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G W Brudvig

Publications and source records attributed to G W Brudvig.

At least 19 recordsLinked to original sources
PubMed2001

Quantifying the ion selectivity of the Ca2+ site in photosystem II: evidence for direct involvement of Ca2+ in O2 formation.

Calcium is an essential cofactor in the oxygen-evolving complex (OEC) of photosystem II (PSII). The removal of Ca2+ or its substitution by any metal ion except Sr2+ inhibits oxygen evolution. We used steady-state enzyme kinetics to measure the rate of O2 evolution in PSII samples treated with an extensive series of mono-, di-, and trivalent metal ions in order to determine the basis for the affinity of metal ions for the Ca2+-binding site. Our results show that the Ca2+-binding site in PSII behaves very similarly to the Ca2+-binding sites in other proteins, and we discuss the implications this has for the structure of the site in PSII. Activity measurements as a function of time show that the binding site achieves equilibrium in 4 h for all of the PSII samples investigated. The binding affinities of the metal ions are modulated by the 17 and 23 kDa extrinsic polypeptides; their removal decreases the free energy of binding of the metal ions by 2.5 kcal/mol, but does not significantly change the time required to reach equilibrium. Monovalent ions are effectively excluded from the Ca2+-binding site, exhibiting no inhibition of O2 evolution. Di- and trivalent metal ions with ionic radii similar to that of Ca2+ (0.99 A) bind competitively with Ca2+ and have the highest binding affinity, while smaller metal ions bind more weakly and much larger ones do not bind competitively. This is consistent with a size-selective Ca2+-binding site that has a rigid array of coordinating ligands. Despite the large number of metal ions that competitively replace Ca2+ in the OEC, only Sr2+ is capable of partially restoring activity. Comparing the physical characteristics of the metal ions studied, we identify the pK(a) of the aqua ion as the factor that determines the functional competence of the metal ion. This suggests that Ca2+ is directly involved in the chemistry of water oxidation and is not only a structural cofactor in the OEC. We propose that the role of Ca2+ is to act as a Lewis acid, binding a substrate water molecule and tuning its reactivity.

Binding, Competitive↗
PubMed2001

Photosynthetic water oxidation in cytochrome b(559) mutants containing a disrupted heme-binding pocket.

The role of cytochrome b(559) in photosynthetic oxygen evolution has been investigated in three chloroplast mutants of Chlamydomonas reinhardtii, in which one of the two histidine axial ligands to the heme, provided by the alpha subunit, has been replaced by the residues methionine, tyrosine, and glutamine. Photosystem two complexes functional for oxygen evolution could be assembled in the methionine and tyrosine mutants up to approximately 15% of wild type levels, whereas no complexes with oxygen evolution activity could be detected in the glutamine mutant. PSII supercomplexes isolated from the tyrosine and methionine mutants were as active as wild type in terms of light-saturated rates of oxygen evolution but in contrast to wild type contained no bound heme despite the presence of the alpha subunit. Oxygen evolution in the tyrosine and methionine mutants was, however, more sensitive to photoinactivation than the WT. Overall, these data establish unambiguously that a redox role for the heme of cytochrome b(559) is not required for photosynthetic oxygen evolution. Instead, our data provide new evidence of a role for cytochrome b(559) in the protection of the photosystem two complex in vivo.

Animals↗
PubMed2001

High-frequency EPR study of a new mononuclear manganese(III) complex:. [(terpy)Mn(N3)3] (terpy = 2,2':6',2''-terpyridine).

