Exchange of copper between serum albumin and bleomycin.

1. The present in vitro study shows that bleomycin is able to take up copper from albumin, which functions as a copper transport protein in plasma. 2. Calculations, based on plasma concentrations of Cu(II)-albumin and bleomycin, suggest that only a small fraction of bleomycin (less than 3%) may directly accept copper from Cu(II)-albumin.     

Intestinal metallothionein and the mutual antagonism between copper and zinc in the rat.

A study has been made of the mechanism of the mutual antagonism between copper and zinc in rats. Dietary zinc concentrations of up to 450 mg/kg had no effect on intestinal 64Cu absorption but 900 mg/kg caused a 40% reduction. This was associated with an increase in the mucosal uptake of 64Cu in the small intestine. This occurred mainly in the form of metallothionein and it appeared that copper displaced zinc from the protein after its synthesis had been induced by zinc. Ths intestinal absorption of 65Zn was decreased by 20% when the dietary copper intake was increased from 3 to 24 mg/kg. Further increases in copper intake to 300 mg/kg did not cause any additional decrease in 65Zn absorption or any change in the association of intestinal 65Zn with metallothionein. Concentrations of this protein in the intestinal mucosa were not influenced by dietary copper intake.     

Interaction between nickel and copper in the rat

Two 42-d experiments were conducted with weanling male rats to study interactions between nickel and copper. In Experiment 1, a low-copper basal diet was supplemented with copper at 0 or 30 ppm and nickel at 0 or 30 ppm. Copper was added in Experiment 2 to a basal copper-deficient diet at a level of 0 or 15 ppm and nickel was supplemented at 0, 15, or 225 ppm. Responses to dietary nickel were dependent upon copper nutriture and experimental duration. Nickel had little effect on growth during the first 21 d of either study when added at low levels (15 or 30 ppm) to copper-deficient diets. Nickel supplementation depressed gains between 21 and 42 d in rats fed copper-deficient, but not copper-adequate, diets. Hematocrits and hemoglobin concentrations were not significantly affected by dietary nickel at 21 d. Nickel supplementation decreased hematocrits and hemoglobin values in copper deficient rats at 42 d in Experiment 1, but not in Experiment 2. Absorption of copper apparently was not reduced by nickel, since tissue copper concentrations were generally not decreased by increasing dietary nickel. Nickel supplementation increased lung and heart copper concentrations in Experiment 2. Liver iron was not affected by nickel, but spleen iron concentrations were reduced by nickel supplementation in copper-deficient rats in Experiment 2. The present studies suggest that nickel acts antagonistically to copper in certain biological processes.     

On the copper transfer between dopamine beta-monooxygenase and Cu-thionein

Metallothionein saturated with copper is able to donate copper to apodopamine beta-monooxygenase. The complete recovery of dopamine beta-monooxygenase activity is observed at the molar ratio Cu-thionein/apoenzyme of 25. On the other hand, apothionein accepts copper easily from the holoenzyme.     

Interaction between pesticides and essential metal copper increases the accumulation of copper in the kidneys of rats

Male Sprague-Dawley rats, weighing 175-200 g, six per group were fed AIN 93M diet (CON) or diets containing 500 mg Ca (LCa), 7 mg Zn (LZn), 2 mg Cu (LCu), 60 mg Zn (HZn), or 12 mg Cu (HCu) per kilogram of diet in the following combinations: control (CON), LCa+LZn (LC+Z), LCa+LZn+LCu (LC+Z+C), or HZn+HCu (HZ+C) without or with a pesticides mixture (PM); Endosulfan, Thiram, and Acephate were added to the diets at 25% of LD50/kg. Rats were fed for 2 wk (small intestinal changes) or 4 wk (tissues changes). Plasma Zn was 47% lower than CON in the experimental groups. Plasma Cu and ceruloplasmin concomitantly decreased in the LC+Z+C group and increased with the addition of PM. Kidney Cu was 40% lower in LC+Z+C group, than CON and increased by 31% with PM; in the HZ+C+PM group, kidney Cu was 38% higher than the HZ+C group. Mucosal and small intestines Ca declined by 47% in all experimental groups; PM increased Zn in the LC+Z+C and HZ+C groups; PM further decreased intestinal and mucosal Cu retention in the LC+Z+C and HZ+C groups. Data suggest that low levels of PM in the diet can induce Cu accumulation in the kidney when dietary Zn and Cu are low or high.     

