lumiflavin in starch (α-amylose) films was studied by absorption and luminescence spectroscopy. Blue-light photo-excitation caused an absorption, fluorescence, and phosphorescence decrease which recovered in the dark. The photo-reduction dark-oxidation cycle could be repeated. The efficiency of photo-reduction decreased with exposed excitation energy, and the speed of re-oxidation in the dark slowed down with time after excitation. The absorption did not fully recover. The fluorescence efficiency after a long time of storage in the dark increased beyond the initial flavin quinone fluorescence efficiency. Flavin photo-excitation is thought to cause starch-flavin restructuring (static fluorescence quenching center formation), enabling enhanced photo-induced starch to flavin electron transfer with subsequent flavin reduction and starch oxidation. In the dark, after light switch-off, thermal reversion of flavin reduction and starch oxidation occurred. [ABSTRACT FROM AUTHOR]

The structure and reactivity of a covalently linked catechol-ortho-benzoquinone (hemiquinone) is studied by UV-Vis and IR absorption spectroscopy. Nanosecond transient absorption spectroscopy of the hemiquinone reveals the formation of bi-radical state consisting of two semiquinone units. It is a long-lived state resulting from proton coupled electron transfer (PCET). [ABSTRACT FROM AUTHOR]

Abstract: Organic radicals are known to be an indispensable component of the humic acids (HA) structure. In HA two forms of radicals, stable (native) and short-lived (transient), are identified. Importantly, these radical forms can be easily differentiated by electron paramagnetic resonance (EPR) spectroscopy. This article provides a DFT-based insight into the electronic and molecular structure of the native radicals. The molecular models including an increase of the radical aromaticity and the hydrogen bonding between the radical and other functional groups of HA are taken under investigation. In consequence the interesting pieces of information on the structure of the native radical centers in HA are revealed and discussed, especially in terms of differences between the electronic structure of the native and transient forms. [Copyright &y& Elsevier]

Based on newly synthesized hydroquinone monomers, two series of halogenated polyaryletherketones, PEEKs and PEEKKs, were synthesized through nucleophilic aromatic substitution polycondensations. The chemical structures of the monomers and the polymers were carefully characterized using Fourier transform infrared and 1H-NMR spectroscopies. Some interesting results about the relationships between the structure and properties of these polymers having comparable chemical structures were revealed for the first time. Despite the similar chemical structure of –F, –Cl and –Br substituted polyaryletherketones, they exhibited different thermal behaviors. The thermogravimetric analysis results showed that chlorinated PEEKK had the best thermal stability and its limited oxygen index (LOI) value was 42.80. The results indicated some of them will be promising high-performance polymers. [ABSTRACT FROM AUTHOR]

Selective 15N isotope labeling of the cytochrome bo3ubiquinol oxidase from Escherichia coli with auxotrophs was used to characterize the hyperfine couplings with the side-chain nitrogens from residues R71, H98, and Q101 and peptide nitrogens from residues R71 and H98 around the semiquinone (SQ) at the high-affinity QH site. The two-dimensional HSEEM (HYSCORE) data have directly identified NE of R71 as an H-bond donor carrying the largest amount of unpaired spin density. In addition, weaker hyperfine couplings with the side-chain nitrogens from all residues around the SQ were determined. These hyperfine couplings reflect a distribution of the unpaired spin density over the protein inTF the SQ state of the QH site and the strength of interaction with different residues. The approach was extended to the virtually inactive D75H mutant, where the intermediate SQ is also stabilized. We found that Nε of a histidine residue, presumably H75,carries most of the unpaired spin density instead of Nε of R71, as in wild-type bo3. However, the detailed characterization of the weakly coupled 15N atoms from selective labeling of R71 and Q101 in D7SH was precluded by overlap of the ISN lines with the, much stronger ~1.6 MHz line from the quadrupole triplet of the strongly coupled 14Nε atom of H75. Therefore, a reverse labeling approach, in which the enzyme was uniformly labeled except for selected amino acid types, was applied to probe the contribution of R71 and Q101 to the 15N signals. Such labeling has shown only weak coupling with ail nitrogens of R71 and Q101. We utilize density functional theory-based calculations to model the available information about 1H, 15N, and 13C hyperfine couplings for the QH site and to describe the protein-substrate interactions in both enzymes. In particular, we identify the factors responsible for the asymmetric distribution of the unpaired spin density and ponder the significance of this asymmetry to the quinone's electron transfer function. [ABSTRACT FROM AUTHOR]

