ABSTRACT
DNA synthesis catalyzed by DNA polymerase is essential for all life forms, and phosphodiester bond formation with phosphorus center inversion is a key step in this process. Herein, by using a single-selenium-atom-modified dNTP probe, we report a novel strategy to visualize the reaction stereochemistry and catalysis. We capture the before- and after-reaction states and provide explicit evidence of the center inversion and in-line attacking SN2 mechanism of DNA polymerization, while solving the diastereomer absolute configurations. Further, our kinetic and thermodynamic studies demonstrate that in the presence of Mg2+ ions (or Mn2+), the binding affinity (Km) and reaction selectivity (kcat/Km) of dGTPαSe-Rp were 51.1-fold (or 19.5-fold) stronger and 21.8-fold (or 11.3-fold) higher than those of dGTPαSe-Sp, respectively, indicating that the diastereomeric Se-Sp atom was quite disruptive of the binding and catalysis. Our findings reveal that the third metal ion is much more critical than the other two metal ions in both substrate recognition and bond formation, providing insights into how to better design the polymerase inhibitors and discover the therapeutics.
Subject(s)
Selenium , DNA-Directed DNA Polymerase/metabolism , Metals/pharmacology , Catalysis , DNA , Ions , KineticsABSTRACT
This paper consists of the evaluation in regards to the ecotoxicological effectiveness of a treatment applied to a coal mining waste. The treatment consisted of separating the particles based on gravimetric concentration in spirals, generating three fractions: heavy, intermediate, and light, with high, moderate, and low pyrite content, respectively. The intermediate fraction represents the larger disposal volume of the waste on soils. To evaluate the effectiveness of the treatment, metal determination and bioassays Eisenia andrei, Folsomia candida, Lactuca sativa, Daphnia similis, and Raphidocelis subcapitata were applied to the intermediary fraction. To evaluate the toxicity to aquatic organisms, elutriates were generated from the unprocessed waste and the intermediate fraction. The intermediate fraction showed a decrease of metal concentrations compared to the untreated waste. Metal concentrations in the intermediate fraction were below the Brazilian thresholds for soil quality. Avoidance bioassay with E. andrei and germination tests of L. sativa showed no significant effects. The bioassay with F. candida indicated a significant reduction in reproduction at the highest doses used (24% and 50%). Bioassays with D. similis and R. subcapitata revealed a reduction in toxicity of the intermediate fraction compared to the untreated waste. However, the toxicity levels of the intermediate fraction to aquatic organisms still require attention, especially in regards to pH that played a crucial role in the toxicity. Finally, the results suggest that the treatment performed on the coal waste was efficient, even though significant toxicity have still been detected in the treated waste and additional steps are still required for adequate final disposal.
Subject(s)
Arthropods , Coal Mining , Soil Pollutants , Animals , Aliivibrio fischeri , Soil , Metals/pharmacology , Soil Pollutants/analysis , MiningABSTRACT
The aim of the present review is to discuss traditional hypotheses on the etiopathogenesis of Alzheimer's disease (AD), as well as the role of metabolic-syndrome-related mechanisms in AD development with a special focus on advanced glycation end-products (AGEs) and their role in metal-induced neurodegeneration in AD. Persistent hyperglycemia along with oxidative stress results in increased protein glycation and formation of AGEs. The latter were shown to possess a wide spectrum of neurotoxic effects including increased Aß generation and aggregation. In addition, AGE binding to receptor for AGE (RAGE) induces a variety of pathways contributing to neuroinflammation. The existing data also demonstrate that AGE toxicity seems to mediate the involvement of copper (Cu) and potentially other metals in AD pathogenesis. Specifically, Cu promotes AGE formation, AGE-Aß cross-linking and up-regulation of RAGE expression. Moreover, Aß glycation was shown to increase prooxidant effects of Cu through Fenton chemistry. Given the role of AGE and RAGE, as well as metal toxicity in AD pathogenesis, it is proposed that metal chelation and/or incretins may slow down oxidative damage. In addition, selenium (Se) compounds seem to attenuate the intracellular toxicity of the deranged tau and Aß, as well as inhibiting AGE accumulation and metal-induced neurotoxicity.