The isolation and structural characterization of [(terpy)Mn(III)(N3)3], complex 1, is reported (terpy = 2,2':6',2' '-terpyridine). Complex 1, a product of the reaction between the mixed-valence dimer [(terpy)(H2O)Mn(III)(O)2Mn(IV)(OH2)(terpy)](NO3)3 and NaN3, crystallizes in a triclinic system, space group P1, a = 8.480(1) A, b = 8.9007(2) A, c = 12.109(2) A, alpha = 93.79(1) degrees, beta = 103.17(1) degrees, gamma = 103.11(1) degrees, and Z = 2. Complex 1 exhibits a Jahn-Teller distortion of the octahedron characteristic of a six-coordinated high-spin Mn(III). A vibrational spectroscopic study was performed. The nu(asym)(N3) mode of complex 1 appears in the IR as a strong band at 2035 cm(-1) with a less intense feature at 2072 cm(-1), and in the FT-Raman as a strong band at 2071 cm(-1) with a weaker broad band at 2046 cm(-1). The electronic properties of complex 1 were investigated using a high-field and high-frequency EPR study (190-475 GHz). The different spin Hamiltonian parameters have been determined (D = -3.29 (+/-0.01) cm(-1), E = 0.48 (+/-0.01) cm(-1), E '= 0.53 (+/-0.01) cm(-1), g(x) = 2.00 (+/-0.005), g(y) = 1.98 (+/-0.005), g(z) = 2.01 (+/-0.005)). These parameters are in agreement with the geometry of complex 1 observed in the crystal structure, a D < 0 related to the elongated distortion, and a value of E/D close to 0.2 as expected from the highly distorted octahedron. The two values of the E-parameter are explained by the presence of two slightly different structural forms of complex 1 in the crystal lattice. A second hypothesis was explored to explain the experimental data. The calculation for the simulation was done taking into account that the g and D tensors are not collinear due to the low symmetry of complex 1. In that case, the spin Hamiltonian parameters found are D = -3.29 (+/-0.01) cm(-1), E = 0.51 (+/-0.01) cm(-1), g(x) = 2.00 (+/-0.005), g(y) = 1.98 (+/-0.005), and g(z) = 2.01 (+/-0.005).

Crystallography, X-Ray↗
PubMed2001

Characterization of the O(2)-evolving reaction catalyzed by [(terpy)(H2O)Mn(III)(O)2Mn(IV)(OH2)(terpy)](NO3)3 (terpy = 2,2':6,2"-terpyridine).

The complex [(terpy)(H(2)O)Mn(III)(O)(2)Mn(IV)(OH(2))(terpy)](NO(3))(3) (terpy = 2,2':6,2' '-terpyridine) (1)catalyzes O(2) evolution from either KHSO(5) (potassium oxone) or NaOCl. The reactions follow Michaelis-Menten kinetics where V(max) = 2420 +/- 490 mol O(2) (mol 1)(-1) hr(-1) and K(M) = 53 +/- 5 mM for oxone ([1] = 7.5 microM), and V(max) = 6.5 +/- 0.3 mol O(2) (mol 1)(-1) hr(-1) and K(M) = 39 +/- 4 mM for hypochlorite ([1] = 70 microM), with first-order kinetics observed in 1 for both oxidants. A mechanism is proposed having a preequilibrium between 1 and HSO(5-) or OCl(-), supported by the isolation and structural characterization of [(terpy)(SO(4))Mn(IV)(O)(2)Mn(IV)(O(4)S)(terpy)] (2). Isotope-labeling studies using H(2)(18)O and KHS(16)O(5) show that O(2) evolution proceeds via an intermediate that can exchange with water, where Raman spectroscopy has been used to confirm that the active oxygen of HSO(5-) is nonexchanging (t(1/2) >> 1 h). The amount of label incorporated into O(2) is dependent on the relative concentrations of oxone and 1. (32)O(2):(34)O(2):(36)O(2) is 91.9 +/- 0.3:7.6 +/- 0.3:0.51 +/- 0.48, when [HSO(5-)] = 50 mM (0.5 mM 1), and 49 +/- 21:39 +/- 15:12 +/- 6 when [HSO(5-)] = 15 mM (0.75 mM 1). The rate-limiting step of O(2) evolution is proposed to be formation of a formally Mn(V)=O moiety which could then competitively react with either oxone or water/hydroxide to produce O(2). These results show that 1 serves as a functional model for photosynthetic water oxidation.

