Electron Nuclear Double Resonance Studies Research Articles

ENDOR study of the chlorinated thiocytosine radical in a crystal matrix

Interaction of protons with the unpaired electron in the σ2σ*1 three-electron S∴Cl bond has been investigated by electron nuclear double resonance (ENDOR) spectroscopy. The S∴Cl bond in the chlorinated thiocytosine radical, imbedded in the crystal lattice of cytosine hydrochloride, interacts significantly with at least seven protons. Five of these interactions have been analyzed. From the nature of the coupling tensors, it is concluded that three couplings are due to protons being constituents of the chlorine–thiocytosine radical, and the remaining two are due to protons belonging to neighboring cytosine molecules. The nature of the dipolar component of the coupling tensors is not consistent with the location of the entire electron spin density in the S∴Cl bond. From isotropic as well as anisotropic hyperfine coupling tensor components, it is concluded that the unpaired electron is somewhat delocalized over the radical, with a major portion being concentrated in the C(2)–S–Cl region but with a significant fraction occupying a π* molecular orbital extending over the cytosine ring atoms. Density functional theory molecular orbital calculations support such a kind of spin distribution. The data also suggest that the radical remains protonated at both the N(1) and N(3) positions.

Continuous Wave and Pulse EPR and ENDOR Study of Oxygenated Cobalt(II) Heme Model Systems

Continuous wave (CW) and pulse electron paramagnetic resonance (EPR), and electron nuclear double resonance (ENDOR) techniques were used to study frozen solutions of (oxyCo)TPP(L) (TPP = tetraphenylporphyrin, L = pyridine, 1-methylimidazole). By using a combination of CW EPR at X-, Q-, and W-band and Davies-ENDOR at Q-band, the g and cobalt hyperfine matrices and their corresponding principal axes were, for the first time, determined in detail. The variation of the g values as a function of temperature was followed by Q-band CW EPR. The existing ambiguity about the assignment of one of the g principal axes to the O−O bound direction was analyzed in detail. By studying the different proton interactions with Mims-ENDOR at X-band, a considerable solvent effect was found. With hyperfine sublevel correlation (HYSCORE) at X-band and two-pulse ESEEM (electron spin-echo envelope modulation) at S-band, the hyperfine and nuclear quadrupole interactions of the nitrogen nuclei of the axial ligand and the TPP ligand w...

Recruitment of a Foreign Quinone into the A1 Site of Photosystem I: II. STRUCTURAL AND FUNCTIONAL CHARACTERIZATION OF PHYLLOQUINONE BIOSYNTHETIC PATHWAY MUTANTS BY ELECTRON PARAMAGNETIC RESONANCE AND ELECTRON-NUCLEAR DOUBLE RESONANCE SPECTROSCOPY

Electron paramagnetic resonance (EPR) and electron-nuclear double resonance studies of the photosystem (PS) I quinone acceptor, A(1), in phylloquinone biosynthetic pathway mutants are described. Room temperature continuous wave EPR measurements at X-band of whole cells of menA and menB interruption mutants show a transient reduction and oxidation of an organic radical with a g-value and anisotropy characteristic of a quinone. In PS I complexes, the continuous wave EPR spectrum of the photoaccumulated Q(-) radical, measured at Q-band, and the electron spin-polarized transient EPR spectra of the radical pair P700(+) Q(-), measured at X-, Q-, and W-bands, show three prominent features: (i) Q(-) has a larger g-anisotropy than native phylloquinone, (ii) Q(-) does not display the prominent methyl hyperfine couplings attributed to the 2-methyl group of phylloquinone, and (iii) the orientation of Q(-) in the A(1) site as derived from the spin polarization is that of native phylloquinone in the wild type. Electron spin echo modulation experiments on P700(+) Q(-) show that the dipolar coupling in the radical pair is the same as in native PS I, i.e. the distance between P700(+) and Q(-) (25.3 +/- 0.3 A) is the same as between P700(+) and A(1)(-) in the wild type. Pulsed electron-nuclear double resonance studies show two sets of resolved spectral features with nearly axially symmetric hyperfine couplings. They are tentatively assigned to the two methyl groups of the recruited plastoquinone-9, and their difference indicates a strong inequivalence among the two groups when in the A(1) site. These results show that Q (i) functions in accepting an electron from A(0)(-) and in passing the electron forward to the iron-sulfur clusters, (ii) occupies the A(1) site with an orientation similar to that of phylloquinone in the wild type, and (iii) has spectroscopic properties consistent with its identity as plastoquinone-9.

