Evidence for a direct manganese-oxygen ligand in water binding to the S2 state of the photosynthetic water oxidation complex

The interaction of water with the water oxidizing Mn complex of photosystem II has been investigated using electron spin-echo envelope modulation spectroscopy in the presence of H(2)(17)O. The spectra show interaction of the (17)O with the preparation in the S(2) state induced by 200 K illumination. The modulation is observed only in the center of the multiline spectrum. The inferred hyperfine coupling terms are compatible with water (not hydroxyl) oxygen bound to a particular quasi-axial Mn(III) center in a coupled Mn cluster.     

Cooperative binding of oxygen to the water-splitting enzyme in the filamentous cyanobacterium Oscillatoria chalybea

In the filamentous cyanobacterium Oscillatoria chalybea photolysis of water does not take place in the complete absence of oxygen. A catalytic oxygen partial pressure of 15x10(-6) Torr has to be present for effective water splitting to occur. By means of mass spectrometry we measured the photosynthetic oxygen evolution in the presence of H(2)(18)O in dependence on the oxygen partial pressure of the atmosphere and analysed the liberations of (16)O(2), (16)O(18)O and (18)O(2) simultaneously. The observed dependences of the light-induced oxygen evolution on bound oxygen yield sigmoidal curves. Hill coefficient values of 3.0, 3.1 and 3.2, respectively, suggest that the binding is cooperative and that four molecules of oxygen have to be bound per chain to the oxygen evolving complex. Oxygen seems to prime the water-splitting reaction by redox steering of the S-state system, putting it in the dark into the condition from which water splitting can start. It appears that in O. chalybea an interaction of oxygen with S(0) and S(1) leads to S(2) and S(3), thus yielding the typical oxygen evolution pattern in which even after extensive dark adaptation substantial amounts of Y(1) and Y(2) are found. The interacting oxygen is apparently reduced to hydrogen peroxide. Mass spectrometry permits to distinguish this highly specific oxygen requirement from the interaction of bulk atmospheric oxygen with the oxygen evolving complex of the cyanobacterium. This interaction leads to the formation H(2)O(2) which is decomposed under O(2) evolution in the light. The dependence on oxygen-partial pressure and temperature is analysed. Structural peculiarities of the cyanobacterial reaction centre of photosystem II referring to the extrinsic peptides might play a role.     

Oxygen ligand exchange at metal sites - implications for the O2 evolving mechanism of photosystem II

The mechanism for photosynthetic O2 evolution by photosystem II is currently a topic of intense debate. Important questions remain as to what is the nature of the binding sites for the substrate water and how does the O-O bond form. Recent measurements of the 18O exchange between the solvent water and the photogenerated O2 as a function of the S-state cycle have provided some surprising insights to these questions (W. Hillier, T. Wydrzynski, Biochemistry 39 (2000) 4399-4405). The results show that one substrate water molecule is bound at the beginning of the catalytic sequence, in the S0 state, while the second substrate water molecule binds in the S3 state or possibly earlier. It may be that the second substrate water molecule only enters the catalytic sequence following the formation of the S3 state. Most importantly, comparison of the observed exchange rates with oxygen ligand exchange in various metal complexes reveal that the two substrate water molecules are most likely bound to separate Mn(III) ions, which do not undergo metal-centered oxidations through to the S3 state. The implication of this analysis is that in the S1 state, all four Mn ions are in the +3 oxidation state. This minireview summarizes the arguments for this proposal.     

Evidence that bicarbonate is not the substrate in photosynthetic oxygen evolution

It is widely accepted that the oxygen produced by photosystem II of cyanobacteria, algae, and plants is derived from water. Earlier proposals that bicarbonate may serve as substrate or catalytic intermediate are almost forgotten, though not rigorously disproved. These latter proposals imply that CO2 is an intermediate product of oxygen production in addition to O2. In this work, we investigated this possible role of exchangeable HCO3- in oxygen evolution in two independent ways. (1) We studied a possible product inhibition of the electron transfer into the catalytic Mn4Ca complex during the oxygen-evolving reaction by greatly increasing the pressure of CO2. This was monitored by absorption transients in the near UV. We found that a 3,000-fold increase of the CO2 pressure over ambient conditions did not affect the UV transient, whereas the S(3) --> S(4) --> S(0) transition was half-inhibited by raising the O2 pressure only 10-fold over ambient, as previously established. (2) The flash-induced O2 and CO2 production by photosystem II was followed simultaneously with membrane inlet mass spectrometry under approximately 15% H2(18)O enrichment. Light flashes that revealed the known oscillatory O2 release failed to produce any oscillatory CO2 signal. Both types of results exclude that exchangeable bicarbonate is the substrate for (and CO2 an intermediate product of) oxygen evolution by photosynthesis. The possibility that a tightly bound carbonate or bicarbonate is a cofactor of photosynthetic water oxidation has remained.     