Abstract: A complete study of Anabaena flavodoxin in the neutral semiquinone state by means of the EPR pulse technique HYSCORE is here presented. The results provide new information about the hyperfine interactions of the unpaired electronic spin and the nuclei in the isoalloxazine ring. This allows a better knowledge of the electronic structure of the neutral flavin radical within the protein. Combination of these results with other previously obtained by using other EPR related techniques allowed producing a very precise mapping of the flavin spin distribution in the neutral semiquinone state. This information can be very useful for determining the relationship between the electronic structure and mechanisms in flavoproteins. An experimental protocol for measuring the electronic structure details available to date is suggested. [Copyright &y& Elsevier]

Little is known about enzymatic quinone-quinol interconversions in the lipid membrane when compared with our knowledge of substrate transformations by globular enzymes. Here, the smallest example of a quinol dehydrogenase in nature, CymA, has been studied. CymA is a monotopic membrane tetraheme c-type cytochrome belonging to the NapC/NirT family and central to anaerobic respiration in Shewanella sp. Using protein-film electrochemistry, it is shown that vesicle-bound menaquinone-7 is not only a substrate for this enzyme but is also required as a cofactor when converting other quinones. Here, we propose that the high concentration of quinones in the membrane negates the evolutionary pressure to create a high affinity active site. However, the instability and reactivity of reaction intermediate, semiquinone, might require a cofactor that functions to minimize damaging side reactions. [ABSTRACT FROM AUTHOR]

The membrane-bound alcohol dehydrogenase of Gluconacetobacter diazotrophicus contains one pyrroloquinoline quinone moiety ( PQQ), one [2 Fe-2 S] cluster, and four c-type cytochromes. Here, we describe a novel and inactive enzyme. ADHi, similarly to ADHa, is a heterodimer of 72- and 44-kDa subunits and contains the expected prosthetic groups. However, ADHa showed a threefold molecular mass as compared to ADHi. Noteworthy, the PQQ, the [2 Fe-2 S] and most of the cytochromes in purified ADHi is in the oxidized form, contrasting with ADHa where the PQQ-semiquinone is detected and the [2 Fe-2 S] cluster as well as the cytochromes c remained fully reduced after purification. Reduction kinetics of the ferricyanide-oxidized enzymes showed that while ADHa was brought back by ethanol to its full reduction state, in ADHi, only one-quarter of the total heme c was reduced. The dithionite-reduced ADHi was largely oxidized by ubiquinone-2, thus indicating that intramolecular electron transfer is not impaired in ADHi. The acidic pH of the medium might be deleterious for the membrane-bound ADH by causing conformational changes leading to changes in the relative orientation of heme groups and shift of corresponding redox potential to higher values. This would hamper electron transfer resulting in the low activity observed in ADHi. [ABSTRACT FROM AUTHOR]

Abstract: The ubihydroquinone: cytochrome c oxidoreductase, or cytochrome bc 1, is a central component of photosynthetic and respiratory energy transduction pathways in many organisms. It contributes to the generation of membrane potential and proton gradient used for cellular energy production (ATP). The three-dimensional structures of cytochrome bc 1 indicate that its two monomers are intertwined to form a symmetrical homodimer. This unusual architecture raises the issue of whether the monomers operate independently, or function cooperatively during the catalytic cycle of the enzyme. In this review, recent progresses achieved in our understanding of the mechanism of function of dimeric cytochrome bc 1 are presented. New genetic approaches producing heterodimeric enzymes, and emerging insights related to the inter monomer electron transfer between the heme b cofactors of cytochrome bc 1 are described. [Copyright &y& Elsevier]