Q-band ENDOR (electron nuclear double resonance) of the high-affinity ubisemiquinone center in cytochrome bo3 from Escherichia coli.

Electron nuclear double resonance (ENDOR) was performed on the protein-bound, stabilized, high-affinity ubisemiquinone radical, QH*-, of bo3 quinol oxidase to determine its electronic spin distribution and to probe its interaction with its surroundings. Until this present work, such ENDOR studies of protein-stabilized ubisemiquinone centers have only been done on photosynthetic reaction centers whose function is to reduce a ubiquinol pool. In contrast, QH*- serves to oxidize a ubiquinol pool in the course of electron transfer from the ubiquinol pool to the oxygen-consuming center of terminal bo3 oxidase. As documented by large hyperfine couplings (>10 MHz) to nonexchangeable protons on the QH*- ubisemiquinone ring, we provide evidence for an electronic distribution on QH*- that is different from that of the semiquinones of reaction centers. Since the ubisemiquinone itself is physically nearly identical in both QH*- and the bacterial photosynthetic reaction centers, this electronic difference is evidently a function of the local protein environment. Interaction of QH*- with this local protein environment was explicitly shown by exchangeable deuteron ENDOR that implied hydrogen bonding to the quinone and by weak proton hyperfine couplings to the local protein matrix.

Magnetic resonance study of Rh complexes in AgCl microcrystals

X-ray or UV irradiation at room temperature of Rh 3+ doped AgCl emulsion powders leads to the production of three paramagnetic Rh 2+ related centres, labeled R4, R5 and R6. A combined X and Q band electron paramagnetic resonance (EPR) and electron nuclear double resonance (ENDOR) study allowed the determination of a nearly complete structural model for these centres. In the X band ENDOR spectra of R4 and R5 interactions of the unpaired electron with nearby protons have been identified, indicating that for these centres Cl − ligands have been exchanged by H 2O or OH −. The R6 centre, identified as a (RhCl 6) 4− complex, has been found to be fundamentally different from the dominant centre in large Rh 2+ doped AgCl single crystals grown from the melt. The results are compared with recent work by other reseachers in the same field.

Decomposition study of the electron paramagnetic resonance spectrum of irradiated alanine

Recent Electron Paramagnetic Resonance (EPR) studies on alanine powders as a function of irradiation dose and temperature on the one hand and single crystal Electron Nuclear DOuble Resonance (ENDOR) studies on the other hand, showed the presence of at least three radicals contributing to the total alanine EPR spectrum. The latter spectrum obtained after irradiation at room temperature (RT), is dominated by the well-known stable-alanine-radical (SAR) CH 3C HCOO −, also denoted R1. Appropriate heating of irradiated alanine causes the relative contribution of R1 to decrease, resulting in a spectrum mainly caused by the H-abstraction radical CH 3C (NH 3 +)COO −, denoted R2. Although the EPR spectrum of these two radicals could be satisfactorily simulated, their influence on dose reconstruction has not been reported yet. Therefore, a detailed Maximum Likelihood Common Factor Analysis (MLCFA) study has been performed on EPR spectra from polycrystalline alanine samples, after irradiation and heat treatments. Conclusions concerning the number of contributing radicals and their influence on the RT irradiated alanine EPR spectrum will be made.