Interaction of ammonia with the water splitting enzyme of photosystem II

The effects of NH3 on the oxygen evolving enzyme have been investigated with EPR and steady-state O2 evolution. The following results were obtained. At low light intensity O2 evolution occurs in all centers even though ammonia is bound. This binding occurs in the S2 state and results in a modification of the multiline signal as reported earlier. However, the oscillations with flash number of the amplitude of the EPR signal are virtually unaffected, indicating that NH3 binding does not prevent S-state advancement. Inhibition of O2 evolution by NH3 measured at light intensities that are nearly saturating for untreated photosystem II is interpreted as being due to a slow down in the rate of S-state cycling. At very high light intensities NH3 is not able to inhibit oxygen evolution presumably because NH3 binding is S state dependent and the susceptible S state (S2) is turned over too quickly. NH3 binding resulting in the modified multiline signal does not occur in S1. When S1 is formed from fully NH3 modified S2 by deactivation or by three further flashes, the S1 state does not have NH3 bound. NH3 thus dissociates easily from S1. Earlier reports of NH3 binding in S1 may be explained by the observation that NH3 binding can occur upon incubation of samples in S2 at temperatures as low as 198 K. Evidence is obtained for an NH3 binding occurring slowly (30 s) in S3. This binding results in a block in S-state advancement as suggested earlier [Velthuys, B. R. (1975) Thesis, University of Leiden]. The results are interpreted in two possible models: (1) NH3 binding in S2 occurs in a substrate site, but it is rapidly exchanged by water upon S4 formation. (2) NH3 binding in S2 is not in a substrate site but instead in a structural site and remains bound while water is oxidized. Inherent in this model is that other NH3 binding sites, i.e., the Cl- site, and the slow NH3 binding site in S3 could be the true substrate sites. Some mechanistic implications are discussed.     

Inhibition of photosynthetic oxygen evolution by calmodulin-type inhibitors and other calcium-antagonists

Several studies have recently implicated a role for Ca2+ in photosynthetic oxygen evolution (9-11). Our previous study indicated that Ca2+ was likely acting at the level of the Cl- cofactor requirement in photosystem II (9). We now demonstrate, through the use of calmodulin-type inhibitors ( calmidazolium and trifluoperazine) and metal Ca2+-antagonists (e.g., Tb+3 and La+3), the function of Ca2+ on the oxidizing side of photosystem II. In addition, the peroxide (H2O2) electron donation site was differentiated from the electron donation site of NH2OH, Mn2+ and diphenyl carbazide in the photosystem II complex.     

Structural and functional dynamics of plant photosystem II

Given the unique problem of the extremely high potential of the oxidant P(+)(680) that is required to oxidize water to oxygen, the photoinactivation of photosystem II in vivo is inevitable, despite many photoprotective strategies. There is, however, a robustness of photosystem II, which depends partly on the highly dynamic compositional and structural heterogeneity of the cycle between functional and non-functional photosystem II complexes in response to light level. This coordinated regulation involves photon usage (energy utilization in photochemistry) and excess energy dissipation as heat, photoprotection by many molecular strategies, photoinactivation followed by photon damage and ultimately the D1 protein dynamics involved in the photosystem II repair cycle. Compelling, though indirect evidence suggests that the radical pair P(+)(680)Pheo(-) in functional PSII should be protected from oxygen. By analogy to the tentative oxygen channel of cytochrome c oxidase, oxygen may be liberated from the two water molecules bound to the catalytic site of the Mn cluster, via a specific pathway to the membrane surface. The function of the proposed oxygen pathway is to prevent O(2) from having direct access to P(+)(680)Pheo(-) and prevent the generation of singlet oxygen via the triplet-P(680) state in functional photosytem IIs. Only when the, as yet unidentified, potential trigger with a fateful first oxidative step destroys oxygen evolution, will the ensuing cascade of structural perturbations of photosystem II destroy the proposed oxygen, water and proton pathways. Then oxygen has direct access to P(+)(680)Pheo(-), singlet oxygen will be produced and may successively oxidize specific amino acids of the phosphorylated D1 protein of photosystem II dimers that are confined to appressed granal domains, thereby targeting D1 protein for eventual degradation and replacement in non-appressed thylakoid domains.     