Catalysis↗
PubMed2001

Characterization of carotenoid and chlorophyll photooxidation in photosystem II.

Photosystem II (PSII) contains two accessory chlorophylls (Chl(Z), ligated to D1-His118, and Chl(D), ligated to D2-His117), carotenoid (Car), and heme (cytochrome b(559)) cofactors that function as alternate electron donors under conditions in which the primary electron-donation pathway from the O(2)-evolving complex to P680(+) is inhibited. The photooxidation of the redox-active accessory chlorophylls and Car has been characterized by near-infrared (near-IR) absorbance, shifted-excitation Raman difference spectroscopy (SERDS), and electron paramagnetic resonance (EPR) spectroscopy over a range of cryogenic temperatures from 6 to 120 K in both Synechocystis PSII core complexes and spinach PSII membranes. The following key observations were made: (1) only one Chl(+) near-IR band is observed at 814 nm in Synechocystis PSII core complexes, which is assigned to Chl(Z)(+) based on previous spectroscopic studies of the D1-H118Q and D2-H117Q mutants [Stewart, D. H., Cua, A., Chisholm, D. A., Diner, B. A., Bocian, D. F., and Brudvig, G. W. (1998) Biochemistry 37, 10040-10046]; (2) two Chl(+) near-IR bands are observed at 817 and 850 nm in spinach PSII membranes which are formed with variable relative yields depending on the illumination temperature and are assigned to Chl(Z)(+), and Chl(D)(+), respectively; (3) the Chl and Car cation radicals have significantly different stabilities at reduced temperatures with Car(+) decaying much faster; (4) in Synechocystis PSII core complexes, Car(+) decays by recombination with Q(A)(-) and not by Chl(Z)/Chl(D) oxidation, with multiphasic kinetics that are attributed to an ensemble of protein conformers that are trapped as the protein is frozen; and (5) in spinach PSII membranes, Car(+) decays mainly by recombination with Q(A)(-), but also partly by formation of the 850 nm Chl cation radical. The greater stability of Chl(Z)(+) at low temperatures enabled us to confirm that resonance Raman bands previously assigned to Chl(Z)(+) are correctly assigned. In addition, the formation and decay of these cations provide insight into the alternate electron-donation pathways to P680(+).

Carotenoids↗
PubMed2001

Mechanism of photosynthetic water oxidation: combining biophysical studies of photosystem II with inorganic model chemistry.

A mechanism for photosynthetic water oxidation is proposed based on a structural model of the oxygen-evolving complex (OEC) and its placement into the modeled structure of the D1/D2 core of photosystem II. The structural model of the OEC satisfies many of the geometrical constraints imposed by spectroscopic and biophysical results. The model includes the tetranuclear manganese cluster, calcium, chloride, tyrosine Z, H190, D170, H332 and H337 of the D1 polypeptide and is patterned after the reversible O2-binding diferric site in oxyhemerythrin. The mechanism for water oxidation readily follows from the structural model. Concerted proton-coupled electron transfer in the S2-->S3 and S3-->S4 transitions forms a terminal Mn(V)=O moiety. Nucleophilic attack on this electron-deficient Mn(V)=O by a calcium-bound water molecule results in a Mn(III)-OOH species, similar to the ferric hydroperoxide in oxyhemerythrin. Dioxygen is released in a manner analogous to that in oxyhemerythrin, concomitant with reduction of manganese and protonation of a mu-oxo bridge.

Crystallography↗
PubMed2001

Carotenoid photooxidation in photosystem II.

