Selenium Protein Research Articles

Selenium in Proteins: Conformational Changes Induced by Se Substitution on Methionine, as Studied in Isolated Model Peptides by Optical Spectroscopy and Quantum Chemistry.

The side-chain of methionine residues is long enough to establish NH⋯S H-bonds with neighboring carbonyl groups of the backbone, giving rise to so-called intra-residue 6δ and inter-residue 7δ H-bonds. The aim of the present article is to document how the substitution of sulfur with a selenium atom affects the H-bonding of the Met system. This was investigated both experimentally and theoretically by conformation-resolved optical spectroscopy, following an isolated molecule approach. The present work emphasizes the similarities of the Met and Sem residues in terms of conformational structures, energetics, NH⋯Se/S H-bond strength and NH stretch spectral shifts, but also reveals subtle behavior differences between them. It provides evidence for the sensitivity of the H-bonding network with the folding type of the Sem/Met side-chains, where a simple flip of the terminal part of the side-chain can induce an extra 50 cm−1 spectral shift of the NH stretch engaged in a 7δ NH⋯S/Se bond.

Stuck between a ROS and a hard place: Analysis of the ubiquitin proteasome pathway in selenocysteine treated Brassica napus reveals different toxicities during selenium assimilation

During the selenium assimilation pathway, inorganic selenate and selenite are reduced to form selenocysteine (Sec). Tolerance to selenium in plants has long been attributable to minimizing the replacement of cysteine with selenocysteine, which can result in nonspecific selenoproteins that are potentially misfolded. Despite this widely accepted assumption, there is no evidence in higher plants demonstrating that selenocysteine induces toxicity by resulting in malformed proteins. In this study, we use Brassica napus to analyze the ubiquitin-proteasome pathway, which is capable of removing misfolded proteins. Sec rapidly increased proteasome activity and levels of ubiquitinated proteins, strongly indicating that selenocysteine induces protein misfolding. Proteasome inhibition increased the amount of selenium in protein in Sec-treated plants. Collectively, these data provide a mechanism that accounts for Sec toxicity. Additionally, Sec did not cause oxidative stress as judged by examining levels of superoxide using fluorescent microscopy. Therefore, the cellular response to Sec is different compared to selenite, which was recently shown to increase antioxidant metabolism in response to elevated mitochondrial superoxide that ultimately impaired proteasome activity. Therefore, plants must contend with two divergent modes of cytotoxicity during selenium assimilation. Selenite can result in oxidative stress, but increased flux of selenite reduction can yield Sec that in turn can cause protein misfolding.

HPLC-ICP-MS analysis of selenium speciation in selenium-enriched Cordyceps militaris

This study examined the distribution of selenium in proteins from selenium-enriched Cordyceps militaris extract using reversed-phase high performance liquid chromatography (RP-HPLC) and size-exclusion chromatography (SEC) coupled to on-line ICP-MS specific for selenium detection. An analytical method was developed for detecting the presence of selenocysteine, selenomethionine and selenopeptides. The results showed that soluble selenium was the major form of selenium in selenium-enriched C. militaris extract. About 84.4% of the selenium was present as low-molecular-weight molecules (MW < 4 kDa), whereas 7.13% of the selenium was present in the form of high-molecular-weight molecules, probably as selenoproteins. Furthermore, analysis of acid-hydrolyzed C. militaris extract revealed a higher proportion of Se-Cys2 than Se-Met (two main selenium-conjugated amino acids) in the extract, although various forms of smaller selenium-proteins or selenium-peptides were also present in low but detectable amounts. The present analysis of selenium thus combined different chromatographic techniques for efficient and rapid separation of selenoproteins and other seleno-conjugated molecules.

Putting Selenium in Proteins

Putting Selenium in Proteins

ChemInform Abstract: Selenium Derivatization of Nucleic Acids for X‐Ray Crystal‐Structure and Function Studies

AbstractReview: 105 refs.

The Analysis of Selenoprotein and Se-Polysaccharide in Selenium-Enriched &lt;i&gt;Lentinan edodes&lt;/i&gt;

The cultivation of Lentinan edodes as a carrier for Se, low concentrations of sodium selenite solution were added directly to the medium to inorganic selenium be absorbed into the mushroom body, and selenium-enriched Lentinan edodes can be got. Using selenium-enriched mushrooms as raw material, it was preliminarily studied that patterns of occurrence of selenium and the contents of Selenoprotein and Se-polysaccharide in their fruiting bodies of mushrooms. Various extraction solvents were used to distill and separation Se-polysaccharide and Se-protein of the Lentinan edodes, and atomic absorption spectrometer was used to determine the selenium contents of all component parts. The results showed that Lentinan edodes have the ability of enrichment selenium and can transform inorganic selenium in the environment into organic selenium. The main group of Selenium is organic state. The selenium in protein occupied the total selenium 40.09%, In the four components of selenium protein, water-soluble proteins of selenium is main (21.65% of the total selenium).The selenium in polysaccharide occupied the total selenium 27.83%. Mass fraction of acid-soluble Se-polysaccharide than in water-soluble Se-polysaccharide and alkali-soluble Se-polysaccharide is higher (13.52% of total selenium). The main distributed form of the selenium is from the organic matter in the analyzed Lentinan edodes, the combined selenium in protein was higher.