Reduction-induced inhibition and Mn(II) release from the photosystem II oxygen-evolving complex by hydroquinone or NH2OH are consistent with a Mn(III)/Mn(III)/Mn(IV)/Mn(IV) oxidation state for the dark-adapted enzyme

Hydroxylamine and hydroquinone were used to probe the oxidation states of Mn in the oxygen-evolving complex of dark-adapted intact (hydroxylamine) and salt-washed (hydroquinone) photosystem II. These preparations were incubated in the dark for 24 h in the presence of increasing reductant/photosystem II ratios, and the loss of oxygen evolution activity and of Mn(II) was determined for each incubation mixture. Monte Carlo simulations of these data yielded models that provide insight into the structure, reactivity, and oxidation states of the manganese in the oxygen-evolving complex. Specifically, the data support oxidation states of Mn(III)(2)/Mn(IV)(2) for the dark stable S(1) state of the O(2)-evolving complex. Activity and Mn(II) loss data were best modeled by assuming an S(1) --> S(-)(1) conversion of intermediate probability, a S(-)(1) --> S(-)(3) reaction of high probability, and subsequent step(s) of low probability. This model predicts that photosystem II Mn clusters that have undergone an initial reduction step become more reactive toward a second reduction, followed by a slower third reduction step. Analysis of the Mn(II) release parameters used to model the data suggests that the photosystem II manganese cluster consists of three Mn atoms that exhibit a facile reactivity with both reductants, and a single Mn that is reducible but sterically trapped at or near its binding site. Activity assays indicate that intact photosystem II centers reduced to S(-)(1) can evolve oxygen upon illumination, but that these centers are inactive in preparations depleted of the extrinsic 23 and 17 kDa polypeptides. Finally, it was found that a substantial population of the tyrosine D radical is reduced by hydroxylamine, but a smaller population reacts with hydroquinone over the course of a 24 h exposure to the reductant.     

Effect of the 17- and 23-kilodalton polypeptides, calcium, and chloride on electron transfer in photosystem II

Electron paramagnetic resonance (EPR) measurements were performed on photosystem II (PSII) membranes that were treated with 2 M NaCl to release the 17- and 23-kilodalton (kDa) polypeptides. By using 75 microM 3-(3,4-dichlorophenyl)-1,1-dimethylurea to limit the photosystem II samples to one stable charge separation in the temperature range of 77-273 K, we have quantitated the EPR signals of the several electron donors and acceptors of photosystem II. It was found that removal of the 17- and 23-kDa polypeptides caused low potential cytochrome b559 to become fully oxidized during the course of dark adaptation. Following illumination at 77-130 K, one chlorophyll molecule per reaction center was oxidized. Between 130 and 200 K, both a chlorophyll molecule and the S1 state were photooxidized and, together, accounted for one oxidation per reaction center. Above 200 K, the chlorophyll radical was unstable. Oxidation of the S1 state gave rise to the S2-state multiline EPR signal, which arises from the Mn site of the O2-evolving center. The yield of the S2-state multiline EPR signal in NaCl-washed PSII membranes was as high as 93% of the control, untreated PSII membranes, provided that both Ca2+ and Cl- were bound. Furthermore, the 55Mn nuclear hyperfine structure of the S2-state multiline EPR signal was unaltered upon depletion of the 17- and 23-kDa polypeptides. In NaCl-washed PSII samples where Ca2+ and/or Cl- were removed, however, the intensity of the S2-state multiline EPR signal decreased in parallel with the fraction of PSII lacking bound Ca2+ and Cl-.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neurophilin-1 is a downstream target of transcription factor Ets-1 in human umbilical vein endothelial cells

