Molecular Weight Metal-binding Proteins Research Articles

Evidence of presence of a low molecular weight, non-metallothionein-like metal-binding protein in the marine gastropod Nassarius reticulatus L.

1. 1. A low molecular weight metal-binding protein was found in the snail Nassarius reticulatus cytosol, which was induced in heavy metal contaminated environments. 2. 2. In our sodium dodecyl sulfate-mercaptoethanol polyacrylamide gel systems it behaved as a protein of 19 kDa mol. wt. 3. 3. Amino acid composition studies definitely established this protein not to be metallothionein (Mt) like, because it had a much lower level of cysteine and substantial amounts of aromatic amino acids and histidine. 4. 4. The metal-binding strength of this protein was concluded to be much weaker than that of Mt. 5. 5. In the crustacean Pagurus bernhardus L. such a protein could not be demonstrated. 6. 6. In both the snail and the crustacean Zn may inhibit the accumulation of Hg. The premise for studying the induction of the metal-binding Nassarius protein as a supplement to environmental metal monitoring purposes is briefly discussed.

A fish hepatocyte model for the investigation of the effects of environmental contaminants

An isolated fish hepatocyte culture system was developed as a models ystem to investigate the mechanisms of action of environmental contaminants. Hepatocytes were isolated from striped bass ( Morone saxatilis) by an adaptation of the two stage perfusion technique of Seglen. 1 The system was used to evaluate metal binding protein (MBP) induction in response to cadmium, a primary inducer of metallothionein (MT) in rat hepatocytes. Striped bass hepatocytes appeared to be refractory to the induction of MBP by cadmium, since there was no significant increase in the synthesis of MBP for any of the doses at any of the time points investigated. However, when a similar experiment was performed using rat hepatocytes there was induction of MBP that was related to both dose and time. These comparative experiments indicate that although there are similarities between the hepatocytes of the two species in regard to 35S incorporation into low molecular weight metal-binding proteins, there appear to be significant quantitative differences as well in regard to MBP kinetics. This in vitro model system could potentially be utilized to investigate the toxicological properties of other environmental contaminants.

Evidence of presence of heavy metal-binding proteins in polychaeta species

1. 1. Four groups of polychaete worms, differing in feeding habits, have been studied with respect to presence of metal-binding proteins. 2. 2. The presence of low molecular weight metal-binding proteins was clearly demonstrated in the species Lumbrineris fragilis as well as in Harmothoë sp. both being carnivorous animals. One of the proteins found in L. fragilis was similar to mammalian metallothionein (Mt).

Partitioning of metals among metal-binding proteins in the bay mussel, Mytilus edulis

Metabolism of metals was assessed in populations of Mytilus edulis exposed to metals in the laboratory and in the field. Chronic exposure to metals in both environments resulted in increased concentrations of metals in both the low molecular weight metal-binding protein fraction, which contains metallothioneins, and in the high molecular weight metal-binding protein fraction, which contains metalloenzymes. Our results from analysis of laboratory-exposed populations and from monitoring indigenous and transplanted populations indicate that the capacity for production of metallothioneins is limited and that the quantities present differ greatly with seasonal changes in the environment.

Nomenclature of metallothionein.

The original recommendations concerning the nomenclature of metallothionein were made by the plenum of the First International Meeting on Metallothionein and Other Low Molecular Weight Metal-binding Proteins in 1978 (Nordberg and Kojima, 1979). The present revised version includes amendments to these recommendations as adopted by the Committee on the Nomenclature of Metallothionein appointed at the General Discussion Session of the Second International Meeting on Metallothionein and Other Low Molecular Weight Metal-binding Proteins in 1985.

Isolation of a novel metal-binding protein from rat testes. Characterization and distinction from metallothionein.

This study was undertaken to further establish the nature of the low molecular weight metal-binding proteins in rat testes. In all cases, control testes were compared to livers of zinc-treated rats, which are known to contain high concentrations of metallothionein. Gel filtration of testicular and hepatic cytosol revealed a major zinc- and/or cadmium-binding protein in the low molecular weight range in both tissues. This protein could be partially purified from either source by a combination of heat treatment and sequential acetone precipitation. When such partially purified preparations were further fractionated by high performance liquid chromatography using a linear gradient of 25%-40% acetonitrile in 0.1% trifluoroacetic acid, two major forms with similar retention times were seen in each tissue. The utility of this high performance liquid chromatography system for separating isoforms of metallothionein was verified by separation of commercially available purified rabbit hepatic metallothionein into a total of five separate forms. Amino acid analysis of the two proteins derived from rat liver was consistent with the known amino acid composition of metallothionein. However, the two testicular forms separated by high performance liquid chromatography were notably different in amino acid composition from metallothionein, with a distinctly lower content of cysteine. These results indicate that the major low molecular weight cadmium/zinc-binding proteins in rat testes are not metallothioneins.