Subject(s)
Alzheimer Disease/metabolism , Alzheimer Disease/physiopathology , Glycation End Products, Advanced/metabolism , Amyloid beta-Peptides/metabolism , Chelating Agents/pharmacology , Copper/metabolism , Glycemic Index/physiology , Humans , Iron/metabolism , Lipid Metabolism/physiology , Metabolic Syndrome/physiopathology , Metals/pharmacology , Neurodegenerative Diseases/etiology , Neurodegenerative Diseases/metabolism , Oxidative Stress/drug effects , Reactive Oxygen Species/pharmacology , Selenium/metabolismABSTRACT
The relative concentration of available inorganic elements is critical for yeast growth and metabolism and has potential to be a tool leading to directed yeast flavour formation during fermentation. This study investigates the influence of essential inorganic elements during alcoholic fermentation of brewers wort, fermented using three independent yeast strains, Saccharomyces pastorianus W34/70, and Saccharomyces cerevisiae strains M2 and NCYC2592 under a range of conditions replicated for each yeast strain. 10 treatments were applied: 1 control and 9 inorganic supplementations: standard brewers wort, ammonia-nitrogen, inorganic phosphate, potassium, magnesium, copper, zinc, iron, manganese and a composite mixture, Twenty-five chemical markers were evaluated by HPLC (ethanol, glycerol), and GC-MS (aroma). There was a significant change in volatile aroma compounds during fermentation, which was more prominent when supplemented with ammonia nitrogen, inorganic phosphate, potassium or magnesium (P < 0.05). Heavy metal ions mostly had a negative effect on the flavour formation.
Subject(s)
Beer/microbiology , Metals/pharmacology , Saccharomyces/metabolism , Beer/analysis , Chromatography, High Pressure Liquid , Ethanol/metabolism , Fermentation/drug effects , Food Microbiology , Gas Chromatography-Mass Spectrometry , Glycerol/metabolism , Metals/metabolism , Odorants , Potassium/metabolism , Potassium/pharmacology , Saccharomyces/drug effects , Saccharomyces cerevisiae/drug effects , Saccharomyces cerevisiae/metabolismABSTRACT
A fibrinolytic enzyme with thrombolytic, anticoagulant activities was purified from fruiting bodies of wild-growing mushroom Boletus pseudocalopus Hongo and homogenized with a two-step procedure with a 6.11-fold increase in specific activity and 3.2% recovery. The molecular weight of the enzyme was estimated to be 63.5 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. The enzyme was active at 40°C and pH 7, and activity was inhibited by zinc metal ion and by serine protease and trypsin inhibitors phenylmethylsulfonyl fluoride and N-α-tosyl-l-lysinyl-chloromethylketone. The enzyme displayed high specificity for Pyro-Glu-Gly-Arg-pNA. In vitro assays showed that the enzyme was able to degrade fibrin and blood clots, inhibit thrombin and activated factor X, and alter the density or structural change of fibrin clots. It could also delay activated partial thromboplastin time and prothrombin time. These results suggest that the enzyme may have characteristics of a trypsin or serine-like enzyme with fibrinolytic and thrombolytic activities and may have potential as an antithrombotic agent for blood clotting disorders.
Subject(s)
Basidiomycota/enzymology , Fibrinolytic Agents/isolation & purification , Fruiting Bodies, Fungal/enzymology , Blood Coagulation Tests , Fibrin Clot Lysis Time , Fibrinolysis/drug effects , Fibrinolytic Agents/pharmacology , Hydrogen-Ion Concentration , Metals/chemistry , Metals/pharmacology , Microscopy, Fluorescence , Molecular Weight , Nephelometry and Turbidimetry , Protease Inhibitors/pharmacology , Republic of Korea , TemperatureABSTRACT
Nanotechnology is an emerging science with a wide array of applications involving the synthesis and manipulation of materials with dimensions in the range of 1-100 nm. Nanotechnological applications include diverse fields such as pharmaceuticals, medicine, the environment, food processing and agriculture. Regarding the latter, applications are mainly focused on plant growth and crop protection against plagues and diseases. In recent years, the biogenic reduction of elements such as Ag, Au, Cu, Cd, Al, Se, Zn, Ce, Ti and Fe with plant extracts has become one of the most accepted techniques for obtaining nanoparticles (NPs), as it is considered an ecological and cost-effective process without the use of chemical contaminants. The objective of this work was to review NPs synthesized by green chemistry using vegetable extracts, as well as their use as antimicrobial agents against phytopathogenic fungi and bacteria. Given the need for alternatives to control and integrate management of phytopathogens, this review is relevant to agriculture, although this technology is barely exploited in this field. © 2020 Society of Chemical Industry.
