The electronic structures of the native Mn(4)O(x)Ca cluster and the biosynthetically substituted Mn(4)O(x)Sr cluster of the oxygen evolving complex (OEC) of photosystem II (PSII) core complexes isolated from Thermosynechococcus elongatus, poised in the S(2) state, were studied by X- and Q-band CW-EPR and by pulsed Q-band (55)Mn-ENDOR spectroscopy. Both wild type and tyrosine D less mutants grown photoautotrophically in either CaCl(2) or SrCl(2) containing media were measured. The obtained CW-EPR spectra of the S(2) state displayed the characteristic, clearly noticeable differences in the hyperfine pattern of the multiline EPR signal [Boussac et al. J. Biol. Chem.2004, 279, 22809-22819]. In sharp contrast, the manganese ((55)Mn) ENDOR spectra of the Ca and Sr forms of the OEC were remarkably similar. Multifrequency simulations of the X- and Q-band CW-EPR and (55)Mn-pulsed ENDOR spectra using the Spin Hamiltonian formalism were performed to investigate this surprising result. It is shown that (i) all four manganese ions contribute to the (55)Mn-ENDOR spectra; (ii) only small changes are seen in the fitted isotropic hyperfine values for the Ca(2+) and Sr(2+) containing OEC, suggesting that there is no change in the overall spin distribution (electronic coupling scheme) upon Ca(2+)/Sr(2+) substitution; (iii) the changes in the CW-EPR hyperfine pattern can be explained by a small decrease in the anisotropy of at least two hyperfine tensors. It is proposed that modifications at the Ca(2+) site may modulate the fine structure tensor of the Mn(III) ion. DFT calculations support the above conclusions. Our data analysis also provides strong support for the notion that in the S(2) state the coordination of the Mn(III) ion is square-pyramidal (5-coordinate) or octahedral (6-coordinate) with tetragonal elongation. In addition, it is shown that only one of the currently published OEC models, the Siegbahn structure [Siegbahn, P. E. M. Acc. Chem. Res.2009, 42, 1871-1880, Pantazis, D. A. et al. Phys. Chem. Chem. Phys.2009, 11, 6788-6798], is consistent with all data presented here. These results provide important information for the structure of the OEC and the water-splitting mechanism. In particular, the 5-coordinate Mn(III) is a potential site for substrate 'water' (H(2)O, OH(-)) binding. Its location within the cuboidal structural unit, as opposed to the external 'dangler' position, may have important consequences for the mechanism of O-O bond formation.

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The Gly-His-Lys (GHK) peptide and the Asp-Ala-His-Lys (DAHK) sequences are naturally occurring high-affinity copper(II) chelators found in the blood plasma and are hence of biological interest. A structural study of the copper complexes of these peptides was conducted in the solid state and in solution by determining their X-ray structures, and by using a large range of spectroscopies, including EPR and HYSCORE (hyperfine sub-level correlation), X-ray absorption and (1)H and (13)C NMR spectroscopy. The results indicate that the structures of [Cu(II)(DAHK)] in the solid state and in solution are similar and confirm the equatorial coordination sphere of NH(2), two amidyl N and one imidazole N. Additionally, a water molecule is bound apically to Cu(II) as revealed by the X-ray structure. As reported previously in the literature, [Cu(II)(GHK)], which exhibits a dimeric structure in the solid state, forms a monomeric complex in solution with three nitrogen ligands: NH(2), amidyl and imidazole. The fourth equatorial site is occupied by a labile oxygen atom from a carboxylate ligand in the solid state. We probe that fourth position and study ternary complexes of [Cu(II)(GHK)] with glycine or histidine. The Cu(II) exchange reaction between different DAHK peptides is very slow, in contrast to [Cu(II)(GHK)], in which the fast exchange was attributed to the presence of a [Cu(II)(GHK)(2)] complex. The redox properties of [Cu(II)(GHK)] and [Cu(II)(DAHK)] were investigated by cyclic voltammetry and by measuring the ascorbate oxidation in the presence of molecular oxygen. The measurements indicate that both Cu(II) complexes are inert under moderate redox potentials. In contrast to [Cu(II)(DAHK)], [Cu(II)(GHK)] could be reduced to Cu(I) around -0.62 V (versus AgCl/Ag) with subsequent release of the Cu ion. These complete analyses of structure and redox activity of those complexes gave new insights with biological impact and can serve as models for other more complicated Cu(II)-peptide interactions.