Abstract: Pectic acids participate in the transport of heavy metal ions in the root apoplasm by establishing interactions that can lead to their partial or total immobilization. The ions accumulated can be mobilized by phenolic compounds and organic acids of the root exudates. In this context, we tested, in aqueous phase, the ability of malic acid and esculetine (ESC) to mobilize the Cu(II) ions accumulated in a Ca-polygalacturonate matrix (Ca-PGA) used as a model of the root apoplasm. The results show that at pH 5.0 and 6.0 malic acid mobilizes about 22% and 34% of the Cu(II) accumulated, respectively, whereas ESC about 12% and 25%. ESC was found to cause the reduction of Cu(II) to Cu(I) with formation of ESC oxidation products. The study of the Cu(II)–ESC binary system evidenced that one molecule of ESC reduces one Cu(II) ion with formation of semiquinonic radicals that couple to form two dimers. The Cu(II) reduction by ESC was found faster in the presence of malic acid. [Copyright &y& Elsevier]

Abstract: 2-hydroxy-1,4-naphthoquinone; Lawsone (Lw) is a natural compound found in henna leaves. The reaction of lawsone with ‘Na’ metal (Lw-1), CH3COONa (Lw-2), NaOH (Lw-3), KOH (Lw-4), K2CO3 (Lw-5) and Tris(hydroxymethyl)aminomethane (Lw-6) were studied. Red orange solids obtained for Lw-1 to Lw-6 are characterized by Elemental Analysis, FTIR, 1HNMR and EPR studies. The results reveal the coordination of alkali metals ‘Na’ and ‘K’ to lawsone anion. The single crystal X-ray structure of Lw-6 was solved and it crystallizes in triclinic space group P-1 with extensive hydrogen bonding network of CO, NO and OO between cations and anions. Polycrystalline powder X-band EPR spectra of Lw-1 to Lw-5 shows signals ∼2.004 at 133K, while Lw-6 is EPR silent. The naphthosemiquinone (NSQ − ) radical formed in Lw-2 to Lw-5, is due to disproportion reaction of catechol and naphthoquinone. [Copyright &y& Elsevier]

Polychlorinated biphenyls (PCBs) can be metabolized toreactivemetabolites, such as PCB semiquinone radical anions (SQ•–), whose structure and role in PCB-induced toxicity are difficultto investigate due to their relative instability. The unrestrictedUB3LYP/6-311G** method was used to investigate several molecular descriptorsof the syn- and anti-like conformation of SQs•–. The bond lengths and angles of the quinone moiety of the SQs•–were in between the values reported for PCBquinones and hydroquinones, which is consistent with the distributionof the α highest occupied molecular orbital (α-HOMO).The dihedral angles between the two ring systems increased in thepresence of orthochlorine substituents and weresmaller compared to the corresponding PCB quinones. The ground-stateenergies indicate that the anti-like conformation of the SQs•–is more favorable than the syn-like conformation. Molecular descriptorused for modeling of quantitative structure–activity relationshipsdisplayed some dependence on the conformation. These findings suggestthat SQs•–in both the syn- and antilikeconformation may interact differently with target molecules, whichmay have implications for the toxicity of PCBs. [ABSTRACT FROM AUTHOR]

Semiquinone-bridged bisdithiazolyls 3 represent a new class of resonance-stabilized neutral radical for use in the design of single-component conductive materials. As such, they display electrochemical cell potentials lower than those of related pyridine-bridged bisdithiazolyls, a finding which heralds a reduced on-site Coulomb repulsion U. Crystallographic characterization of the chloro-substituted derivative 3a and its acetonitrile solvate 3a∙MeCN, both of which crystallize in the polar orthorhombic space group Pna21, revealed the importance of intermolecular oxygen-to-sulfur (CO⋯SN) interactions in generating rigid, tightly packed radical π-stacks, including the structural motif found for 3a∙MeCN in which radicals in neighboring π-stacks are locked into slipped-ribbon-like arrays. This architecture gives rise to strong intra- and interstack overlap and hence a large electronic bandwidth W. Variable-temperature conductivity measurements on 3a and 3a∙MeCN indicated high values of σ(300 K) (>10-3 S cm-1) with correspondingly low thermal activation energies Eact, reaching 0.11 eV in the case of 3a∙MeCN. Overall, the strong performance of these materials as ƒ = ½ conductors is attributed to a combination of low U and large W. Variable-temperature magnetic susceptibility measurements were performed on both 3a and 3a∙MeCN. The unsolvated material 3a orders as a spin-canted antiferromagnet at 8 K, with a canting angle ϕ = 0.14° and a coercive field Hc = 80 Oe at 2 K. [ABSTRACT FROM AUTHOR]