Biochemical and ultrastructural observations of long-term silver accumulation in the mussel, Mytilus edulis

The cellular distribution and chemical forms of Ag were determined in mussels after accumulation of the metal from sea water. The major accumulations were either in the vacuoles of connective tissue macrophages, where it was associated with S, or in deposits in the fibrillar layer of the basement membranes of the digestive diverticulum and kidney epithelia, where it was bound to the sulphydryl and sulphate groups of glycoproteins and proteoglycans. In acutely exposed animals, about 10 % of the accumulated Ag was found in a (Ag, Cu)-binding protein with characteristics indicating that it may be a metallothionein. There was a concomitant increase in the body Cu levels after Ag exposure and 40 % of this was bound to this low molecular weight metal-binding protein.

Time course of gold-induced accumulation of copper and zinc and the effects of dimercaptosuccinate and cadmium on gold metabolism in rat kidney

After the s.c. administration of sodium aurothiomalate (SATM, 10 mg Au/kg) to male rats, the gold content of the kidney increased to a maximum after 4 days and thereafter declined slowly. The total copper content of the kidney increased at least until 11 days after SATM treatment but was not simply a function of the renal gold content. Gold and copper accumulated in the metallothionein-like, low molecular weight protein fraction and the initial uptake of gold by this fraction appeared to be related to the accumulation of copper. The Zn 2+ content of the kidney was initially unchanged but later increased with the additional Zn 2+ bound predominantly to the non-soluble components. The accumulation of low molecular weight protein-bound copper was not related either to the Zn 2+ content of this fraction or to the total Zn 2+ content of the kidney. Daily administration of dimercaptosuccinic acid (DMSA, 50 mg/kg, i.p.) for 2 weeks to SATM-pretreated rats resulted in a 50% reduction in the concentration of gold in the non-soluble fraction and in both the high and low molecular weight protein components of the soluble fraction. Copper, which accumulated in the non-soluble components and in the soluble low molecular weight protein, was lost only from the non-soluble components. Uptake of gold by the kidneys was increased when rats were pretreated with Cd 2+. The accumulation of gold in the low molecular weight protein fraction was also increased, but most of the additional gold in the kidneys was accumulated by the non-soluble components. SATM had no effect on the renal concentration or subcellular distribution of Cd 2+. It was concluded that the soluble low molecular weight metal binding protein does not have prominent regulatory or protective functions in the renal metabolism of gold.

Metal accumulation inAgaricus bisporus: Influence of Cd and Cu on growth and tyrosinase activity

To investigate heavy metal uptake in the common mushroomAgaricus bisporus, commercial cultures of the fungus were exposed to different amounts of copper ad cadmium. In contrast to copper, cadmium already exerted pronounced toxic effects at low concentrations (>10 μM) in the compost and concomitantly enhanced the uptake of copper. Tyrosinase activity measured in the fruit bodies increased sharply, when small amounts of Cd (10 μM) were added to copper-rich compost. Gel filtration experiments with crude extracts from fruit bodies demonstrated the absence of a low molecular weight metal binding protein.

重金属とメタロチオネイン

Metallothionein is named for its characteristic structure, a low molecular weight metal-binding protein (61 amino acid residues) rich in cysteinyl residues (20 residues) and heavy metals (7 metals for zinc and cadmium). Heavy metals which can induce and can be bound to metallothionein are restricted to seven heavy metals (zinc, copper, cadmium, silver, mercury, gold, and bismuth). Namely, heavy metals with higher affinity than zinc can be bound in vivo to metallothionein.Other heavy metals such as nickel, manganese, chromium, and so on can also induce metallothionein but can not be bound to metallothionein and the induced metallothionein contains only zinc or zinc and a small amount of copper. A great diversity of stresses other than heavy metals such as starvation, operation, administrations of alkylating agents and inflammatory drugs also induces metallothionein. Glucocorticohormone which may be related to the stresses also induces metallothionein and high concentrations of metallothionein are observed in the livers of foetus and neonatus. Biological functions of metallothionein have been postulated to be a protective protein from harmful heavy metals and a regulating protein for the homeostasis of zinc and copper.