Adenosine 5′-phosphosulfate reductase (APR2) mutation inArabidopsisimplicates glutathione deficiency in selenate toxicity

APR2 is the dominant APR (adenosine 5'-phosphosulfate reductase) in the model plant Arabidopsis thaliana, and converts activated sulfate to sulfite, a key reaction in the sulfate reduction pathway. To determine whether APR2 has a role in selenium tolerance and metabolism, a mutant Arabidopsis line (apr2-1) was studied. apr2-1 plants had decreased selenate tolerance and photosynthetic efficiency. Sulfur metabolism was perturbed in apr2-1 plants grown on selenate, as observed by an increase in total sulfur and sulfate, and a 2-fold decrease in glutathione concentration. The altered sulfur metabolism in apr2-1 grown on selenate did not reflect typical sulfate starvation, as cysteine and methionine levels were increased. Knockout of APR2 also increased the accumulation of total selenium and selenate. However, the accumulation of selenite and selenium incorporation in protein was lower in apr2-1 mutants. Decreased incorporation of selenium in protein is typically associated with increased selenium tolerance in plants. However, because the apr2-1 mutant exhibited decreased tolerance to selenate, we propose that selenium toxicity can also be caused by selenate's disruption of glutathione biosynthesis leading to enhanced levels of damaging ROS (reactive oxygen species).

Selenium speciation in different organs of African catfish (Clarias gariepinus) enriched through a selenium-enriched garlic based diet

Speciation of Se in fish is needed to elucidate the metabolism of this element in living organisms in the marine environment. In this paper, selenium concentration and its species distribution in several organs and tissues (liver, gills, kidney, muscle and gastrointestinal tract) of African catfish fed with a selenium-enriched garlic based diet was studied. The intention of this paper is focused on both the investigation of selenium distribution in the soluble protein fraction and the detection of selenoaminoacids. Thus, two different procedures have been developed. In the first procedure, screening of selenium in proteins in the Tris-buffer soluble fraction of different tissues was carried out by size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) and by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) after sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) separation and electroblotting onto membranes. For the amino acid analysis, several sample treatments for Se-species extraction, based on enzymatic hydrolysis, were compared. The best results were obtained for incubation at controlled temperature. Application of several sample treatments in conjunction with different chromatographic techniques (reverse phase, anion exchange and ion exchange/size exclusion) was crucial to unambiguous Se-species identification. In Se-enriched African catfish a noticeable increase in the content of selenium in different organs was observed, except for the liver, where the Se level remained unaltered. The kidney was the Se-target organ in animals fed with enriched Se food. Selenomethionine (SeMet) was the main Se species identified in fillet extracts, whereas the presence of selenocysteine (SeCys) was detected in the liver and both SeMet and SeCys were present in the kidney.

Screening of selenium containing proteins in the Tris-buffer soluble fraction of African catfish (Clarias gariepinus) fillets by laser ablation-ICP-MS after SDS-PAGE and electroblotting onto membranes

The anti-carcinogenic properties of selenium against certain types of cancer when present in organic forms justify the increasing interest in development of selenium fortified food. In this particular study, African catfish (Clarias gariepinus) were fed with a Se-enriched diet in order to enhance the selenium concentration in the fish fillet up to 0.85 ± 0.06 mg Se kg−1 (0.26 ± 0.02 mg Se kg−1 in control). Selenium distribution in proteins separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was studied for better understanding of the metabolism of this element in the organism. Soluble proteins (Tris buffer) were investigated by conventional application of a glass homogenizer in comparison to application of ultrasound probe sonication, and the latter was convincing due to higher protein extraction efficiency and the ease and speed of application. Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) was successfully applied for screening of selenium in proteins in fillets of African catfish separated by one-dimensional SDS-PAGE and after electroblotting onto nitrocellulose membranes, with a total Se amount below 0.4 ng loaded into a well. Selenium was detected in more than 11 protein spots. In order to improve the protein separation of one-dimensional SDS-PAGE, pre-fractionation of the Tris buffer soluble protein extract by size exclusion was carried out. Tryptic digestion of the Se-containing bands was performed for protein identification by nano-HPLC coupled to electrospray-MS/MS. A database search revealed several proteins which are located in muscle tissue. However, the very low Se concentration circumvents the detection of selenium containing amino acids in the tryptic peptides by electrospray-MS/MS.