Photosystem II complex (PSII) of thylakoid membranes uses light energy to oxidise extremely stable water and produce oxygen (2H(2)O-->O(2)+4H(+)+4e(-)). PSII is compared with cytochrome c oxidase that catalyses the opposite reaction coupled to proton translocation. Cytochrome c oxidase has proton and water channels, and a tentative oxygen channel. I propose that functional PSII complexes also need a specific oxygen channel to direct O(2) from the water molecules bound to specific Mn atoms of the Mn cluster within PSII out to the membrane surface. The function of this channel will be to prevent oxygen being accessible to the radical pair P680(+)Pheo(-), thereby preventing singlet oxygen generation from the triplet P680 state in functional PSII. The important role of singlet oxygen in structurally perturbed non-functional photosystem II is also discussed.     

The function of the chloride ion in photosynthetic oxygen evolution

The involvement of Cl(-) and several other monovalent anions in photosynthetic oxygen evolution was studied using photosystem II membranes depleted of Cl(-) by dialysis. The results of these studies differ significantly from results obtained using other depletion methods. Binding studies with glycerol as a cryoprotectant confirm our previous observations with sucrose of two interconvertible binding states of photosystem II with similar activities and with slow or fast exchange, respectively, of the bound ion. With glycerol, Cl(-) depletion decreased the oxygen evolution rate to 55% of that with Cl(-) present without decreasing the quantum efficiency of the reaction, supporting our previous conclusion that oxygen evolution can proceed at high rates in the absence of Cl(-). Further, after Cl(-) depletion the S(2) state multiline signal displayed the same periodic appearance with the same signal yield after consecutive laser flashes as with Cl(-) present. Br(-), I(-), and NO(3)(-), although with different capacities to reactivate oxygen evolution, also showed two binding modes. I(-) inhibited when bound in the low-affinity, fast-exchange mode but activated in the high-affinity mode. A comparison of the EPR properties of the S(2) state with these anions suggests that the nature of the ion or the binding mode only has a minor influence on the environment of the manganese. In contrast, F(-) completely inhibited oxygen evolution by preventing the S(2) to S(3) transition and shifted the equilibrium between the g = 4.1 and multiline S(2) forms toward the former, which suggests a considerable perturbation of the manganese cluster. To explain these and earlier observations, we propose that the role of chloride in the water-splitting mechanism is to participate together with charged amino acid side chains in a proton-relay network, which facilitates proton transfer from the manganese cluster to the medium. The structural requirements likely to be involved may explain the sensitivity of oxygen evolution to Cl(-) depletion or other perturbations.     

Comparison of the structure of the manganese complex in the S1 and S2 states of the photosynthetic O2-evolving complex: an x-ray absorption spectroscopy study

A Mn-containing enzyme complex is involved in the oxidation of H2O to O2 in algae and higher plants. X-ray absorption spectroscopy is well suited for studying the structure and function of Mn in this enzyme complex. Results of X-ray K-edge and extended X-ray absorption fine structure (EXAFS) studies of Mn in the S1 and S2 states of the photosynthetic O2-evolving complex in photosystem II preparations from spinach are presented in this paper. The S2 state was prepared by illumination at 190 K or by illumination at 277 K in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU); these are protocols that limit the photosystem II reaction center to one turnover. Both methods produce an S2 state characterized by a multiline electron paramagnetic resonance (EPR) signal. An additional protocol, illumination at 140 K, produces as a state characterized by the g = 4.1 EPR signal. We have previously observed a shift to higher energy in the X-ray absorption K-edge energy of Mn upon advancement from the dark-adapted S1 state to the S2 state produced by illumination at 190 K [Goodin, D. B., Yachandra, V. K., Britt, R. D., Sauer, K., & Klein, M. P. (1984) Biochim. Biophys. Acta 767, 209-216]. The Mn K-edge spectrum of the 277 K illuminated sample is similar to that produced at 190 K, indicating that the S2 state is similar when produced at 190 or 277 K.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetics of O2 evolution from H2O2 catalyzed by the oxygen-evolving complex: investigation of the S1-dependent reaction