Energetics of copper trafficking between the Atx1 metallochaperone and the intracellular copper transporter, Ccc2

The Atx1 metallochaperone protein is a cytoplasmic Cu(I) receptor that functions in intracellular copper trafficking pathways in plants, microbes, and humans. A key physiological partner of the Saccharomyces cerevisiae Atx1 is Ccc2, a cation transporting P-type ATPase located in secretory vesicles. Here, we show that Atx1 donates its metal ion cargo to the first N-terminal Atx1-like domain of Ccc2 in a direct and reversible manner. The thermodynamic gradient for metal transfer is shallow (K(exchange) = 1.4 +/- 0.2), establishing that vectorial delivery of copper by Atx1 is not based on a higher copper affinity of the target domain. Instead, Atx1 allows rapid metal transfer to its partner. This equilibrium is unaffected by a 50-fold excess of the Cu(I) competitor, glutathione, indicating that Atx1 also protects Cu(I) from nonspecific reactions. Mechanistically, we propose that a low activation barrier for transfer between partners results from complementary electrostatic forces that ultimately orient the metal-binding loops of Atx1 and Ccc2 for formation of copper-bridged intermediates. These thermodynamic and kinetic considerations suggest that copper trafficking proteins overcome the extraordinary copper chelation capacity of the eukaryotic cytoplasm by catalyzing the rate of copper transfer between physiological partners. In this sense, metallochaperones work like enzymes, carefully tailoring energetic barriers along specific reaction pathways but not others.     

Wilson disease: not just a copper disorder. Analysis of a Wilson disease model demonstrates the link between copper and lipid metabolism

Copper is an essential nutrient required for normal growth and development in many organisms. In humans, the disruption of normal copper absorption and excretion is associated with two severe disorders, known as Menkes disease and Wilson disease, respectively. The consequences of insufficient copper supply that is characteristic of Menkes disease have been largely linked to the inactivation of key metabolic enzymes, although other non-enzymatic processes may also be involved. In contrast, the consequences of copper accumulation in Wilson disease have been generally ascribed to copper-induced radical-mediated damage. Recent studies suggest that the cellular response to copper overload, particularly at the early stages of copper accumulation, involves more specific mechanisms and specific pathways. Genetic and metabolic characterization of animal models of Wilson disease has provided new insights into the pre-symptomatic effects of copper that is accumulated in the liver. The studies have uncovered unexpected links between copper metabolism, cell-cycle machinery, and cholesterol biosynthesis. We discuss these new findings along with the earlier reports on dietary effects of copper. Together these experiments suggest a tight link between lipid and copper metabolism and identify several candidate proteins that may mediate the cross-talk between copper status and lipid metabolism.     

Mapping of genes for copper efficiency in rye and the relationship between copper and iron efficiency

A 4B/5R wheat-rye translocation line derived from the Danish wheat variety Viking was revealed to be highly copper efficient. The chromosomal exchange includes a very small terminal segment of chromosome arm 5RL of rye which was physically mapped by genomic DNA: DNA in situ hybridization and chromosome analysis. The gene for Cu efficiency (Ce) is linked to a dominant hairy neck character from rye (Ha1) and to two rye-specific leaf esterase loci (Est6, Est7), all of which are postulated to map to the distal part of 5RL. Genes coding for mugineic acid synthetase and 3-hydroxymugineic acid synthetase also on chromosome 5R are not included in the 4B/5R translocation and hence map outside the terminal 5R region. These genetic and molecular markers can be useful tools for large-scale screening in wheat breeding programmes.     

Complex formation between the copper protein, azurin and the cytochrome c peroxidase of Pseudomonas aeruginosa.