Carotenoids are known to function as light-harvesting pigments and they play important roles in photoprotection in both plant and bacterial photosynthesis. These functions are also important for carotenoids in photosystem II. In addition, beta-carotene recently has been found to function as a redox intermediate in an alternate pathway of electron transfer within photosystem II. This redox role of a carotenoid in photosystem II is unique among photosynthetic reaction centers and stems from the very highly oxidizing intermediates that form in the process of water oxidation. In this minireview, an overview of the electron-transfer reactions in photosystem II is presented, with an emphasis on those involving carotenoids. The carotenoid composition of photosystem II and the physical methods used to study the structure of the redox-active carotenoid are reviewed. Possible roles of carotenoid cations in photoprotection of photosystem II are discussed.

Carotenoids↗
PubMed2001

Factors that determine the unusually low reduction potential of cytochrome c550 in cyanobacterial photosystem II.

A new purification protocol for cytochrome c550 (cyt c550) from His-tagged SYnechocYstis PCC 6803 photosystem II (PSII) was developed which allows the protein to be isolated in high yield and purity. Electron paramagnetic resonance spectroscopy of cyt c550, both free in solution and in intact PSII preparations, yields identical spectra with g values at 1.50, 2.23, and 2.87, which are characteristic for a ferric low-spin bis-histidine coordinated heme. The resonance Raman spectrum of the isolated protein exhibits features characteristic of bis-histidine axial ligation of the iron and a slight ruffling of the heme macrocycle. Together, these results indicate that the heme structure is not very different from most c-type cytochromes, and thus the structure of the heme does not account for its unusually low reduction potential. A direct electrochemical measurement of the reduction potential was performed using square wave voltammetry on a pyrolytic graphite edge electrode, yielding E1.2=-108 mV (vs. NHE) with a peak separation of 5 mV. This value is 150 mV more positive than that previously measured by redox titrations. Because the behavior of the protein in the electrochemistry experiments is indicative of adsorption to the electrode surface, we surmise that binding of the protein to the electrode excludes solvent water from the heme-binding site. We conclude that the degree of solvent exposure makes a significant contribution to the heme reduction potential. Similarly, the binding of cyt c550 to PSII may also reduce the solvent exposure of the heme, and so the direct electrochemical value of the reduction potential may be relevant to the protein in its native state.

Cyanobacteria↗
PubMed2001

Pulsed electron paramagnetic resonance methods for macromolecular structure determination.

Pulsed electron paramagnetic resonance (EPR) distance measurement techniques target macromolecular structure elucidation at both the local and global level. Recent developments in pulse microwave technology and high-field EPR have led to the development of a variety of pulsed EPR distance measurement techniques. These methods have emerged as powerful tools for the determination of structure/function relationships in macromolecular systems. In this review article, we discuss recent applications of long-range and short-range EPR distance measurements.

Animals↗
PubMed2001

Use of EPR spectroscopy to study macromolecular structure and function.

Electron paramagnetic resonance (EPR) spectroscopy is now part of the armory available to probe the structural aspects of proteins, nucleic acids and protein-nucleic acid complexes. Since the mobility of a spin label covalently attached to a macromolecule is influenced by its microenvironment, analysis of the EPR spectra of site-specifically incorporated spin labels (probes) provides a powerful tool for investigating structure-function correlates in biological macromolecules. This technique has become readily amenable to address various problems in biology in large measure due to the advent of techniques like site-directed mutagenesis, which enables site-specific substitution of cysteine residues in proteins, and the commercial availability of thiol-specific spin-labeling reagents (Figure 1). In addition to the underlying principle and the experimental strategy, several recent applications are discussed in this review.

Animals↗
PubMed2000

Assignment of the Qy absorbance bands of photosystem II chromophores by low-temperature optical spectroscopy of wild-type and mutant reaction centers.

