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X-ray absorption spectroscopy was used to measure the damage caused by exposure to x-rays to the Mn4Ca active site in single crystals of photosystem II as a function of dose and energy of x-rays, temperature, and time. These studies reveal that the conditions used for structure determination by x-ray crystallography cause serious damage specifically to the metal-site structure. The x-ray absorption spectra show that the structure changes from one that is characteristic of a high-valent Mn4(III2,IV2) oxo-bridged Mn4Ca cluster to that of Mn(II) in aqueous solution. This damage to the metal site occurs at a dose that is more than one order of magnitude lower than the dose that results in loss of diffractivity and is commonly considered safe for protein crystallography. These results establish quantitative x-ray dose parameters that are applicable to redox-active metalloproteins. This case study shows that a careful evaluation of the structural intactness of the active site(s) by spectroscopic techniques can validate structures derived from crystallography and that it can be a valuable complementary method before structure–function correlations of metalloproteins can be made on the basis of high-resolution x-ray crystal structures. [ABSTRACT FROM AUTHOR]

Abstract: The antenna proteins in photosystem II (PSII) not only promote energy transfer to the photosynthetic reaction center (RC) but provide also an efficient cation sink to re-reduce chlorophyll a if the electron transfer (ET) from the Mn-cluster is inhibited. Using the newest PSII dimer crystal structure (3.0 Å resolution), in which 11 β-carotene molecules (Car) and 14 lipids are visible in the PSII monomer, we calculated the redox potentials (E m) of one-electron oxidation for all Car (E m(Car)) by solving the Poisson–Boltzmann equation. In each PSII monomer, the D1 protein harbors a previously unlocated Car (CarD1) in van der Waals contact with the chlorin ring of ChlZ(D1). Each CarD1 in the PSII dimer complex is located in the interface between the D1 and CP47 subunits, together with another four Car of the other PSII monomer and several lipid molecules. The proximity of Car bridging between CarD1 and plastoquinone/QA may imply a direct charge recombination of Car+QA −. The calculated E m(CarD1) and E m(ChlZ(D1)) are, respectively, 83 and 126 mV higher than E m(CarD2) and E m(ChlZ(D2)), which could explain why CarD2 + and ChlZ(D2) + are observed rather than the corresponding CarD1 + and ChlZ(D1) +. [Copyright &y& Elsevier]

Oxygenic photosynthesis takes place in the thylakoid membrane of cyanobacteria, algae, and higher plants. Initially light is absorbed by an oligomeric pigment-protein complex designated as photosystem II (PSII), which catalyzes light-induced water cleavage under release of molecular oxygen for the biosphere on our planet. The membrane-extrinsic manganese stabilizing protein (PsbO) is associated on the lumenal side of the thylakoids close to the redox-active (Mn)4Ca cluster at the catalytically active site of PSII. Recombinant PsbO from the thermophilic cyanobacterium Thermosynechococcus elongatus was expressed in Escherichia coli and spectroscopically characterized. The secondary structure of recombinant PsbO (recPsbO) was analyzed in the absence and presence of Ca2+ using Fourier transform infrared spectroscopy (FTIR) and circular dichroism spectropolarimetry (CD). No significant structural changes could be observed when the PSII subunit was titrated with Ca2+ in vitro. These findings are compared with data for spinach PsbO. Our results are discussed in the light of the recent 3D-structural analysis of the oxygen-evolving PSII and structural/thermodynamic differences between the two homologous proteins from thermophilic cyanobacteria and plants. [ABSTRACT FROM AUTHOR]

Presents a study which used the light-harvesting proteins of photosystem II to investigate the relationship between chlorophyll fluorescence quenching and the binding of dicyclohexylcarbodiinide (DCCD). Information on photosystem II's light-harvesting system; Details on DCCD; Method used for the study; What was shown by the results of the study.