Hiroki Nagashima

Electronic structure of S2 state of the oxygen-evolving complex of photosystem II studied by PELDOR

Photosynthetic water splitting is catalyzed by a Mn(4)CaO(5) cluster in photosystem II, whose structure was recently determined at a resolution of 1.9Å [Umena, Y. et al. 2011, Nature, 473:55-60]. To determine the electronic structure of the Mn(4)CaO(5) cluster, pulsed electron-electron double resonance (PELDOR) measurements were performed for the tyrosine residue Y(D)() and S(2) state signals with non-oriented and oriented photosystem II (PS II) samples. Based on these measurements, the spin density distributions were calculated by comparing with the experimental results. The best fitting parameters were obtained with a model in which Mn1 has a large positive projection, Mn3 has a small positive projection, and Mn2 and Mn4 have negative projections (the numbering of Mni (i=1-4) is based on the crystal structure at a 1.9Å resolution), which yielded spin projections of 1.97, -1.20, 1.19 and -0.96 for Mn1-4 ions. The results show that the Mn1 ion, which is coordinated by H332, D342 and E189, has a valence of Mn(III) in the S(2) state. The sign of the exchange interactions J(13) is positive, and the other signs are negative.

Highly resolved proton matrix ENDOR of oriented photosystem II membranes in the S2 state

Proton matrix ENDOR was performed to investigate the protons close to the manganese cluster in oriented samples of photosystem II (PS II). Eight pairs of ENDOR signals were detected in oriented PS II membranes. At an angle of θ=0° between the membrane normal vector n and the external field H0, five pairs of ENDOR signals were exchangeable in D2O medium and three pairs were not exchangeable in D2O medium. The hyperfine splitting of 3.60MHz at θ=0° increased to 3.80MHz at θ=90°. The non-exchangeable signals with 1.73MHz hyperfine splitting at θ=0°, which were assigned to a proton in an amino acid residue, were not detected at θ=90° in oriented PS II or in non-oriented PS II. Highly resolved spectra show that only limited numbers of protons were detected by CW-ENDOR spectra, although many protons were located near the CaMn4O5 cluster. The detected exchangeable protons were proposed to arise from the protons belonging to the water molecules, labeled W1-W4 in the 1.9Å crystal structure, directly ligated to the CaMn4O5 cluster, and nearby amino-acid residue.

Location of Methanol on the S2 State Mn Cluster in Photosystem II Studied by Proton Matrix Electron Nuclear Double Resonance

Proton matrix electron nuclear double resonance (ENDOR) spectroscopy was performed to specify the location of the methanol molecule near the manganese cluster in photosystem II. Comparison of the ENDOR spectra in the presence of CH3OH and CD3OH revealed two pairs of hyperfine couplings, 1.2 MHz for A⊥ and 2.5 MHz for A//, arising from the methyl group in methanol. On the basis of the crystal structure, the possible location of methanol close to the manganese cluster was discussed.

Regulated Electron Tunneling of Photoinduced Primary Charge-Separated State in the Photosystem II Reaction Center

In initial events of the photosynthesis by higher plants, the photosystem II (PSII) generates photoinduced primary charge-separated (CS) state composed of reduced pheophytin (PheoD1-•) and oxidized special pair (P+•) in chlorophyll a (Chla) PD1/PD2 in the D1/D2 heterodimer, ultimately leading to the water oxidation at the oxygen-evolving Mn4CaO5 cluster by P+•. To understand the mechanism of the efficient generation of initially localized CS state (PD1+• PheoD1-•), we have characterized cofactor geometries and electronic coupling of the photoinduced primary CS state in quinone prereduced membrane of PSII from spinach using the time-resolved electron paramagnetic resonance method. It has been revealed that the electronic coupling between the charges is significantly weak in the CS state separated by 1.5 nm, showing the importance of regulated cofactor-cofactor electronic interaction between a vinyl substituent in PheoD1 and an accessory chlorophyll to inhibit the energy-wasting charge recombination after the primary electron-transfer processes.

Magnetic structure of manganese cluster in photosystem II investigated by electron paramagnetic resonance

The electronic structure of manganese (Mn) cluster in photosystem II was investigated by electron paramagnetic resonance (EPR) spectroscopy. In order to determine the spin density distribution in magnetically coupled Mn in the S2 state Mn cluster, pulsed electron–electron double resonance (PELDOR) measurement was performed. The local environment of the Mn cluster was investigated by electron-nuclear double resonance (ENDOR). Using spin projections determined by PELDOR, ENDOR signals were assigned to the water molecules ligated to the Mn cluster. The location of a high-affinity Mn2+ site in apo-photosystem II, which is the initial site of photoactivation of the Mn cluster, was determined by PELDOR.

Hyperfine Sublevel Correlation Spectroscopy Studies of Iron-Sulfur Cluster in Rieske Protein from Green Sulfur Bacterium Chlorobaculum tepidum

The magnetic properties of the Rieske protein purified from Chlorobaculum tepidum were investigated using electron paramagnetic resonance and hyperfine sublevel correlation spectroscopy (HYSCORE). The g-values of the Fe2S2 center were gx = 1.81, gy = 1.90, and gz = 2.03. Four classes of nitrogen signals were obtained by HYSCORE. Nitrogens 1 and 2 had relatively strong magnetic hyperfine couplings and were assigned as the nitrogen directly ligated to Fe. Nitrogens 3 and 4 had relatively weak magnetic hyperfine couplings and were assigned as the other nitrogen of the His ligands and peptide nitrogen connected to the sulfur atom via hydrogen bonding, respectively. The anisotropy of nitrogen 3 reflects the different spin density distributions on the His ligands, which influences the electron transfer to quinone.