Subject(s)
Anti-Infective Agents/chemical synthesis , Anti-Infective Agents/pharmacology , Metal Nanoparticles/chemistry , Metal Nanoparticles/toxicity , Plant Extracts/chemistry , Anti-Infective Agents/chemistry , Bacteria/drug effects , Bacteria/growth & development , Fungi/drug effects , Fungi/growth & development , Metals/chemistry , Metals/pharmacologyABSTRACT
Terrestrial gastropods express metal-selective metallothioneins (MTs) by which they handle metal ions such as Zn2+ , Cd2+ , and Cu+ /Cu2+ through separate metabolic pathways. At the same time, they depend on the availability of sufficient amounts of Cu as an essential constituent of their respiratory protein, hemocyanin (Hc). It was, therefore, suggested that in snails Cu-dependent MT and Hc pathways might be metabolically connected. In fact, the Cu-specific snail MT (CuMT) is exclusively expressed in rhogocytes, a particular molluscan cell type present in the hemocoel and connective tissues. Snail rhogocytes are also the sites of Hc synthesis. In the present study, possible interactions between the metal-regulatory and detoxifying activity of MTs and the Cu demand of Hc isoforms was explored in the edible snail Cornu aspersum, one of the most common European helicid land snails. This species possesses CdMT and CuMT isoforms involved in metal-selective physiological tasks. In addition, C. aspersum expresses three different Hc isoforms (CaH ÉD, CaH ÉN, CaH ß). We have examined the effect of Cd2+ and Cu2+ exposure on metal accumulation in the midgut gland and mantle of C. aspersum, testing the impact of these metals on transcriptional upregulation of CdMT, CuMT, and the three Hc genes in the two organs. We found that the CuMT and CaH ÉD genes exhibit an organ-specific transcriptional upregulation in the midgut gland of Cu-exposed snails. These results are discussed in view of possible interrelationships between the metal-selective activity of snail MT isoforms and the synthesis and metabolism of Hc isoforms.
Subject(s)
Cadmium/pharmacology , Copper/pharmacology , Hemocyanins/metabolism , Snails/drug effects , Animals , Base Sequence , Cadmium/metabolism , Copper/metabolism , DNA, Complementary , Gene Expression Regulation/drug effects , Hemocyanins/genetics , Metallothionein , Metals/metabolism , Metals/pharmacology , Snails/metabolismABSTRACT
Bioactive metal complexes represent a promising and rapidly evolving area of pharmacotherapy. After the first part of our survey on metallopharmaceuticals dealing with antimicrobial activity of metal complexes and their application in diagnostics and the second part dedicated to anticancer properties of these compounds, this third and last part of the review focuses on several other applications of metals in therapy (mainly on the therapy of rheumatoid arthritis, some mental diseases, diabetes, as well as on chelation therapy). Following a brief account of the historical development of clinical use of the respective category of drugs, their chemical properties, toxicity, clinical applications and mechanism of action are discussed. The aim of this brief survey is to provide basic outline of the area of metallopharmacy, aimed at specialists in pharmacy and chemistry as well as at the general educated public.
Subject(s)
Coordination Complexes/pharmacology , Medicine/trends , Pharmacy/trends , Metals/pharmacologyABSTRACT
BACKGROUND: Methylxanthines, including caffeine, theobromine and theophylline, are natural and synthetic compounds in tea, which could be metabolized by certain kinds of bacteria and fungi. Previous studies confirmed that several microbial isolates from Pu-erh tea could degrade and convert caffeine and theophylline. We speculated that these candidate isolates also could degrade and convert theobromine through N-demethylation and oxidation. In this study, seven tea-derived fungal strains were inoculated into various theobromine agar medias and theobromine liquid mediums to assess their capacity in theobromine utilization. Related metabolites with theobromine degradation were detected by using HPLC in the liquid culture to investigate their potential application in the production of 3-methylxanthine. RESULTS: Based on theobromine utilization capacity, Aspergillus niger PT-1, Aspergillus sydowii PT-2, Aspergillus ustus PT-6 and Aspergillus tamarii PT-7 have demonstrated the potential for theobromine biodegradation. Particularly, A. sydowii PT-2 and A. tamarii PT-7 could degrade theobromine significantly (p < 0.05) in all given liquid mediums. 3,7-Dimethyluric acid, 3-methylxanthine, 7-methylxanthine, 3-methyluric acid, xanthine, and uric acid were detected in A. sydowii PT-2 and A. tamarii PT-7 culture, respectively, which confirmed the existence of N-demethylation and oxidation in theobromine catabolism. 3-Methylxanthine was common and main demethylated metabolite of theobromine in the liquid culture. 3-Methylxanthine in A. sydowii PT-2 culture showed a linear relation with initial theobromine concentrations that 177.12 ± 14.06 mg/L 3-methylxanthine was accumulated in TLM-S with 300 mg/L theobromine. Additionally, pH at 5 and metal ion of Fe2+ promoted 3-methylxanthine production significantly (p < 0.05). CONCLUSIONS: This study is the first to confirm that A. sydowii PT-2 and A. tamarii PT-7 degrade theobromine through N-demethylation and oxidation, respectively. A. sydowii PT-2 showed the potential application in 3-methylxanthine production with theobromine as feedstock through the N-demethylation at N-7 position.