Influence of selenised dairy proteins on biomarkers of colon cancer risk

AbstractSelenium is an essential trace element for mammals, and a component of at least 25 selenoproteins which incorporate the amino acid selenocysteine (Sec). These proteins include a number with oxidoreductase functions. An examination of the selenoproteins and their influence on carcinogenesis in an animal model may assist in determining their relevance in chemoprevention.Food sources offer a number of organic forms of selenium, with selenomethionine a common component, as in selenised yeasts. A selenium‐rich dairy protein product has been developed (TaturaBioSe, Tatura Milk Industries, Tatura, Victoria) which could improve selenium status in populations considered marginal or deficient. It could also provide higher intakes (e.g. several fold above recommended dietary intake recommendations) of potential benefit as chemopreventive to those at greater risk of some selenium‐responsive diseases, such as some sporadic colorectal cancers. Clinical studies are showing it to be a safe, palatable product for consumption in the form provided.Dairy protein selenium at 1 ppm in diet had a significant chemopreventive effect compared with control (0.05 ppm Se) in an azoxymethane model of colon cancer. Colon tumour incidence in rats was down by 29%, and tumour burden (colon tumours/rat) was halved, effects not observed when an equivalent Se concentration was provided as yeast selenium. When assessed by plasma Se concentration, this dairy protein form of selenium showed greater bioavailability (as assessed by plasma selenium) as well as efficacy in chemoprevention. Programmed cell death (apoptosis) was increased in colonic crypts and crypt height significantly diminished. The influence on early changes in carcinogenesis provides an insight into possible mechanism(s) of action. Histological and biochemical assays (e.g. monitoring oxidoreductase enzymes) could potentially provide early biomarkers with clinical relevance.

Expression of a mouse selenocysteine lyase in Brassica juncea chloroplasts affects selenium tolerance and accumulation

Selenium is an essential nutrient for many organisms, as part of certain selenoproteins. However, selenium is toxic at high levels, which is thought to be due to non‐specific replacement of cysteine by selenocysteine leading to disruption of protein function. In an attempt to prevent non‐specific incorporation of selenocysteine into proteins and to possibly enhance plant selenium tolerance and accumulation, a mouse selenocysteine lyase was expressed in Brassica juncea (Indian mustard) chloroplasts, the site of selenocysteine synthesis. This selenocysteine lyase specifically breaks down selenocysteine into elemental selenium and alanine. The transgenic cpSL plants showed normal growth under standard conditions. Selenocysteine lyase activity in the cpSL transgenics was up to 6‐fold higher than in wild‐type plants. The cpSL transgenics contained up to 40% less selenium in protein compared to wild‐type plants, indicating that Se flow in the plant was successfully redirected. Surprisingly, the selenium tolerance of the transgenic cpSL plants was reduced, perhaps due to interference of produced elemental selenium with chloroplastic sulphur metabolism. Shoot selenium levels were enhanced up to 50% in the cpSL transgenics, but only during the seedling stage.

Detection and quantification of selenium in proteins by means of gel electrophoresis and electrothermal vaporization ICP-MS

A method for the separation of selenium-containing proteins and subsequent detection and quantification of selenium was developed. First, the proteins are fractionated by means of sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE); subsequently, the bands of the gel, containing the proteins, are analyzed by electrothermal vaporization (ETV)-ICP-MS to detect and quantify selenium. External standardization with the use of an internal reference (Te) was applied. A detection limit of ca. 40 pg Se per band and a recovery of ca. 98% were obtained. A single measurement is accomplished in less than 4 min and thus a gel lane after a separation of ca. 1.5 h, can be entirely analyzed with ETV-ICP-MS in 3.5 h. If limited to 10 bands, a gel lane is analyzed in 2 h (calibration included). The analysis is directly carried out on the stained gel, without blotting, which makes the analysis even more practical. This method was optimized using the selenoprotein glutathione peroxidase as model. Then, it was applied to the fractionation of proteins from a selenium–yeast candidate reference material. The reconstructed Se electropherograms are presented and compared with the stained gels. The major advantages of this method are the high resolution of the protein fractionation and the straightforward quantification of selenium.

Interaction of peroxynitrite with selenoproteins and glutathione peroxidase mimics

Peroxynitrite is an oxidant generated under inflammatory conditions, acting in defense against invading microorganisms. There is a need for protection of the organism from damage inflicted by peroxynitrite. Selenium-containing compounds, notably ebselen, have a high second-order reaction rate constant (approx. 2 × 10 6 M −1 s −1), which makes them candidates for efficient protection. This applies also for selenium in proteins, occurring as selenocysteine or selenomethionine residues. Glutathione peroxidases, thioredoxin reductase, and selenoprotein P have been shown to play a potential role in protection against peroxynitrite. Tellurium-containing compounds also react with peroxynitrite.