The evolution of O2 from H2O2 catalyzed by the oxygen-evolving complex (OEC) in darkness was examined with photosystem II reaction center complex preparations from spinach. Flash illumination of dark-adapted reaction centers was used to make S0-enriched or S1-enriched complexes. The membranes catalyzed O2 evolution from H2O2 when preset to either the S0 or S1 state. However, only the S0-state reaction was inhibited by carbonyl cyanide m-chlorophenylhydrazone and dependent on chloride. These results indicate that (1) the S0-dependent and S1-dependent catalytic cycles can be separated by flash illumination, (2) the S0-dependent reaction involves the formation of the S2 state, and (3) the S1-dependent reaction does not involve the formation of the S2 or S3 states. A kinetic study of the S1-dependent reaction revealed a rapid equilibrium ordered mechanism in which (1) the binding of Ca(II) must precede the binding of H2O2 to the OEC and (2) the reaction of Ca(II) with the free enzyme is at thermodynamic equilibrium such that Ca(II) does not necessarily dissociate after each catalytic cycle.     

Too much light? How beta-carotene protects the photosystem II reaction centre.

The photosystem II reaction centre of all oxygenic organisms is subject to photodamage by high light i.e. photoinhibition. In this review I discuss the reasons for the inevitable and unpreventable oxidative damage that occurs in photosystem II and the way in which beta-carotene bound to the reaction centre significantly mitigates this damage. Recent X-ray structures of the photosystem II core complex (reaction centre plus the inner antenna complexes) have revealed the binding sites of some of the carotenoids known to be bound to the complex. In the light of these X-ray structures and their known biophysical properties it is thus possible to identify the two beta-carotenes present in the photosystem II reaction centre. The two carotenes are both bound to the D2 protein and this positioning is discussed in relation to their ability to act as quenchers of singlet oxygen, generated via the triplet state of the primary electron donor. It is proposed that their location on the D2 polypeptide means there is more oxidative damage to the D1 protein and that this underlies the fact that this latter protein is continuously re-synthesised, at a far greater rate than any other protein involved in photosynthesis. The relevance of a cycle of electrons around photosystem II, via cytochrome b(559), in order to re-reduce the beta-carotenes when they are oxidised and hence restore their ability to quench singlet oxygen, is also discussed.     

Electron spin-lattice relaxation of the S0 state of the oxygen-evolving complex in photosystem II and of dinuclear manganese model complexes

The temperature dependence of the electron spin-lattice relaxation time T1 was measured for the S0 state of the oxygen-evolving complex (OEC) in photosystem II and for two dinuclear manganese model complexes by pulse EPR using the inversion-recovery method. For [Mn(III)Mn(IV)(mu-O)2 bipy4]ClO4, the Raman relaxation process dominates at temperatures below 50 K. In contrast, Orbach type relaxation was found for [Mn(II)Mn(III)(mu-OH)(mu-piv)2(Me3 tacn)2](ClO4)2 between 4.3 and 9 K. For the latter complex, an energy separation of 24.7-28.0 cm(-1) between the ground and the first excited electronic state was determined. In the S0 state of photosystem II, the T1 relaxation times were measured in the range of 4.3-6.5 K. A comparison with the relaxation data (rate and pre-exponential factor) of the two model complexes and of the S2 state of photosystem II indicates that the Orbach relaxation process is dominant for the S0 state and that its first excited state lies 21.7 +/- 0.4 cm(-1) above its ground state. The results are discussed with respect to the structure of the OEC in photosystem II.     

Temporal Changes in State Transitions and Photosystem Organization in the Unicellular,Diazotrophic Cyanobacterium Cyanothece sp. ATCC 51142

The unicellular Cyanobacterium Cyanothece sp. ATCC 51142, grown under alternating 12-h light/12-h dark conditions, temporally separated N2 fixation from photosynthesis. The regulation of photosynthesis was studied using fluorescence spectra and kinetics to determine changes in state transitions and photosystem organization. The redox poise of the plastoquinone (PQ) pool appeared to be central to this regulation. Respiration supported N2 fixation by oxidizing carbohydrate granules, but reduced the PQ pool. This induced state 2 photosystem II monomers and lowered the capacity for O2 evolution. State 2 favored photosystem I trimers and cyclic electron transport, which could stimulate N2 fixation; the stimulation suggested an ATP limitation to N2 and CO2 fixation. The exhaustion of carbohydrate granules at around 6 h in the dark resulted in reduced respiratory electron flow, which led to a more oxidized PQ pool and produced a sharp transition from state 2 to state 1. This transient state 1 returned to state 2 in the remaining hours of darkness. In the light phase, photosystem II dimerization correlated with increased phycobilisome coupling to photosystem II (state 1) and increased rates of O2 evolution. However, dark adaptation did not guarantee state 2 and left photosystem I centers in a mostly monomeric state at certain times.     