Structure of dimeric radical cations of benzene and toluene in halocarbon matrices: an EPR, ENDOR and MO study

The structure of dimeric cations of benzene and toluene formed in X-irradiated halocarbon matrices containing relatively high concentration of the solutes was investigated. EPR and ENDOR (electron nuclear double resonance) spectra of these dimer cations were observed and accurate values of the hf coupling constants were obtained. The ENDOR spectrum of the dimeric radical cation of benzene, (C6H6)2+, exhibited hf couplings due to twelve equivalent protons and the isotropic coupling (aiso) was almost one-half of that in the monomer cation. ENDOR transitions with a rhombic symmetry were observed in a CFCl3 matrix, whereas clear axially symmetric transitions with ∣A‖∣<∣A⊥∣ were obtained in CF3CCl3 even at 50 K. The rhombic dipolar coupling tensor was used as a parameter to evaluate the distance between the two partner rings. As regards the toluene dimer cation, (CH3C6H5)2+, ENDOR transitions of the CH3 and H(4,4′) protons were observed. The isotropic hf coupling of the CH3 protons deviated even more strongly from the half value, being rather close to one-third of the value of the monomer. The hf coupling of the H(4,4′) protons was almost half the coupling of the monomer. It was suggested that the anomalous hf couplings of the CH3 protons were due to the interaction between the two rings through the σ-bond. Density functional theory (DFT) calculations were employed to obtain the optimized geometries and hf coupling tensors and suggested sandwich structures in both dimers. Furthermore, the distances between the two rings and the anomalous CH3 protons hf coupling in the (CH3C6H5)2+ were successfully evaluated.

ENDOR Studies of Pyruvate:Ferredoxin Oxidoreductase Reaction Intermediates

Electron−nuclear double resonance (ENDOR) studies of radical intermediates formed by the oxidative decarboxylation of pyruvate by pyruvate:ferredoxin oxidoreductase were carried out to characterize their electronic structure and elucidate aspects of the recently proposed catalytic mechanism (Menon, S.; Ragsdale, S. W. Biochemistry 1997, 36, 8484−8494). The EPR spectrum of the PFOR/pyruvate adduct at 4 K displays a narrow resonance centered at g = 2.008 that has been attributed to a hydroxyethyl thiamine pyrophosphate (HE-TPP) radical. This spectral feature is superimposed on a broad, complex line shape characteristic of magnetically coupled [Fe4S4] clusters. The ENDOR spectrum at g = 2.008 reveals a broad peak with a complex line shape that can be analyzed, assuming that it arises from a composite of two axially symmetric proton hyperfine couplings. The principle coupling values for these two hyperfine tensors were: A∥(1) = 18.9 MHz, A⊥(1) = 12.6 MHz; and A∥(2) = 20.3 MHz, A⊥(2) = 14.9 MHz. The assignment of these features to the methyl protons of the pyruvate substrate was made using isotopic substitution. The temperature independence of these 1H ENDOR line shapes from 4 to 200 K indicates that the methyl group of pyruvate undergoes rapid rotation even at 4 K. The ENDOR spectrum at g = 2.008 also shows a pair of derivative peaks centered about the 31P Larmor frequency that are assigned to a weak hyperfine coupling with the phosphorus nuclei of the TPP cofactor. Two models for the electronic structure of the radical intermediate are discussed. A σ radical model which postulates a pyruvate-derived acetyl-type radical where little unpaired spin density resides on the TPP cofactor, and a π radical model that calls for more extensive delocalization of the unpaired electron spin over the HE-TPP framework. Both models require association of the radical center with the pyrophosphate group of TPP to interpret the observed 31P hyperfine coupling.

