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51.
Marcus Bär Joachim Klaer Lothar Weinhardt Regan G. Wilks Stefan Krause Monika Blum Wanli Yang Clemens Heske Hans‐Werner Schock 《Liver Transplantation》2013,3(6):777-781
The surface properties of CuInS2 (CIS) thin‐film solar cell absorbers are investigated by a combination of electron and soft X‐ray spectroscopies. Spatially separated regions of varying colors are observed and identified to be dominated by either CuS or Cu2S surface phases. After their removal by KCN etching, the samples cannot be distinguished by eye and the CIS surface is found to be Cu‐deficient in both regions. However, a significantly more pronounced off‐stoichiometry in the region initially covered by Cu2S can be identified. In this region, the resulting surface band gap is also significantly larger than the EgSurf of the initially CuS‐terminated region. Such variations may represent a hidden parameter which, if overlooked, induces irreproducibility and thus prevents systematic optimization efforts. 相似文献
52.
J. Esclapez B. ZafrillaR.M. Martínez-Espinosa M.J. Bonete 《Biochimica et Biophysica Acta - Proteins and Proteomics》2013,1834(6):1003-1009
The green Cu-NirK from Haloferax mediterranei (Cu-NirK) has been expressed, refolded and retrieved as a trimeric enzyme using an expression method developed for halophilic Archaea. This method utilizes Haloferax volcanii as a halophilic host and an expression vector with a constitutive and strong promoter. The enzymatic activity of recombinant Cu-NirK was detected in both cellular fractions (cytoplasmic fraction and membranes) and in the culture media. The characterization of the enzyme isolated from the cytoplasmic fraction as well as the culture media revealed important differences in the primary structure of both forms indicating that Hfx. mediterranei could carry out a maturation and exportation process within the cell before the protein is exported to the S-layer. Several conserved signals found in Cu-NirK from Hfx. mediterranei sequence indicate that these processes are closely related to the Tat system. Furthermore, the N-terminal sequence of the two Cu-NirK subunits constituting different isoforms revealed that translation of this protein could begin at two different points, identifying two possible start codons. The hypothesis proposed in this work for halophilic Cu-NirK processing and exportation via the Tat system represents the first approximation of this mechanism in the Halobacteriaceae family and in Prokarya in general. 相似文献
53.
54.
AbstractSublethal concentrations of copper in water cause the degeneration of olfactory receptors in rainbow trout (Oncorhynchus mykiss). Receptor cell loss has been correlated to the loss of olfaction in fish and may cause difficulties in olfactory mediated behaviors such as migration. This study investigated the effects of three levels of copper (100, 75 and 50 mg L?1) on the olfactory epithelium of rainbow trout. Twenty fish randomly allocated between three exposure groups and one control were exposed for 24 hours under static renewal conditions. Light and scanning electron microscopic observations of olfactory tissue were taken to determine the extent of degeneration of receptors. In addition, levels of copper and zinc in the brain tissues were analyzed to determine if the olfactory route was a significant route of copper exposure and transfer to fish brain tissue. Results indicate that degeneration of receptors is related to the concentration of copper. Levels of copper in brain were found to be below detection of the instrument. Levels of zinc were extremely variable ranging from 52 to 132 ng zinc g?1 brain tissue. 相似文献
55.
Copper complexing capacity in fresh-waters using the catechol-cathodic stripping voltammetric method
AbstractThe catechol-cathodic stripping voltammetric (CSV) method for measurement of the copper complexing capacity of marine and estuarine waters, has been modified and applied to fresh-waters. The optimum catechol concentration needed was the major difference between the two methods. It was found necessary to separately optimise the catechol concentration for each of the three fresh-water samples tested, presumably because of the much greater differences in the concentration of complexing ligands in fresh-water compared with marine or estuarine waters. This limits the usefulness of the CSV method for the determination of the complexing capacity of fresh-waters.The total ligand concentration ([Lt]) and conditional stability constants (*K) derived for the three fresh-waters are compared with similar data obtained using an anodic stripping voltammetric (ASV) method. The set of copper-binding ligands determined using the CSV method were slightly lower in concentration, but stronger binding, than those obtained using the ASV method (CSV: [Lt] 0.04–0.06 μM, log K(pH 6) 8.9–9.4; ASV: [Lt] 0.11–0.17 μM, log *K(pH 6) 7.9–8.1). 相似文献
56.
