RESUMO
Photosystem II (PSII) is a huge membrane-protein complex consisting of 20 different subunits with a total molecular mass of 350 kDa for a monomer. It catalyses light-driven water oxidation at its catalytic centre, the oxygen-evolving complex (OEC). The structure of PSII has been analysed at 1.9 Å resolution by synchrotron radiation X-rays, which revealed that the OEC is a Mn4CaO5 cluster organized in an asymmetric, 'distorted-chair' form. This structure was further analysed with femtosecond X-ray free electron lasers (XFEL), providing the 'radiation damage-free' structure. The mechanism of O=O bond formation, however, remains obscure owing to the lack of intermediate-state structures. Here we describe the structural changes in PSII induced by two-flash illumination at room temperature at a resolution of 2.35 Å using time-resolved serial femtosecond crystallography with an XFEL provided by the SPring-8 ångström compact free-electron laser. An isomorphous difference Fourier map between the two-flash and dark-adapted states revealed two areas of apparent changes: around the QB/non-haem iron and the Mn4CaO5 cluster. The changes around the QB/non-haem iron region reflected the electron and proton transfers induced by the two-flash illumination. In the region around the OEC, a water molecule located 3.5 Å from the Mn4CaO5 cluster disappeared from the map upon two-flash illumination. This reduced the distance between another water molecule and the oxygen atom O4, suggesting that proton transfer also occurred. Importantly, the two-flash-minus-dark isomorphous difference Fourier map showed an apparent positive peak around O5, a unique µ4-oxo-bridge located in the quasi-centre of Mn1 and Mn4 (refs 4,5). This suggests the insertion of a new oxygen atom (O6) close to O5, providing an O=O distance of 1.5 Å between these two oxygen atoms. This provides a mechanism for the O=O bond formation consistent with that proposed previously.
Assuntos
Cristalografia/métodos , Elétrons , Lasers , Luz , Oxigênio/química , Oxigênio/efeitos da radiação , Complexo de Proteína do Fotossistema II/química , Complexo de Proteína do Fotossistema II/efeitos da radiação , Biocatálise/efeitos da radiação , Cianobactérias/química , Transporte de Elétrons/efeitos da radiação , Análise de Fourier , Manganês/química , Manganês/metabolismo , Modelos Moleculares , Ferroproteínas não Heme/química , Ferroproteínas não Heme/metabolismo , Ferroproteínas não Heme/efeitos da radiação , Oxigênio/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Prótons , Temperatura , Fatores de Tempo , Água/química , Água/metabolismoRESUMO
The inactive, nitrosyl bound form of Fe-type nitrile hydratase (NHase) contains two active site cysteine residues that are post-translationally modified to sulfenate (SO-) and sulfinate (SO2-) ligands. DFT and INDO/S calculations support the hypothesis that these unusual modifications play a key role in modulating the electronic absorption spectra and photoreactivity of the Fe(III) centre in the enzyme.
Assuntos
Hidroliases/efeitos da radiação , Processamento de Proteína Pós-Traducional/fisiologia , Sítios de Ligação , Cisteína/metabolismo , Hidroliases/química , Hidroliases/metabolismo , Luz , Modelos Moleculares , Ferroproteínas não Heme/química , Ferroproteínas não Heme/metabolismo , Ferroproteínas não Heme/efeitos da radiação , Ácidos Sulfênicos , Ácidos SulfínicosRESUMO
We examined the influence of the level of dietary protein or vitamin E (VE) on oxidative damage to DNA, lipids, and protein in the liver after total body irradiation (TBI) with X-rays at 1 or 4 Gy. Levels of 8-hydroxydeoxyguanosine, thiobarbituric acid-reactive substances, and protein carbonyls in the liver did not differ among the groups that did not receive TBI. However, oxidative damage to lipids and protein was increased by TBI only in the 1% protein group. DNA damage, lipid peroxidation, or protein oxidation in the liver was increased by TBI in a dose-dependent manner, and the damage was consistently higher in the 1% than in the 20% protein group. In the 1% protein group, a greater decrease in relative spleen weight by TBI was also observed. Concentrations of antioxidants (vitamins C and E and glutathione) in the liver were lower and the concentration of nonheme iron in the liver was higher in the 1% than in the 20% protein group. Mice fed a 1% protein diet became susceptible to TBI-induced oxidative damage, and decreases in antioxidant levels and an increase in iron level were involved in the mechanism of this susceptibility. These results suggest that dietary VE and protein can prevent oxidative damage to DNA, lipid, and protein in mice subjected to TBI. Consumption of a VE-free diet significantly increased 8-hydroxydeoxyguanosine levels in DNA from mice fed the 1% protein diet with TBI, but such changes were not detected in DNA from mice fed the 20% protein diet.