EPR and ENDOR studies of NOx and Cu2+ in zeolites: bonding and diffusion

The diffusion and bonding of NOx (x=1, 2) and Cu2+ species in zeolites are reviewed, based mainly on our own research. The molecular motion of adsorbed NO2 has been examined with EPR in several zeolite samples and analyzed using the slow-motional EPR theory. In X- and Y-type zeolites the broadening of the spectra with temperature could be analyzed by simulations using a rotational diffusion model in agreement with earlier results in Vycor glass and Cu-metal. For the diffusion of NO2 in Na-mordenite and Na-ZSM-5 the broadening of the spectra with increased temperature could be better simulated with the Heisenberg type of spin exchange model. The exchange was attributed to the interaction between NO2 molecules diffusing along the zeolite channels. In Na-ZSM-5 the spin exchange rate increased rapidly with an increasing Si/Al ratio of the zeolite. The effect was attributed to the hindrance against diffusion by Na+, the amount of which increases with a decreased Si/Al ratio. The rates increased with increasing number of H2O molecules adsorbed on the zeolite because of weaker interaction between NO2 and the surface. A more detailed slow-motional analysis indicated that at each temperature a distribution of diffusion rates occurs. NO adsorbed on Na-A zeolite was found to be present as monomer at low pressure ( 100 mbar). The triplet was absent in calcium ion-exchanged A-type zeolite indicating that the NO–NO species depends on Na+ for its stability. The detailed structure of a Cu2+ complex with water molecules in Cu-ZSM-5 zeolite was characterised with ENDOR at 4 K. The complex has an axial distorted octahedral structure with two water molecules at a longer distance in axial position and four molecules in the equatorial positions. The Cu2+ complexes with ammonia and deuterated ammonia have been investigated with electron nuclear double resonance (ENDOR). Simulations of the ENDOR spectra with 1H, 2H and 14N hyperfine interactions have been undertaken. The analysis indicates that the complex has a square planar structure with four ammonia ligands.

Electron nuclear double resonance study of pure and aquated rhodium complexes in silver chloride emulsions

Detailed information has been obtained on the structure of pure and aquated complexes in AgCl using multifrequency EPR and powder ENDOR spectroscopy. Because of the anisotropy of the centre g matrix it was possible to selectively excite centres with a specific angle between the z axis of the g matrix and the magnetic field axis. As a result, almost single-crystalline-like ENDOR spectra were obtained and the angular dependence of the ENDOR spectra could be obtained by scanning the magnetic field over the range of the EPR spectra. Three different centres were detected that are distinguished by different thermal stabilities, different parameters of the g matrix and the hyperfine matrices of the central Rh nucleus and ligand H, Cl and Ag nuclei. A pure centre without ligands could only be generated in an emulsion where great care was taken to prevent the aquation of the dopant ion in the dopant solution and during the precipitation. This centre had the highest thermal stability. In an emulsion prepared conventionally two centres were found that are associated with one and two ligands bound to the ion in the plane perpendicular to the g matrix z axis. Using the results of the EPR and ENDOR analysis in combination with the results of a total energy calculation two models are proposed that contain the positions of the ligands and the charge compensating vacancies.

Electron paramagnetic resonance and electron nuclear double resonance of NaCl:Rh2+: A comparison between similarly doped Bridgman- and solution-grown single crystals

In a recent electron paramagnetic resonance (EPR) and electron nuclear double resonance (ENDOR) study several new rhodium related centres in a Bridgman-grown crystal were reported. For two centres a detailed microscopic model was established. The primary Rh2+ centre was identified as a [RhCl6]4− complex with a single charge compensating Na+ vacancy in a next nearest neighbour position. A second centre was identified as a rhodium dimer. Two other Rh2+ centres, with apparent axial symmetry, remained unidentified. In the present study a detailed comparison between Rh2+ defects in Bridgman- and solution-grown NaCl crystals is presented. One of the previously unidentified Rh2+ centres is shown not to be axial but to have orthorhombic-I symmetry and is identified as a cis-[RhCl6]4− complex with two next nearest neighbour Na+ vacancies. For the primary centre additional ENDOR interactions confirm the model mentioned above. Finally a comparison is made between the temperature stability of the primary and orthorhombic-I centre in Bridgman-grown and similarly doped solution-grown NaCl crystals.