Reduced azurin reacts with the resting, oxidized cytochrome c peroxidase of Pseudomonas aeruginosa to yield time courses observed at 420 nm, which consist of the sum of two exponential processes. Each process exhibits a hyperbolic dependence of the observed rate constant on the reduced azurin concentration. The fraction of the total optical density change which each process contributes is found to be dependent on the reduced azurin concentration. This pattern of reactivity is maintained at pH values between 5.5 and 8.0. The data has been analyzed in terms of a complex formation between the two proteins followed by an intramolecular electron exchange reaction. This analysis yields values for the binding constants at each pH value. The intramolecular exchange reaction is independent of pH, whilst the pH dependence of the binding reaction suggests the involvement of a histidine residue in this process.     

Understanding copper trafficking in bacteria: interaction between the copper transport protein CopZ and the N-terminal domain of the copper ATPase CopA from Bacillus subtilis

In this paper the interaction of cytoplasmic CopZ and the N-terminal domain of the CopA ATPase from Bacillus subtilis has been studied by NMR through (15)N-(1)H HSQC experiments in order to understand the role of the two proteins in the whole copper trafficking mechanism of the bacteria. It appears that the two proteins interact in a fashion similar to that of the yeast homologue proteins [Arnesano, F., Banci, L., Bertini, I., Cantini, F., Ciofi-Baffoni, S., Huffman, D. L., and O'Halloran, T. V. (2001) J. Biol. Chem. 276, 41365-41376], although the surface potentials are reversed. A structural model for the interaction is proposed. (15)N mobility studies on the free proteins and on their complex are also reported. From these data, it appears that copper is largely transferred from CopZ to CopA, thus suggesting their possible involvement in a detoxification process. Comparing functional data of homologous proteins of other bacteria, it can be concluded that this class of proteins is involved in copper homeostasis but the specific roles are species dependent.     

A study of copper treatment and tissue copper levels in the murine congenital copper deficiency, mottled.

The injection of copper chloride overcomes the lethality and pigment deficiency in the brindled ($\text{Mo}$^br) mouse mutant but copper levels remain depressed in the liver and brain, and a further accumulation occurs in the kidney. The copper-dependent synthesis of brain noradrenaline returns to normal but the activity of brain cytochrome c oxidase, although increased, remains depressed. Significant changes in tissue copper content of female brindled heterozygotes are reported and in each case, the changes exceed those expected on the basis of X-inactivation. The significance of these results to the development of a satisfactory treatment regime for this disease is discussed.     

The relation between selenium, zinc and copper concentration and the trace element dependent antioxidative status.

An imbalance in the antioxidative system was connected with the development of a number of pathological processes. In order to receive values of a healthy group and to evaluate pathological changes of the trace element dependent antioxidative status in future, we investigated 99 healthy volunteers (45 male and 54 female, mean age 37.4 +/- 11.7 years). We determined the concentrations of Se, Cu and Zn, the concentrations of malondialdehyde (MDA) and the activities of the Se dependent glutathione peroxidase (GSH-Px) and the Zn/Cu dependent superoxide dismutase (SOD). The plasma concentrations (mean +/- SD) for Se, Cu and Zn were 0.84 +/- 0.10 micromol/l, 15.6 +/- 2.78 micromol/l and 12.6 +/- 1.80 micromol/l, resp., and for non protein-bound and protein bound MDA 0.27 +/- 0.07 micromol/l and 1.11 +/- 0.25 micromol/l, resp. The activity of GSH-Px in plasma and erythrocytes was 130 +/- 20.8 U/l and 19.8 +/- 4.18 U/mg Hb, resp. and of SOD in erythrocytes 3,159 +/- 847.2 U/g Hb. In plasma positive correlations have been found between Se concentrations and GSH-Px activities (p < 0.002, r = 0.31) and between GSH-Px activities and concentrations of non protein-bound MDA (p = 0.004, r = 0.28). A negative correlation has been observed between GSH-Px activities in plasma and in erythrocytes. The higher the concentrations of Cu in erythrocytes, the higher were the activities of SOD (p = 0.03, r = 0.22) and GSH-Px in erythrocytes (r = 0.26, p = 0.01), while an increasing activity of GSH-Px in these cells correlated with a decreasing concentration of non protein-bound MDA (r = -0,31, p = 0.002). An increase in BMI was connected with an increase in protein-bound MDA and a decrease in GSH-Px activities in pLasma (p = 0.002 and r = 0.23). As the results demonstrate, Se and Cu concentrations in erythrocytes can improve the trace element dependent antioxidative status.