The electronic properties of the Mn(4)O(x)Ca cluster in the S(2) state of the oxygen-evolving complex (OEC) were studied using X- and Q-band EPR and Q-band (55)Mn-ENDOR using photosystem II preparations isolated from the thermophilic cyanobacterium T. elongatus and higher plants (spinach). The data presented here show that there is very little difference between the two species. Specifically it is shown that: (i) only small changes are seen in the fitted isotropic hyperfine values, suggesting that there is no significant difference in the overall spin distribution (electronic coupling scheme) between the two species; (ii) the inferred fine-structure tensor of the only Mn(III) ion in the cluster is of the same magnitude and geometry for both species types, suggesting that the Mn(III) ion has the same coordination sphere in both sample preparations; and (iii) the data from both species are consistent with only one structural model available in the literature, namely the Siegbahn structure [Siegbahn, P. E. M. Accounts Chem. Res.2009, 42, 1871-1880, Pantazis, D. A. et al., Phys. Chem. Chem. Phys.2009, 11, 6788-6798]. These measurements were made in the presence of methanol because it confers favorable magnetic relaxation properties to the cluster that facilitate pulse-EPR techniques. In the absence of methanol the separation of the ground state and the first excited state of the spin system is smaller. For cyanobacteria this effect is minor but in plant PS II it leads to a break-down of the S(T)=½ spin model of the S(2) state. This suggests that the methanol-OEC interaction is species dependent. It is proposed that the effect of small organic solvents on the electronic structure of the cluster is to change the coupling between the outer Mn (Mn(A)) and the other three Mn ions that form the trimeric part of the cluster (Mn(B), Mn(C), Mn(D)), by perturbing the linking bis-μ-oxo bridge. The flexibility of this bridging unit is discussed with regard to the mechanism of O-O bond formation.

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Premium bonds: The pH‐dependent coordination of CuII to the Alzheimer′s disease amyloid‐β peptide has been studied by NMR spectroscopy. Several equivalent ligands are in equilibrium for CuII binding near pH 6.6 and 8.7. Fewer conformers are detected at high pH, in line with a reshuffling of the CuII binding site induced by deprotonation of the Asp1Ala2 peptide bond (see picture).

The Kae1 (Kinase-associated endopeptidase 1) protein is a member of the recently identified transcription complex EKC and telomeres maintenance complex KEOPS in yeast. Kae1 homologues are encoded by all sequenced genomes in the three domains of life. Although annotated as putative endopeptidases, the actual functions of these universal proteins are unknown. Here we show that the purified Kae1 protein (Pa-Kae1) from Pyrococcus abyssi is an iron-protein with a novel type of ATP-binding site. Surprisingly, this protein did not exhibit endopeptidase activity in vitro but binds cooperatively to single and double-stranded DNA and induces unusual DNA conformational change. Furthermore, Pa-Kae1 exhibits a class I apurinic (AP)-endonuclease activity (AP-lyase). Both DNA binding and AP-endonuclease activity are inhibited by ATP. Kae1 is thus a novel and atypical universal DNA interacting protein whose importance could rival those of RecA (RadA/Rad51) in the maintenance of genome integrity in all living cells.

Multifrequency (95, 190, and 285 GHz) high-field electron paramagnetic resonance (EPR) spectroscopy has been used to characterize radical intermediates in wild-type and Trp191Gly mutant cytochrome c peroxidase (CcP). The high-field EPR spectra of the exchange-coupled oxoferryl--trytophanyl radical pair that constitutes the CcP compound I intermediate [(Fe(IV)=O) Trp*(+)] were analyzed using a spin Hamiltonian that incorporated a general anisotropic spin-spin interaction term. Perturbation expressions of this Hamiltonian were derived, and their limitations under high-field conditions are discussed. Using numerical solutions of the completely anisotropic Hamiltonian, its was possible to simulate accurately the experimental data from 9 to 285 GHz using a single set of spin parameters. The results are also consistent with previous 9 GHz single-crystal studies. The inherent superior resolution of high-field EPR spectroscopy permitted the unequivocal detection of a transient tyrosyl radical that was formed 60 s after the addition of 1 equiv of hydrogen peroxide to the wild-type CcP at 0 degrees C and disappeared after 1 h. High-field EPR was also used to characterize the radical intermediate that was generated by hydrogen peroxide addition to the W191G CcP mutant. The g- values of this radical (g(x)= 2.00660, g(y) = 2.00425, and g(z)= 2.00208), as well as the wild-type transient tyrosyl radical, are essentially identical to those obtained from the high-field EPR spectra of the tyrosyl radical generated by gamma-irradiation of crystals of tyrosine hydrochloride (g(x)= 2.00658, g(y) = 2.00404, and g(z) = 2.00208). The low g(x)-value indicated that all three of the tyrosyl radicals were in electropositive environments. The broadening of the g(x) portion of the HF-EPR spectrum further indicated that the electrostatic environment was distributed. On the basis of these observations, possible sites for the tyrosyl radical(s) are discussed.