A Pulsed Electron Nuclear Double Resonance Study of the Lewis Acid Site−Nitric Oxide Complex in Zeolite H-ZSM-5

ADVERTISEMENT RETURN TO ISSUEPREVCommunicationNEXTA Pulsed Electron Nuclear Double Resonance Study of the Lewis Acid Site−Nitric Oxide Complex in Zeolite H-ZSM-5Andreas Pöppl, Thomas Rudolf, and Dieter MichelView Author Information Fakultät für Physik und Geowissenschaften Universität Leipzig, Linnéstrasse 5 D-04103 Leipzig, Germany Cite this: J. Am. Chem. Soc. 1998, 120, 19, 4879–4880Publication Date (Web):May 2, 1998Publication History Received9 December 1997Published online2 May 1998Published inissue 1 May 1998https://doi.org/10.1021/ja9741685Copyright © 1998 American Chemical SocietyRIGHTS & PERMISSIONSArticle Views215Altmetric-Citations29LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit Read OnlinePDF (62 KB) Get e-AlertsSUBJECTS:Adsorption,Aluminum,Chemical structure,Electron paramagnetic resonance spectroscopy,Zeolites Get e-Alerts

Pulsed ENDOR and ESEEM Study of [Bis(maleonitriledithiolato)nickel](-): An Investigation into the Ligand Electronic Structure.

By a combination of Q-band pulsed ENDOR (electron nuclear double resonance) and X-band ESEEM (electron stimulated echo envelope modulation) techniques, we have determined the hyperfine tensors for ethylene (C1) and cyano (C2) carbons and N, of [Ni(mnt)(2)](-), along with the quadrupole tensor for nitrogen. These measurements give pi electron spin densities of rho(C1) approximately 0.03 in the C1 2p(z)() orbital, rho(C2) < 0.003, rho(N) approximately 0.01, such that in total, approximately 0.15 of the spin resides on the ligand atoms C and N, while the rest resides in the NiS(4) core, giving rho(NiS(4)(-)) = 0.85. These results are compared with extended Hückel and density functional (BLYP) MO calculations, as well as with Xalpha calculations reported earlier.

Electron Paramagnetic Resonance and Electron Nuclear Double Resonance Studies of X-Irradiated Crystals of Cytosine Hydrochloride. Part I: Free Radical Formation at 10 K after High Radiation Doses

Anhydrous single crystals of cytosine hydrochloride (protonated at N3) have been X-irradiated at 10 K and studied using K-band EPR, ENDOR and FSE spectroscopy. At least seven radicals were present at 10 K after X irradiation with a dose of about 150 kGy. Two different protonation states of the one-electron reduced cytosine cation were observed: an amino-protonated species (R1) and the pristine one-electron reduced species (R2) with zero net charge. Apparently three deprotonated versions of the one-electron oxidized cytosine cation were formed: the amino-deprotonated cation (R3), an N3-deprotonated cation (R4) and an N1-deprotonated cation (R5). Finally, two products formed by net hydrogen addition to the cytosine base were observed: a C5 hydrogen-addition radical (R6) and a C6 hydrogen-addition radical (R7). The crystalline lattice of cytosine hydrochloride is characterized in part by a cytosine base initially protonated at the N3-position, thus forming a cytosine base cation, and in part by an extended network of hydrogen bonding involving the chlorine anions. Proton transfer properties of pristine one-electron oxidation and reduction base products in this lattice are discussed and are suggested as explanations of the unusual multitude of positions for deprotonation of the one-electron oxidized species as well as for the two protonation states of the reduction product observed. The magnetic parameters for the amino-protonated species R1 agree well with those extracted from previous studies of cytosine derivatives in frozen solutions and in various glasses.