Purification of Low Molecular Weight Metal-Binding Proteins by Preparative Polyacrylamide Gel Electrophoresis: Properties of Electrophoretically Purified Rat Liver (Cd, Zn)-Metallothioneins

A method is proposed for purification of metallothionein by preparative polyacrylamide gel electrophoresis. The method enables purification of 100-700 mg of a preparation containing (Cd, Zn) - metallothionein yielding preparations of considerably higher purity as compared with those obtained by ion-exchange chromatography.

Metallothionein and cadmium, zinc and copper distribution in blood of rats.

Plasma and red blood cells were prepared from rats given daily subcutaneous injection of 0.5 mg Cd/kg body weight for 26 weeks. The plasma and hemolysate of the cells were separated by Sephadex G-75 gel filtration. Main cadmium-con-taining (metallothionein) fractions were collected and subjected to DEAE Sephadex A-25 chromatography.DEAE Sephadex A-25 chromatographic profile of the cadmium-containing frac-tion from the plasma showed four cadmium peaks having corresponding copper peaks, but no zinc. DEAE Sephadex A-25 chromatogram of the sample from the hemolysate showed also four cadmium peaks corresponding to the four peaks from the plasma; each peak had an absorption peak at 250 nm. A small amount of zinc was observed at every peak. The separation profiles of two main cadmium peaks of the anion exchange chromatograms from the plasma and hemolysate were consistent with those of two forms of cadmium-induced hepatic and renal metallothionein.These results indicate that the low molecular-weight metal-binding proteins in the blood include metallothionein as a main component and strongly suggest that the proteins affect the turnover of zinc and copper as well as cadmium in the blood.

Alaskan malamute chondrodysplasia—VII. Isolation and characterization of copper, zinc and iron binding proteins in Canis familiaris

1. 1. Three low molecular weight (12,000, 10,000 and 7,000) metal binding proteins have been isolated from the livers of normal and chondrodysplastic Alaskan Malamutes. 2. 2. Comparison studies between kennel (mixed breed) dogs and both adult and immature Alaskan Malamutes suggested that the disturbance in trace mineral metabolism found in the Malamutes is almost entirely reflected in the 12,000 mol. wt species. 3. 3. The major copper-inducible protein (10,000 mol. wt) observed in kennel dogs was not found to be inducible in Malamutes and contained constant ratios of both copper and zinc to protein in metal binding proteins isolated from the livers of both normal and dwarf Malamutes. 4. 4. The copper and zinc found in the UM2 concentrates ( mol. wt > 2000 ) of immature Malamutes showed very little affinity to the proteins and these metals were found chiefly in a peptide fraction which apparently serves as a reservoir from which the storage proteins obtain the metals that they bind. 5. 5. Regression analysis indicated a statistically significant correlation between both copper and zinc concentrations and the carbohydrate concentration in the proteins investigated.

Isolation of a non-thionein copper-binding protein from liver of copper-injected rats

1. The characterization of a low molecular weight, non-thionein, Cubinding protein isolated from rat liver is reported. The protein was isolated following chronic administration of Cu(NO 3) 2 using a combination of Sephadex G-75 and Sephadex DEAE A-25 chromatography. The protein did not bind to fully equilibrated Sephadex DEAE which formed the basis of the isolation procedure. 2. The final protein pereparation was found to be homogeneous by a variety of electrophoretic techniques and was distinguished from metallothionein on the basis of its behaviour on ion exchange and electrophoretic systems, spectral properties, and amino acid composition and metal content. It contains 6.8% cysteine and was found to bind Cu in a ratio of 1.5:1 based on a molecular weight of 11 000. 3. These results confirm the necessity to use techniques other than gel filtration alone to obtain adequate separation of low molecular weight metal-binding protein fractions.

607. Problems in assessing exposure to mercury: Berlin, M. (1963). On estimating threshold limits for mercury in biological material. Acta med. scand. Suppl. 396

607. Problems in assessing exposure to mercury: Berlin, M. (1963). On estimating threshold limits for mercury in biological material. Acta med. scand. Suppl. 396