Subject(s)
Aspergillus/metabolism , Theobromine/metabolism , Xanthines/metabolism , Aspergillus/drug effects , Biotransformation , Culture Media/chemistry , Culture Media/metabolism , Hydrogen-Ion Concentration , Metals/pharmacology , Methylation , Mycology/methods , Oxidation-Reduction , Teas, Herbal/microbiologyABSTRACT
Classification of chemical compounds of plants as a source of medicaments for neurodegenerative diseases through computer screening is an efficient process in drug discovery, in advance of laboratory testing and clinical trials. The onset of neurodegenerative disorders incarcerates both sufferers and their families mentally and financially. This investigation emphasises the search for potent compounds via a computational approach, as an initial path towards the treatment of the neurodegenerative diseases Alzheimer's (AD), Parkinson's (PD), prion, and Huntington's (HD) diseases. The therapeutic strategy considered here is chelation therapy, emanated from the heightened levels of metal ions, which play an imperative role in the pathogenesis of all four neurodegenerative disorders mentioned. Hence, potent compounds from Sri Lankan plants to function as lead compounds have been identified for Cu(II), Fe(III), Zn(II), and Al(III) ions, from a library of around 200 chemical compounds, using an umbrella sampling molecular dynamics computational approach where the chelating ability of compounds for the metal ion is assessed in terms of binding free energy. Calculations reveal that 12 Sri Lankan plants possess compounds that could be considered as starting points of leads for AD, PD and prion disease. However, no compound was potentially useful for the HD category, according to the study. Potential of mean force of Al3+ binding to (-)-5-methylmellin found in Semecarpus walkeri with two representative configurations.
Subject(s)
Chelating Agents/pharmacology , Chelation Therapy/methods , Computer Simulation , Metals/pharmacology , Neurodegenerative Diseases/drug therapy , Plant Extracts/pharmacology , Plants/chemistry , Chelating Agents/chemistry , Humans , Metals/chemistryABSTRACT
Aureobasidium pullulans is a ubiquitous and widely distributed fungus in the environment, and exhibits substantial tolerance against toxic metals. However, the interactions between metals and metalloids with the copious extracellular polymeric substances (EPS) produced by A. pullulans and possible relationships to tolerance are not well understood. In this study, it was found that mercury (Hg) and selenium (Se), as selenite, not only significantly inhibited growth of A. pullulans but also affected the composition of produced EPS. Lead (Pb) showed little influence on EPS yield or composition. The interactions of EPS from A. pullulans with the tested metals and metalloids depended on the specific element and their concentration. Fluorescence intensity measurements of the EPS showed that the presence of metal(loid)s stimulated the production of extracellular tryptophan-like and aromatic protein-like substances. Examination of fluorescence quenching and calculation of binding constants revealed that the fluorescence quenching process for Hg; arsenic (As), as arsenite; and Pb to EPS were mainly governed by static quenching which resulted in the formation of a stable non-fluorescent complexes between the EPS and metal(loid)s. Se showed no significant interaction with the EPS according to fluorescence quenching. These results provide further understanding of the interactions between metals and metalloids and EPS produced by fungi and their contribution to metal(loid) tolerance. KEY POINTS: ⢠Metal(loid)s enhanced production of tryptophan- and aromatic protein-like substances. ⢠Non-fluorescent complexes formed between the EPS and tested metal(loid)s. ⢠EPS complexation and binding of metal(loid)s was dependent on the tested element. ⢠Metal(loid)-induced changes in EPS composition contributed to metal(loid) tolerance.