Electron Spin Resonance, Electron-Nuclear Double Resonance and General Triple Resonance Studies on the Contributions to Line Broadening of CO2- in A-type Carbonated Apatites and Biocarbonates

Electron spin resonance (ESR), electron-nuclear double resonance (ENDOR) and general triple resonance were combined to study linewidth differences observed in the ESR spectra of CO2- species in calcium carbonate and carbonated apatite. In the latter compound, the motion and a distribution of g-factors (g-strain) are dominant contributions to the observed linewidth. The linewidth temperature behavior follows previous models for CO2- rotating in calcite. In apatite, these radicals rotate more freely and movement is not restricted to gxx-gzz plane. In carbonates theoretical fit of the linewidth at 9.5 and 34.0 GHz shows the existence of two orthorhombic spectra with slightly different g values and linewidths, which contribute to the observed linewidth difference between narrow and broad line species. Superhyperfine interactions with protons of water molecules, located probably near the second and the third nearest neighbor lattice sites, are responsible for minor contributions to line broadening of CO2- spectrum.

Electronic structure of the deep boron acceptor in boron-doped6H-SiC

A high-frequency (95 GHz) and conventional-frequency (9.3 GHz) pulsed electron paramagnetic resonance and electron-nuclear double resonance (ENDOR) study is reported on the deep boron acceptor in $6H$-SiC. The results support a model in which the deep boron acceptor consists of a boron on a silicon position with an adjacent carbon vacancy. The carbon vacancy combines with a boron along the hexagonal $c$ axis. It is concluded that 70--90% of the spin density resides in the silicon dangling bonds surrounding the vacancy and another 9% on the neighboring carbon atoms. The spin-density distribution is more localized than in the case of the shallow boron acceptor as deduced from the ENDOR experiments.

Electron paramagnetic resonance and electron nuclear double resonance studies of photoinduced charge separation from N-methylphenothiazine doped into poly(ethylene oxide) and poly(propylene oxide) polymer matrices

Download options Please wait... Article information DOI https://doi.org/10.1039/A804974B Article type Paper Download Citation J. Chem. Soc., Faraday Trans., 1998,94, 3247-3250 BibTex EndNote MEDLINE ProCite ReferenceManager RefWorks RIS Permissions Request permissions Electron paramagnetic resonance and electron nuclear double resonance studies of photoinduced charge separation from N-methylphenothiazine doped into poly(ethylene oxide) and poly(propylene oxide) polymer matrices Y. Soo Kang, J. A. Jung and L. Kevan, J. Chem. Soc., Faraday Trans., 1998, 94, 3247 DOI: 10.1039/A804974B To request permission to reproduce material from this article, please go to the Copyright Clearance Center request page. If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given. If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page. Read more about how to correctly acknowledge RSC content. Search articles by author Young Soo Kang Jung A. Jung Larry Kevan

Electron Spin Resonance and Electron Nuclear Double Resonance Study of X-Irradiated Deoxyadenosine: Proton Transfer Behavior of Primary Ionic Radicals

A study of deoxyadenosine crystals (anhydrous form) after irradiation at 10 K found four base-centered radicals and one sugar-centered radical. Radical R1, thermally stable to about 100 K and photobleachable easily with white light, was the product of deprotonation at the amino group by the primary radical cation. Radical R2, also thermally stable to about 100 K, was the product of protonation at N3 of the primary radical anion. Radical R3, stable to about 170 K, was centered in the deoxyribose moiety and evidently was the result of net hydrogen abstraction from C4'. Radicals R4 and R5 were the C2 and C8 H-addition products with couplings typical of those species. Both R4 and R5 were formed at 10 K and were stable at room temperature. The behavior of R1 in several systems provides additional evidence for significant involvement of the hydrogen-bonding environment in controlling the stabilization (or formation) of radicals resulting directly from ionization, as described previously (Radiat. Res. 131, 272-284, 1992). From comparison of amino-group hydrogen-bonding environments in which radicals with the structure of R1 were stabilized, we conclude that oxygen atoms as proton acceptors are important in permitting the charge and spin separation necessary for radical stabilization. In particular, oxygens of ROH structures seem most efficient by readily permitting a multi-proton shuffle through a mechanism amounting to proton exchange. The collective results show that stabilization of these products is unlikely unless the charge and spin can be separated by at least one intervening molecule.