Maricarmen Hernández‐Rodríguez José Correa‐Basurto Claudia G. Benitez‐Cardoza Aldo Arturo Resendiz‐Albor Martha C. Rosales‐Hernández 《Protein science : a publication of the Protein Society》2013,22(10):1320-1335
The formation of fibrils and oligomers of amyloid beta (Aβ) with 42 amino acid residues (Aβ1–42) is the most important pathophysiological event associated with Alzheimer''s disease (AD). The formation of Aβ fibrils and oligomers requires a conformational change from an α-helix to a β-sheet conformation, which is encouraged by the formation of a salt bridge between Asp 23 or Glu 22 and Lys 28. Recently, Cu2+ and various drugs used for AD treatment, such as galanthamine (Reminyl®), have been reported to inhibit the formation of Aβ fibrils. However, the mechanism of this inhibition remains unclear. Therefore, the aim of this work was to explore how Cu2+ and galanthamine prevent the formation of Aβ1–42 fibrils using molecular dynamics (MD) simulations (20 ns) and in vitro studies using fluorescence and circular dichroism (CD) spectroscopies. The MD simulations revealed that Aβ1–42 acquires a characteristic U-shape before the α-helix to β-sheet conformational change. The formation of a salt bridge between Asp 23 and Lys 28 was also observed beginning at 5 ns. However, the MD simulations of Aβ1−42 in the presence of Cu2+ or galanthamine demonstrated that both ligands prevent the formation of the salt bridge by either binding to Glu 22 and Asp 23 (Cu2+) or to Lys 28 (galanthamine), which prevents Aβ1−42 from adopting the U-characteristic conformation that allows the amino acids to transition to a β-sheet conformation. The docking results revealed that the conformation obtained by the MD simulation of a monomer from the 1Z0Q structure can form similar interactions to those obtained from the 2BGE structure in the oligomers. The in vitro studies demonstrated that Aβ remains in an unfolded conformation when Cu2+ and galanthamine are used. Then, ligands that bind Asp 23 or Glu 22 and Lys 28 could therefore be used to prevent β turn formation and, consequently, the formation of Aβ fibrils. 相似文献
57.
Abstract Copper is a vital component of electron transfer reactions mediated by proteins such as superoxide dismutase, cytochrome c oxidase and plastocyanin, but its concentrations in the cells needs to be maintained at low levels. In fact, the same ability of this essential metal ion to transfer electrons can also make it toxic to cells when present in excess. In vitro cultured explants of Nicotiana have been extensively used as a model to analyse metal-DNA interactions. In this report, we examined the effect of copper (1, 10 and 100 μM CuSO4) on callus growth and protein synthesis of in vitro-cultured pith explants of Nicotiana glauca. In addition, a N. glauca cDNA library from Cu-treated (100 μM CuSO4) pith explants cultured in vitro for 24 h was analysed by mRNA differential screening. The copper treatments inhibited callus growth of pith explants. The extent of inhibition was directly correlated to metal concentration. One and 10 μM CuSO4 induced a notable increase of proteins synthesis relative to control explants. By contrast, 100 μM CuSO4 inhibited protein synthesis relative to control extracts. The SDS-PAGE fluorography of pith proteins revealed, in Cu-treated extracts qualitative and/or quantitative differences in the synthesis of some polypeptides compared with control explants. Copper-modulated patterns of gene expression were also analysed by mRNA differential screening. The N. glauca genes isolated from Cu-treated pith explants shared common identities with other genes known to be elicited by diverse stresses, including pathogenesis and abiotic stress. In particular, the cDNAs were homologues to genes encoding cell wall proteins (i.e., extensin, and arabinogalactan-protein) and pathogenesis-related proteins (i.e., osmotin, endochitinase and a member of the Systemic Acquired Resistance gene family). In addition, an MD-2-related lipid-recognition (ML) domain protein and the enzyme S-adenosyl-L-homocysteine (AdoHcy) hydrolase appeared involved in the response to copper stress. In animal cells, AdoHcy hydrolase is a copper binding protein in vivo, which suggests that, also in plant tissues, this enzyme may play an important role in regulating the levels and intracellular distribution of copper. 相似文献
58.