Subject(s)
Aureobasidium/drug effects , Fluorescence , Metalloids/pharmacology , Metals/pharmacology , Aureobasidium/growth & development , Extracellular Polymeric Substance Matrix/chemistry , Mercury/pharmacology , Selenium/pharmacologyABSTRACT
Trace metals such as iron and zinc are vital nutrients known to play key roles in prokaryotic processes including gene regulation, catalysis, and protein structure. Metal sequestration by hosts often leads to metal limitation for the bacterium. This limitation induces bacterial gene expression whose protein products allow bacteria to overcome their metal-limited environment. Characterization of such genes is challenging. Bacteria must be grown in meticulously prepared media that allows sufficient access to nutritional metals to permit bacterial growth while maintaining a metal profile conducive to achieving expression of the aforementioned genes. As such, a delicate balance must be established for the concentrations of these metals. Growing a nutritionally fastidious organism such as Neisseria gonorrhoeae, which has evolved to survive only in the human host, adds an additional level of complexity. Here, we describe the preparation of a defined metal-limited medium sufficient to allow gonococcal growth and the desired gene expression. This method allows the investigator to chelate iron and zinc from undesired sources while supplementing the media with defined sources of iron or zinc, whose preparation is also described. Finally, we outline three experiments that utilize this media to help characterize the protein products of metal-regulated gonococcal genes.
Subject(s)
Gene Expression Regulation, Bacterial , Metals/metabolism , Metals/pharmacology , Neisseria gonorrhoeae/growth & development , Bacterial Proteins/genetics , Humans , Iron/metabolism , Ligands , Neisseria gonorrhoeae/genetics , Neisseria gonorrhoeae/metabolism , Zinc/metabolismABSTRACT
Cichoric acid (CA) belongs to the group of polyphenols, which occurs in a variety of plant species and it is characterized by anticancer, antibacterial, and antiviral properties. Selected polyphenols have the ability to combine with metal ions to form chelate complexes that reveal greater biological activity than free compounds. In order to study possible antimicrobial and anticancer effect of CA and its complexes with copper(II)/zinc(II)/nickel(II)/cobalt(II) we decided to conduct cytotoxicity tests to estimate the most effective concentrations of tested compounds. The results of the presented study demonstrated, for the first time, that the treatment with newly synthesized CA-metal complexes has anticancer and antimicrobial effects, which were examined in seven different cell lines: MCF-7, MDA-MB-231, and ZR-75-1 breast cancer cell lines, A375 melanoma cell line, DLD-1 cell line, LN-229 cell line, FN cell line; five bacterial strains: Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Proteus vulgaris, Lactobacillus rhamnosus, yeast Sacchcaromyces boulardii, and pathogenic yeast-like fungi Candida albicans. The presented study indicates that CA-metal complexes could be considered as a potential supplementary tool in anticancer therapy, however, because of their possible toxic activity on fibroblasts, they should be used with caution. Some of the tested complexes have also preservative properties and positive influence on normal non-pathogenic microorganisms, which was demonstrated in selected microbial strains, therefore they may serve as food preservatives of natural origin with cytoprotective properties.
Subject(s)
Anti-Infective Agents/administration & dosage , Antineoplastic Agents/administration & dosage , Bacteria/drug effects , Caffeic Acids/pharmacology , Coordination Complexes/administration & dosage , Plant Extracts/pharmacology , Succinates/pharmacology , Yeasts/drug effects , Anti-Infective Agents/adverse effects , Anti-Infective Agents/pharmacology , Anti-Infective Agents/therapeutic use , Antineoplastic Agents/adverse effects , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Caffeic Acids/therapeutic use , Candida albicans/drug effects , Cell Line, Tumor , Coordination Complexes/adverse effects , Coordination Complexes/pharmacology , Coordination Complexes/therapeutic use , Fibroblasts/drug effects , Humans , Ions , Metals/pharmacology , Metals/therapeutic use , Microbial Sensitivity Tests , Neoplasms/drug therapy , Plant Extracts/therapeutic use , Saccharomyces boulardii/drug effects , Succinates/therapeutic useABSTRACT