Photosystem II (PSII) contains a collection of pheophytins (Pheo) and chlorophylls (Chl) that have unique absorbance spectra depending on their electronic structure and the surrounding protein environment. Despite numerous efforts to identify the spectra of each cofactor, differing assignments of the chromophore absorbance bands and electrochromic effects have led to conflicting models of pigment organization and chromophore interactions in PSII. We have utilized low-temperature measurements on well-defined redox states, together with the use of site-directed mutants, to make spectral assignments of several reaction center (RC) chromophores. Cryogenic (77 K) optical spectroscopy has been used to trap the bound redox-active quinone, Q(A), in the reduced form and measure the effect of the redox state of Q(A) on PSII chromophores without interference from other redox-active cofactors. The Q(A)(-) minus Q(A) difference spectrum contains a number of features that represent the perturbation of Pheo and Chl absorbance bands upon Q(A) reduction. Using site-directed mutants in which the axial ligand of the D1-side monomeric core Chl, P(A), is changed (D1-H198Q) or the hydrogen-bonding environment of the D1-side Pheo is modified (D1-Q130E), we have assigned the Q(y)() absorbance bands of four chromophores shifted by Q(A) reduction including both RC Pheos, the D1-side monomeric accessory Chl (B(A)), and one other Chl in PSII. The absorbance maximum of B(A) was identified at 683.5 nm from least-squares fits of the D1-H198Q minus wild type (WT) Q(A)(-) minus Q(A) double-difference spectrum; this assignment provides new evidence of a secondary effect of site-directed mutation on a RC chromophore. The other chromophores were assigned from simultaneous fits of the WT and D1-Q130E spectra in which the parameters of only the D1-side Pheo were allowed to vary. The D1-side and D2-side Pheos were found to have lambda(max) values at 685.6 and 669.3 nm, respectively, and another Chl influenced by Q(A)(-) was identified at 678.8 nm. These assignments are in good agreement with previous spectral analyses of intact PSII preparations and reveal that the number of chromophores affected by Q(A) reduction has been underestimated previously. In addition, the assignments are generally consistent with chromophore positions that are similar in the PSII RC and the bacterial photosynthetic RC.

Bacteriochlorophylls↗
PubMed1999

Location of the iron-sulfur clusters FA and FB in photosystem I: an electron paramagnetic resonance study of spin relaxation enhancement of P700+.

Photosystem I (PS I) mediates electron-transfer from plastocyanin to ferredoxin via a photochemically active chlorophyll dimer (P700), a monomeric chlorophyll electron acceptor (A0), a phylloquinone (A1), and three [4Fe-4S] clusters (FX/A/B). The sequence of electron-transfer events between the iron-sulfur cluster, FX, and ferredoxin is presently unclear. Owing to the presence of a 2-fold symmetry in the PsaC protein to which the iron-sulfur clusters F(A) and F(B) are bound, the spatial arrangement of these cofactors with respect to the C2-axis of symmetry in PS I is uncertain as well. An unequivocal determination of the spatial arrangement of the iron-sulfur clusters FA and FB within the protein is necessary to unravel the complete electron-transport chain in PS I. In the present study, we generate EPR signals from charge-separated spin pairs (P700+-FredX/A/B) in PS I and characterize them by progressive microwave power saturation measurements to determine the arrangement of the iron-sulfur clusters FX/A/B relative to P700. The microwave power at half saturation (P1/2) of P700+ is greater when both FA and FB are reduced in untreated PS I than when only FA is reduced in mercury-treated PS I. The experimental P1/2 values are compared to values calculated by using P700-FA/B crystallographic distances and assuming that either FA or FB is closer to P700+. On the basis of this comparison of experimental and theoretical values of spin relaxation enhancement effects on P700+ in P700+ [4Fe-4S]- charge-separated pairs, we find that iron-sulfur cluster FA is in closer proximity to P700 than the FB cluster.

Chlorophyll↗
PubMed1999

Orientation of the tetranuclear manganese cluster and tyrosine Z in the O(2)-evolving complex of photosystem II: An EPR study of the S(2)Y(Z)(*) state in oriented acetate-inhibited photosystem II membranes.