Different selenium-containing proteins in the extracellular and intracellular media of leucocytes cultivated in vitro.

The purpose of this communication is to elucidate if selenium plays a role in the function of granulocytes and lymphocytes. Thus, the incorporation of selenium in proteins from granulocytes and lymphocytes cultured with 1 microCi/mL radioactive Na2(75)SeO3 was studied. The protein peaks containing 75Se from two columns of Heparin Sepharose CL-6B and Sephacryl S-200 HR were separated further by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The results showed that the incorporation of 75Se into granulocytes was about six times higher than that of lymphocytes during a 96-h cultivation, however, the GSH-Px activity in granulocytes did not change significantly. On the other hand, the GSH-Px activity of lymphocytes rose significantly after three days cultivation. These data indicated that the main chemical form of selenium in granulocytes was not GSH-Px. Results from SDS-PAGE revealed a strongly 75Se-labeled protein band with subunit molecular weight of 15 kDa in the supernatant of granulocyte homogenate. However, the main chemical forms of selenium in the culture media of granulocytes and lymphocytes were found to be selenoprotein P. The different forms of selenium-containing proteins in the intracellular and extracellular media of granulocytes indicated the different functions of these proteins.

Biological utilization of some selenium- and sulfur-containing amino acids.

It is now generally recognized that selenium is an essential micronutrient for mammals, birds and several bacteria (1). Covalen-tly bound selenium is naturally present in some proteins; it is an essential constituent of at least four enzymes, the well known mammalian glutathione peroxidase (2,3,4,8) and the bacterial formate dehydrogenase (5), glycine reductase (6) and nicotinic acid hydroxylase (7). In the first three enzymes selenium is present in a polypeptide chain as a selenocysteine residue (6,8,9). Besides these examples, in which the catalytic activity is strictly dependent on the presence of selenium, the unspecific occurrence of selenium in proteins is also well documented (1). For example E.coli can utilize selenate in substitution of sulfate, and the majority of selenium is recovered in cellular proteins as selenomethionine (10); selenomethionine containing proteins are active enough to support cell growth (11). Selenomethionine can substitute methionine as growth factor for a methionine requiring E.coli mutant, and extensive replacement of protein methionine by selenomethionine occurs (12). Thus some proteins may contain selenium in substitution of sulfur without great impairment of their biological function. A typical example is the β-galactosidase of E.coli.KeywordsFormate DehydrogenaseSelenium AtomProtein LysineBiological UtilizationProline AnalogThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Selenium biochemistry.

The toxicity of selenium to animals and plants has been known and extensively documented since the 1930's, but it is only during the past 15 years that selenium has also been shown to be an essential micronutrient for animals and bacteria. Very little is known about the specific role or roles of selenium and, to date, there are only three enzyme-catalyzed reactions that have been shown to require the participation of a selenium-containing protein. These are the reactions catalyzed by (i) formate dehydrogenase of bacteria, (ii) glycine reductase of clostridia, and (iii) glutathione peroxidase of erythrocytes. The common denominator of these selenium-dependent processes is that they are all oxidation-reduction reactions. A fourth selenoprotein has been isolated from skeletal muscle of sheep but its catalytic function has not been identified. The form in which selenium occurs in these selenoproteins is unknown. The selenoprotein of clostridial glycine reductase contains selenium in a covalently bound form. Studies in progress indicate that this may be an organoselenium compound not previously detected in nature. Identification of the chemical nature of selenium in proteins participating in electron transport processes should enable us to determine its specific role and to understand the basic defects in certain cardiac and skeletal muscle degenerative diseases which are selenium-deficiency syndromes. The greater availability and ease of isolation of the selenoprotein of the bacterial glycine reductase system makes this the biological material of choice for studies on the mechanism of action of selenium. An added attractive feature of this system is that it can conserve the energy made available by the reductive deamination of glycine in a biologically useful form by synthesizing ATP.

Selenium in Proteins from Toxic Foodstuffs: IV. The Effect of Feeding Toxic Proteins, Toxic Protein Hydrolysates, and Toxic Protein Hydrolysates from which the Selenium Has Been Removed: Two Figures

Selenium in Proteins from Toxic Foodstuffs: IV. The Effect of Feeding Toxic Proteins, Toxic Protein Hydrolysates, and Toxic Protein Hydrolysates from which the Selenium Has Been Removed: Two Figures

SELENIUM IN PROTEINS FROM TOXIC FOODSTUFFS: III. THE REMOVAL OF SELENIUM FROM TOXIC PROTEIN HYDROLYSATES

SELENIUM IN PROTEINS FROM TOXIC FOODSTUFFS: III. THE REMOVAL OF SELENIUM FROM TOXIC PROTEIN HYDROLYSATES