Roehl M. Cinco

X-ray spectroscopy-based structure of the Mn cluster and mechanism of photosynthetic oxygen evolution

The mechanism by which the Mn-containing oxygen evolving complex (OEC) produces oxygen from water has been of great interest for over 40 years. This review focuses on how X-ray spectroscopy has provided important information about the structure of this Mn complex and its intermediates, or S-states, in the water oxidation cycle. X-ray absorption near-edge structure spectroscopy and high-resolution Mn Kbeta X-ray emission spectroscopy experiments have identified the oxidation states of the Mn in the OEC in each of the intermediate S-states, while extended X-ray absorption fine structure experiments have shown that 2.7 A Mn-Mn di-mu-oxo and 3.3 A Mn-Mn mono-mu-oxo motifs are present in the OEC. X-ray spectroscopy has also been used to probe the two essential cofactors in the OEC, Ca2+ and Cl-, and has shown that Ca2+ is an integral component of the OEC and is proximal to Mn. In addition, dichroism studies on oriented PS II membranes have provided angular information about the Mn-Mn and Mn-Ca vectors. Based on these X-ray spectroscopy data, refined models for the structure of the OEC and a mechanism for oxygen evolution by the OEC are presented.

Counting the number of disulfides and thiol groups in proteins and a novel approach for determining the local pKa for cysteine groups in proteins in vivo

X-ray Absorption Spectroscopy (XAS) is a powerful tool to investigate sulfur in biological molecules. The spectral features are sensitive to the local electronic and geometric environment of the atom; thus, they constitute a fingerprint of the different chemical forms in which the sulfur is present. This allows straightforward detection of the ratio between free thiols and disulfides. Intra- or inter-molecular disulfide bond formation between residues plays an important role in structural and conformational changes in proteins, and such changes can be investigated using sulfur XAS. Also, a thiolate-disulfide equilibrium is involved in the regulation of the redox potential in the cells by means of modulating the concentrations of the reduced (thiolate) and oxidized (disulfide) form of the tripeptide glutathione. Thus, we can monitor the redox state of a cell by means of sulfur XAS. Thiols also exhibit an acid-base equilibrium, and sulfur XAS can be used to determine the local pKa of the -SH group. Here we report examples of how sulfur XAS has been used for these applications.

Oxidation States and Structure of the Manganese Cluster in the S 0 State of the Oxygen Evolving Complex

Photosystem II (PS II) catalyzes the light driven oxidation of water to molecular oxygen and the reduction of plastoquinone to plastohydroquinone. Water oxidation occurs in the oxygen evolving complex (OEC) of PS II that is known to cycle through five different redox states, referred to as the S states (S0,..,S4). A cluster of four Mn, one Ca and Cl− is thought to form the central unit of the OEC which stores most of the oxidizing equivalents and binds the substrate water.

Influence of the 33 KDa Manganese-Stabilizing Protein on the Structure And Substrate Accessibility of the Oxygen-Evolving Complex of Photosystem II

The 33 kDa manganese-stabilizing extrinsic protein binds to the lumenal side of photosystem II (PS II) close to the Mn(4)Ca cluster of the oxygen-evolving complex, where it limits access of small molecules to the metal site. Our previous finding that the removal of this protein did not alter the magnetic coupling regime within the manganese cluster, measured by electron spin-echo envelope modulation [Gregor, W., and Britt, R. D. (2000) Photosynth. Res. 65, 175-185], prompted us to examine whether this accessibility control is also true for substrate water, using the same pulsed EPR technique. Comparing the deuteron modulation of the S(2)-state multiline signal of PS II membranes, equilibrated with deuterated water (D(2)O) after removal or retention of the 33 kDa protein, we observed no change in the number and the distance of deuterons magnetically coupled to manganese, indicating that the number and distance of water molecules bound to the manganese cluster are independent of bound 33 kDa protein in the S(1) state, in which the sample was poised prior to cryogenic illumination. A simple modulation depth analysis revealed a distance of 2.5-2.6 A between the closest deuteron and manganese. These results are in agreement with our refined X-ray absorption analysis. The manganese K-edge positions, reflecting their oxidation states, and the extended X-ray absorption fine structure amplitudes and distances between the manganese ions and their oxygen and nitrogen ligands (1.8, 2.7, and 3.3-3.4 A) were independent of bound 33 kDa protein.

Proximity of calcium to the manganese cluster of the photosynthetic oxygen-evolving complex determined from strontium XAFS

Working on Photosystem II membranes from spinach, we substituted strontium for calcium and probed using Sr EXAFS for any nearby Mn. Sr EXAFS results indicate major differences between the functional (intact) and inactive (hydroxylamine-treated) samples. In intact samples, the Fourier transform of the Sr EXAFS shows a major peak that is missing in inactive samples. This interaction is best simulated by two Mn neighbors at a distance of ~3.5 A.

Calcium and Chloride Cofactors of the Oxygen Evolving Complex - X-Ray Absorption Spectroscopy Evidence for A Mn/Ca/Cl Heteronuclear Cluster

The oxygen-evolving complex (OEC) of photosystem II (PSII) in green plants and algae contains a cluster of four Mn atoms in the active site, which catalyzes the oxidation of water to dioxygen. Along with Mn, Cl− and Ca2+ are essential cofactors for oxygen evolution (1).

S K- and Mo L-edge X-ray absorption spectroscopy to determine metal-ligand charge distribution in molybdenum-sulfur compounds.

Mo L-edge and S K-edge X-ray absorption spectroscopy were applied to investigate the charge distribution between Mo and S in a series of Mo thiolate compounds, which serve as amide-sulfur H-bonding models and exhibit different redox potentials arising from polar group effects and ligand hydrogen bonds near the redox center. For all oxidized complexes, the S K-edge spectra exhibit a thiolate-based pre-edge feature centered at 2470.2 eV and the inflection point oCCurs at 2472.0 eV. No intense pre-edge feature is observed in the spectra for the reduced Mo model compounds and the energy shift of the S K-edge position depends on the S-ligand. Correlations between ligand charge density and the redox potential of the Mo-S cores are observed.

Refined Model of the Oxidation States and Structures of the Mn/Ca/Cl Cluster of the Oxygen Evolving Complex of Photosystem II

Central to the problem of photosynthetic oxygen evolution is the structure and function of the Mn/Ca/Cl complex that appears to be the locus of charge accumulation and water splitting. In the recent past our group has presented a topological model for the structure of the tetranuclear Mn cluster, the oxidation state assignments of the S-states of the Kok cycle, the orientation of the Mn-Mn vectors relative to the membrane normal, and evidence for the proximity of Ca to the Mn (1–3).

Recent Advances Toward A Structural Model for the Photosynthetic Oxygen-Evolving Manganese Cluster

Photosynthetic water oxidation occurs in the oxygen evolving complex (OEC) of photosystem II (PS II). One-electron photo-oxidations in the reaction center of PS II are coupled to the four-electron oxidation of water in the OEC. As the PS II reaction center sequentially extracts electrons, the OEC cycles through five intermediate oxidation states (So-S4) where So is the least oxidized and S4 is a transient state which decays to So with the release of a dioxygen molecule. A complex of four manganese atoms has been shown to function in charge accumulation and is thought to form the catalytic site for the water oxidation reaction. The structure of this manganese complex and the mechanism of water oxidation have been the subject of vigorous inquiry by a great many research groups (Sauer et al., 1992).