ENDOR study of γ-irradiated hydrated testosterone orthorhombic single crystals

The molecular structure of free radicals formed in γ-irradiated orthorhombic single crystals of hydrated testosterone was investigated by Electron Nuclear Double Resonance (ENDOR) spectroscopy. Only one kind of radical was observed, which is formed by addition of hydrogen atom to oxygen atom O(3). We observed interaction of the unpaired electron, which is delocalized on the carbons C(3), C(4) and C95), with one α-proton in position 4 and with four unequivalent β-protons connected with the carbon atoms C(2) and C(6). The matrices of the hyperfine couplings and the g-factor of the radical are given.

Metal-Ion Valencies of the FeMo Cofactor in CO-Inhibited and Resting State Nitrogenase by 57Fe Q-Band ENDOR

The resting state of nitrogenase shows an S = 3/2 electron paramagnetic resonance (EPR) signal resulting from the FeMo-cofactor (M-center; inorganic portion, [Mo, Fe7, S9]) of the MoFe-protein. When the enzyme undergoes turnover under a CO atmosphere, this signal disappears and two new ones appear: one under low pressure of CO (denoted lo-CO; 0.08 atm) with g = [2.09, 1.97, 1.93] and the other under high pressure of CO (denoted hi-CO; 0.5 atm) with g = [2.06, 2.06, 2.17]. Our recent Q-band (35 GHz) 57Fe and 13C electron nuclear double resonance (ENDOR) studies clearly identified [FeMo-cofactor][CO]n, as the origin of the EPR signals from both lo-CO (n = 1) and hi-CO (n = 2) [Christie, P. D.; Lee, H. I.; Cameron, L. M.; Hales, B. J.; Orme-Johnson, W. H.; Hoffman, B. M. J. Am. Chem. Soc. 1996, 118, 8707−8709 and Pollack, R. C.; Lee, H. I.; Cameron, L. M.; Derose, V. J.; Hales, B. J.; Orme-Johnson, W. H.; Hoffman, B. M. J. Am. Chem. Soc. 1995, 117, 8686−8687], and a previous paper discusses CO binding in detail [Lee, H. I.; Cameron, L. M.; Hales, B. J.; Hoffman, B. M. J. Am. Chem. Soc. 1997, 119, 10121−10126]. We now present complete orientation-selective 57Fe ENDOR measurements of the CO-bound FeMo-cofactor in both lo- and hi-CO forms of the MoFe-protein from Azotobacter vinelandii. The 57Fe ENDOR signals associated with the seven Fe ions of the FeMo-cofactor of lo-CO can be completely assigned and interpreted in terms of four magnetically distinct iron signals. Analysis of these signals following the procedures of Mouesca et al. [Mouesca, J.-M.; Noodleman, L.; Case, D. A.; Lamotte, B. Inorg. Chem. 1995, 34, 4347−4359] has led us to propose valence assignments and charges for the cofactor cluster, [Mo, Fe7, S9]+ = [Mo4+, Fe3+1, Fe2+6, S2-9]+, organized into one Fe2.5+ pair and five Fe2+ ions, [Mo4+, (2Fe2.5+)1, Fe2+5, S2-9]+. The result is a formal d-electron count of 43. ENDOR and functional studies indicate that the lo-CO, hi-CO, and resting states of the M-center are all at the same oxidation level. Hence, the proposed valency assignments apply to all three states.