Linear free-energy relationships for a series of functionalized semiquinone ligands and their MnII- and CuIIhydro-tris(3-cumenyl-5-methylpyrazolyl) borate complexes were examined. Quinone-semiquinone cycle half-wave reduction potentials and semiquinone hydrogen hyperfine coupling constants (aH) were determined and their correlation with Hammett σ parameters reported. A new σ parameter, σaH, has been proposed. MnII and CuII metal complex metal-ligand charge transfer and n → π* UV transitions were found to be modulated by substituents. Satisfactory Hammett correlations between UV transitions and various σ values have been determined and compared in a number of instances. Copyright © 2011 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

An electronic and vibrational spectroscopic analysis of p-coumaric acid (HCou) and its deprotonated species was performed by UV–vis and Raman, respectively, and the results were supported by density functional theory (DFT) calculations. Electronic UV–vis spectral data of HCou solutions show that the deprotonation of the carboxyl group (Cou−) leads to a blue shift of the lowest energy electronic transition in comparison to the neutral species, whereas the subsequent deprotonation of the phenolic moiety (Cou2−) carries out to a more delocalized chromophore. The DFT geometric parameters calculations suggest that the variation in the electronic delocalization for the three organic species is due to different contribution of a quinoid structure that is significantly distorted in the case of Cou2−. The Raman data of HCou and its sodium salts show that the main spectral features that allow to differentiate the three organic species are those involving the styrene ν(C=C)sty vibration at 1600 cm−1 region. Even though the Raman spectra of the sodium salts of Cou− and Cou2− anions show subtle differences, the appearing of a band at ca. 1598 cm−1 in the Na2Cou spectrum, assigned to a mode involving the carboxylate asymmetric stretching, νas(COO), and the styrene stretching, ν(C=C)sty, is quite characteristic, as confirmed by the theoretical Raman spectrum. Considering that p-coumaric acid is an archetypical phenolic compound with several biological activities that essentially depend upon the medium pH, Raman spectroscopy results reported in this work can provide a proper way to characterize such important phytochemical compound in different protonation states. In order to complement the characterization of the sodium salts, X-ray diffraction (XRD) and thermal analysis were performed. [Copyright &y& Elsevier]

Electronic structure calculations are performed on models of the phyllosemiquinone (PhSQ) free radical in the A1Aand A1Bsites of photosystem I. Partial geometry optimization of each site is performed, and from the resultant geometry spin densities and hyperfine couplings are calculated. We exploit the ONIOM methodology to progressively build up a model of the A1Asite and monitor the effect on the spin density distribution of the PhSQ and its hyperfine couplings. For the A1Asite, we show that while the O1 atom of the PhSQ is not involved in direct hydrogen bonding, the 17O anisotropic hyperfine coupling for this position is sensitive to interactions with neighboring groups, especially Trp A697 and Phe A689. The results obtained are in agreement with experimental determinations which indicate small differences in 17O hyperfine couplings for both oxygen atoms. Good agreement between calculated and experimental 1H and 13C hyperfine couplings is also found. In addition, we find that a significant 14N isotropic coupling of 1.4 MHz is calculated for the peptide NH group of Leu A722. The 14N isotropic hyperfine coupling obtained for the indole nitrogen atom of Trp A697 is calculated to be zero in disagreement with a previous experimental assignment. The spin density distribution of the PhSQ in the A1Bsite is calculated to be very similar to that in the A1Asite. The presence of just one relatively weak hydrogen bond to the photosytem I quinone is proposed to contribute substantially to its relatively low redox potential when compared with the more strongly hydrogen bonded quinone acceptors present in type II reaction centers. [ABSTRACT FROM AUTHOR]