Inhibitory treatment by acetate, followed by illumination and rapid freezing, is known to trap the S(2)Y(Z)(*) state of the O(2)-evolving complex (OEC) in photosystem II (PS II). An EPR spectrum of this state exhibits broad split signals due to the interaction of the tyrosyl radical, Y(Z)(*), with the S = 1/2 S(2) state of the Mn(4) cluster. We present a novel approach to analyze S(2)Y(Z)(*) spectra of one-dimensionally (1-D) oriented acetate-inhibited PS II membranes to determine the magnitude and relative orientation of the S(2)Y(Z)(*) dipolar vector within the membrane. Although there exists a vast body of EPR data on isolated spins in oriented membrane sheets, the present study is the first of its kind on dipolar-coupled electron spin pairs in such systems. We demonstrate the feasibility of the technique and establish a rigorous treatment to account for the disorder present in partially oriented 1-D membrane preparations. We find that (i) the point-dipole distance between Y(Z)(*) and the Mn(4) cluster is 7.9 +/- 0.2 A, (ii) the angle between the interspin vector and the thylakoid membrane normal is 75 degrees, (iii) the g(z)()-axis of the Mn(4) cluster is 70 degrees away from the membrane normal and 35 degrees away from the interspin vector, and (iv) the exchange interaction between the two spins is -275 x 10(-)(4) cm(-)(1), which is antiferromagnetic. Due to the sensitivity of EPR line shapes of oriented spin-coupled pairs to the interspin distance, the present study imposes a tighter constraint on the Y(Z)-Mn(4) point-dipole distance than obtained from randomly oriented samples. The geometric constraints obtained from the 1-D oriented sample are combined with published models of the structure of Mn-depleted PS II to propose a location of the Mn(4) cluster. A structure in which Y(Z) is hydrogen bonded to a manganese-bound hydroxide ligand is consistent with available data and favors maximal orbital overlap between the two redox center that would facilitate direct electron- and proton-transfer steps.

Electron Spin Resonance Spectroscopy↗
PubMed1999

Competitive binding of acetate and chloride in photosystem II.

The binding of chloride and acetate to photosystem II (PSII) was examined to elucidate the mechanism of acetate inhibition. The mode of inhibition was studied, and individual binding sites were assigned by steady-state O2 evolution measurements in correlation with electron paramagnetic resonance (EPR) results. Two binding sites were found for acetate, one chloride-sensitive on the electron donor side and one chloride-insensitive on the electron acceptor side. The respective binding constants were as follows: KCl = 0.5 +/- 0.2 mM (chloride binding to the donor side), KI = 16 +/- 5 mM (acetate binding to the donor side), and KI' = 130 +/- 40 mM (acetate binding to the acceptor side). When acetate was bound to the acceptor side of PSII, 200 K illumination induced a narrowed form of the QA-FeII EPR signal, the yield of which was independent of the chloride concentration. When acetate was bound to the donor side, room-temperature illumination produced the S2YZ* state. EPR measurements showed that both the yield and formation rate of this state increased with acetate concentration. Increasing chloride concentrations slowed the rate of formation of the S2YZ* state, but did not affect the steady-state yield of the S2YZ* state. These findings indicate that the light-induced reactions in acetate-inhibited PSII are modulated by both donor side and acceptor side binding of acetate, while the steady-state yield of the S2YZ* state at the high PSII concentrations used for EPR measurements depends primarily on acceptor side turnover. Our data further support a close proximity of chloride to YZ*, indicating a possible role for chloride in the electron-transfer mechanism at the O2-evolving complex.

Acetates↗
PubMed1999

A functional model for O-O bond formation by the O2-evolving complex in photosystem II.

The formation of molecular oxygen from water in photosynthesis is catalyzed by photosystem II at an active site containing four manganese ions that are arranged in di-mu-oxo dimanganese units (where mu is a bridging mode). The complex [H2O(terpy)Mn(O)2Mn(terpy)OH2](NO3)3 (terpy is 2,2':6', 2"-terpyridine), which was synthesized and structurally characterized, contains a di-mu-oxo manganese dimer and catalyzes the conversion of sodium hypochlorite to molecular oxygen. Oxygen-18 isotope labeling showed that water is the source of the oxygen atoms in the molecular oxygen evolved, and so this system is a functional model for photosynthetic water oxidation.