Michal S. Shoshan Edit Y. Tshuva Deborah E. Shalev 《Journal of visualized experiments : JoVE》2013,(82)
Copper (I) binding by metallochaperone transport proteins prevents copper oxidation and release of the toxic ions that may participate in harmful redox reactions. The Cu (I) complex of the peptide model of a Cu (I) binding metallochaperone protein, which includes the sequence MTCSGCSRPG (underlined is conserved), was determined in solution under inert conditions by NMR spectroscopy.NMR is a widely accepted technique for the determination of solution structures of proteins and peptides. Due to difficulty in crystallization to provide single crystals suitable for X-ray crystallography, the NMR technique is extremely valuable, especially as it provides information on the solution state rather than the solid state. Herein we describe all steps that are required for full three-dimensional structure determinations by NMR. The protocol includes sample preparation in an NMR tube, 1D and 2D data collection and processing, peak assignment and integration, molecular mechanics calculations, and structure analysis. Importantly, the analysis was first conducted without any preset metal-ligand bonds, to assure a reliable structure determination in an unbiased manner. 相似文献
59.
《MABS-AUSTIN》2013,5(5):901-911
Fragmentation in the hinge region of an IgG1 monoclonal antibody (mAb) can affect product stability, potentially causing changes in potency and efficacy. Metals ions, such as Cu2+, can bind to the mAb and undergo hydrolysis or oxidation, which can lead to cleavage of the molecule. To better understand the mechanism of Cu2+-mediated mAb fragmentation, hinge region cleavage products and their rates of formation were studied as a function of pH with and without Cu2+. More detailed analysis of the chemical changes was investigated using model linear and cyclic peptides (with the sequence of SCDKTHTC) derived from the upper hinge region of the mAb. Cu2+ mediated fragmentation was determined to be predominantly via a hydrolytic pathway in solution. The sites and products of hydrolytic cleavage are pH and strain dependent. In more acidic environments, rates of Cu2+ induced hinge fragmentation are significantly slower than at higher pH. Although the degradation reaction rates between the linear and cyclic peptides are not significantly different, the products of degradation vary. mAb fragmentation can be reduced by modifying His, which is a potential metal binding site and a known ligand in other metalloproteins. These results suggest that a charge may contribute to stabilization of a specific molecular structure involved in hydrolysis, leading to the possible formation of a copper binding pocket that causes increased susceptibility of the hinge region to degradation. 相似文献
60.
Structure and coordination of CuB in the Acidianus ambivalens aa
3 quinol oxidase heme–copper center
Tiago M. Bandeiras Manuel M. Pereira Miguel Teixeira Pierre Moenne-Loccoz Ninian J. Blackburn 《Journal of biological inorganic chemistry》2005,10(6):625-635
The coordination environment of the CuB center of the quinol oxidase from Acidianus ambivalens, a type B heme–copper oxygen reductase, was investigated by Fourier transform (FT) IR and extended X-ray absorption fine
structure (EXAFS) spectroscopy. The comparative structural chemistry of dinuclear Fe–Cu sites of the different types of oxygen
reductases is of great interest. Fully reduced A. ambivalens quinol oxidase binds CO at the heme a
3 center, with ν(CO)=1,973 cm−1. On photolysis, the CO migrated to the CuB center, forming a CuBI–CO complex with ν(CO)=2,047 cm−1. Raising the temperature of the samples to 25°C did not result in a total loss of signal in the FTIR difference spectrum
although the intensity of these signals was reduced sevenfold. This observation is consistent with a large energy barrier
against the geminate rebinding of CO to the heme iron from CuB, a restricted limited access at the active-site pocket for a second binding, and a kinetically stable CuB–CO complex in A. ambivalens aa
3. The CuB center was probed in a number of different states using EXAFS spectroscopy. The oxidized state was best simulated by three
histidines and a solvent O scatterer. On reduction, the site became three-coordinate, but in contrast to the bo
3 enzyme, there was no evidence for heterogeneity of binding of the coordinated histidines. The CuB centers in both the oxidized and the reduced enzymes also appeared to contain substoichiometric amounts (0.2 mol equiv) of
nonlabile chloride ion. EXAFS data of the reduced carbonylated enzyme showed no difference between dark and photolyzed forms.
The spectra could be well fit by 2.5 imidazoles, 0.5 Cl− and 0.5 CO ligands. This arrangement of scatterers would be consistent with about half the sites remaining as unligated Cu(his)3 and half being converted to Cu(his)2Cl−CO, a 50/50 ratio of Cu(his)2Cl− and Cu(his)3CO, or some combination of these formulations.
Electronic Supplementary Material Supplementary material is available for this article at . 相似文献