Removal of the 33, 23 and 18 kDa extrinsic proteins of photosystem II by sulfite treatment at alkaline pH

We found that sulfite incubation of photosystem II submembrane fractions can induce selective depletion of the 18, 23 and 33 kDa polypeptides of the PSII oxygen evolving complex. When the sulfite treatment was done at pH 8.0, the 18 and 23 kDa proteins were removed efficiently from the PSII oxygen evolving complex. Under the same conditions, the 33 kDa subunit remained bound (even when 2 M sodium sulfite was used). However, in more alkaline conditions (pH 9.8), we show extensive removal of the 33 kDa in the presence of a low sulfite concentration (50 mM). The different extraction affinity for the 18, 23 and 33 kDa of the photosystem II complex was interpreted to mean that the 33 kDa polypeptides are bound to photosystem II by both electrostatic and hydrogen bonding forces.     

Reduction of the Mn cluster of the water-oxidizing enzyme by nitric oxide: formation of an S(-2) state

The manganese cluster of the oxygen-evolving enzyme of photosystem II is chemically reduced upon interaction with nitric oxide at -30 degrees C. The state formed gives rise to an S = 1/2 multiline EPR signal [Goussias, Ch., Ioannidis, N., and Petrouleas, V. (1997) Biochemistry 36, 9261] that is attributed to a Mn(II)- Mn(III) dimer [Sarrou, J., Ioannidis, N., Deligiannakis, Y., and Petrouleas, V. (1998) Biochemistry 37, 3581]. In this work, we sought to establish whether the state could be assigned to a specific, reduced S state by using flash oxymetry, chlorophyll a fluorescence, and electron paramagnetic resonance spectroscopy. With the Joliot-type O(2) electrode, the first maximum of oxygen evolution was observed on the sixth or seventh flash. Three saturating pre-flashes were required to convert the flash pattern characteristic of NO-reduced samples to that of the untreated control (i.e., O(2) evolution maximum on the third flash). Measurements of the S state-dependent level of chlorophyll fluorescence in NO-treated PSII showed a three-flash downshift compared to untreated controls. In the EPR study, the maximum S(2) multi-line EPR signal was observed after the fourth flash. The results from all three methods are consistent with the Mn cluster being in a redox state corresponding to an S(-2) state in a majority of centers after treatment with NO. We were unable to generate the Mn(II)-Mn(III) multi-line signal using hydrazine as a reductant; it appears that the valence distribution and possibly the structure of the Mn cluster in the S(-2) state are dependent on the nature of the reductant that is used.     

Reaction pattern and mechanism of light induced oxidative water splitting in photosynthesis

This mini review is an attempt to briefly summarize our current knowledge on light driven oxidative water splitting in photosynthesis. The reaction leading to molecular oxygen and four protons via photosynthesis comprises thermodynamic and kinetic constraints that require a balanced fine tuning of the reaction coordinates. The mode of coupling between electron (ET) and proton transfer (PT) reactions is shown to be of key mechanistic relevance for the redox turnover of Y(Z) and the reactions within the WOC. The WOC is characterized by peculiar energetics of its oxidation steps in the WOC. In all oxygen evolving photosynthetic organisms the redox state S(1) is thermodynamically most stable and therefore this general feature is assumed to be of physiological relevance. Available information on the Gibbs energy differences between the individual redox states S(i+1) and S(i) and on the activation energies of their oxidative transitions are used to construct a general reaction coordinate of oxidative water splitting in photosystem II (PS II). Finally, an attempt is presented to cast our current state of knowledge into a mechanism of oxidative water splitting with special emphasis on the formation of the essential O-O bond and the active role of the protein environment in tuning the local proton activity that depends on time and redox state S(i). The O-O linkage is assumed to take place within a multistate equilibrium at the redox level of S(3), comprising both redox isomerism and proton tautomerism. It is proposed that one state, S(3)(P), attains an electronic configuration and nuclear geometry that corresponds with a hydrogen bonded peroxide which acts as the entatic state for the generation of complexed molecular oxygen through S(3)(P) oxidation by Y(Z)(ox).