Catalysis↗
PubMed1999

Mapping RNA-protein interactions in ribonuclease P from Escherichia coli using electron paramagnetic resonance spectroscopy.

Ribonuclease P (RNase P) is a catalytic ribonucleoprotein (RNP) essential for tRNA biosynthesis. In Escherichia coli, this RNP complex is composed of a catalytic RNA subunit, M1 RNA, and a protein cofactor, C5 protein. Using the sulfhydryl-specific reagent (1-oxyl-2,2,5, 5-tetramethyl-Delta3-pyrroline-3-methyl)methanethiosulfonate (MTSL), we have introduced a nitroxide spin label individually at six genetically engineered cysteine residues (i.e., positions 16, 21, 44, 54, 66, and 106) and the native cysteine residue (i.e., position 113) in C5 protein. The spin label covalently attached to any protein is sensitive to structural changes in its microenvironment. Therefore, we expected that if the spin label introduced at a particular position in C5 protein was present at the RNA-protein interface, the electron paramagnetic resonance (EPR) spectrum of the spin label would be altered upon binding of the spin-labeled C5 protein to M1 RNA. The EPR spectra observed with the various MTSL-modified mutant derivatives of C5 protein indicate that the spin label attached to the protein at positions 16, 44, 54, 66, and 113 is immobilized to varying degrees upon addition of M1 RNA but not in the presence of a catalytically inactive, deletion derivative of M1 RNA. In contrast, the spin label attached to position 21 displays an increased mobility upon binding to M1 RNA. The results from this EPR spectroscopy-based approach together with those from earlier studies identify residues in C5 protein which are proximal to M1 RNA in the RNase P holoenzyme complex.

Amino Acid Sequence↗
PubMed1998

Characterization of the interaction between manganese and tyrosine Z in acetate-inhibited photosystem II.

When acetate-inhibited photosystem II (PSII) membranes are illuminated at temperatures above 250 K and quickly cooled to 77 K, a 240 G-wide electron paramagnetic resonance (EPR) signal is observed at 10 K. This EPR signal arises from a reciprocal interaction between the spin 1/2 ground state of the S2 state of the Mn4 cluster, for which a multiline EPR signal with shifted 55Mn hyperfine peaks is observed, and the oxidized tyrosine residue, YZ*, for which a broadened YZ* EPR spectrum is observed. The S2YZ* EPR signal in acetate-inhibited PSII is the first in which characteristic spectral features from both paramagnets can be observed. The observation of distinct EPR signals from each of the paramagnets together with the lack of a half-field EPR transition indicates that the exchange and dipolar couplings are weak. Below 20 K, the S2YZ* EPR signal in acetate-inhibited PSII is in the static limit. Above 20 K, the line width narrows dramatically as the broad low-temperature S2YZ* EPR signal is converted to a narrow YZ* EPR signal at room temperature. The line width narrowing is interpreted to be due to averaging of the exchange and dipolar interactions between YZ* and the S2 state of the Mn4 cluster by rapid spin-lattice relaxation of the Mn4 cluster as the temperature is increased. Decay of the S2YZ* intermediate at 200 K shows that the g = 4.1 form of the S2 state is formed and that a noninteracting S2-state multiline EPR signal is not observed as an intermediate in the decay. This result shows that a change in the redox state of YZ induces a spin-state change in the Mn4 cluster in acetate-inhibited PSII. The interconversion between spin states of the Mn4 cluster in acetate-inhibited PSII supports the idea that YZ oxidation or YZ* reduction is communicated to the Mn4 cluster through a direct hydrogen-bonding pathway, possibly involving a ligand bound to the Mn4 cluster.

Electron Spin Resonance Spectroscopy↗