Prostate cancer (PCa) is the second most frequent cancer in men and the fifth most common cause of death worldwide, with an estimated 378,553 deaths in 2020. Prostate cancer shows a strong tendency to form metastatic foci in the bones. A number of interactions between cancer cells attacking bones and cells of the bone matrix lead to destruction of the bone and growth of the tumour. The last few decades have seen increased interest in the precise role of minerals in human health and disease. Tumour cells accumulate various minerals that promote their intensive growth. Bone, as a storehouse of elements, can be a valuable source of them for the growing tumour. There are also reports suggesting that the presence of some tumours, e.g., of the breast, can adversely affect bone structure even in the absence of metastasis to this organ. This paper presents the effect of chronic dietary intake of calcium, iron and zinc, administered in doses corresponding maximally to twice their level in a standard diet, on homeostasis of selected elements (Ca, K, Zn, Fe, Cu, Sr, Ni, Co, Mn and Mo) in the femoral bones of healthy rats and rats with implanted cancer cells of the LNCaP line. The experiment was conducted over 90 days. After the adaptation period, the animals were randomly divided into four dietary groups: standard diet and supplementation with Zn, Fe and Ca. Every dietary group was divided into experimental group (with implanted cancer cells) and control group (without implanted cancer cells). The cancer cells (LnCaP) were implanted intraperitoneally in the amount 1 × 106 to the rats at day 90 of their lifetime. Bone tissue was dried and treated with microwave-assisted mineral digestation. Total elemental content was quantified by ICP-MS. Student's t-test and Anova or Kruskal-Wallis tests were applied in order to compare treatment and dietary groups. In the case of most of the diets, especially the standard diet, the femoral bones of rats with implanted LNCaP cells showed a clear downward trend in the content of the elements tested, which may be indicative of slow osteolysis taking place in the bone tissue. In the group of rats receiving the standard diet, there were significant reductions in the content of Mo (by 83%), Ca (25%), Co (22%), Mn (13%), K (13%) and Sr (9%) in the bone tissue of rats with implanted LNCaP cells in comparison with the control group receiving the same diet but without LNCaP implantation. Supplementation of the rat diet with calcium, zinc and iron decreased the frequency of these changes relative to the standard diet, which may indicate that the diet had an inhibitory effect on bone resorption in conditions of LNCaP implantation. The principal component analysis (PCA) score plot confirms the pronounced effect of implanted LNCaP cells and the standard diet on bone composition. At the same time, supplementation with calcium, zinc and iron seems to improve bone composition. The microelements that most often underwent quantitative changes in the experimental conditions were cobalt, manganese and molybdenum.
Subject(s)
Bone Density/drug effects , Dietary Supplements , Femur/metabolism , Metals/pharmacology , Prostatic Neoplasms , Animals , Cell Line, Tumor , Femur/pathology , Humans , Male , Neoplasm Transplantation , Prostatic Neoplasms/diet therapy , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Rats , Rats, Sprague-DawleyABSTRACT
A 61.3 kDa Phenol hydroxylase (PheA) was purified and characterized from Pseudomonas sp. KZNSA (PKZNSA). Cell free extract of the isolate grown in mineral salt medium supplemented with 600 ppm phenol showed 21.58 U/mL of PheA activity with a specific activity of 7.67 U/mg of protein. The enzyme was purified to 1.6-fold with a total yield of 33.6%. The purified PheA was optimally active at pH 8 and temperature 30 °C, with ≈95% stability at pH 7.5 and temperature 30 °C after 2 h. The Lineweaver-Burk plot showed the vmax and Km values of 4.04 µM/min and 4.03 µM, respectively, for the substrate phenol. The ES-MS data generated from the tryptic digested fragments of pure protein and PCR amplification of a ≈600 bp gene from genomic DNA of PKZNSA lead to the determination of complete amino acid and nucleotide sequence of PheA. Bioinformatics tools and homology modelling studies indicated that PheA from PKZNSA is likely a probable protein kinase UbiB (2-octaprenylphenol hydroxylase) involving Lys and Asp at positions 153 and 288 for binding and active site, respectively. Characterization and optimization of PheA activity may be useful for a better understanding of 2,4-dichlorophenol degradation by this organism and for potential industrial application of the enzyme.
Subject(s)
Mixed Function Oxygenases/chemistry , Mixed Function Oxygenases/isolation & purification , Models, Molecular , Pseudomonas/enzymology , Amino Acid Sequence , Base Sequence , Biophysical Phenomena , Enzyme Inhibitors/pharmacology , Enzyme Stability/drug effects , Hydrogen-Ion Concentration , Ions , Metals/pharmacology , Phylogeny , Pseudomonas/genetics , RNA, Ribosomal, 16S/genetics , Substrate Specificity/drug effects , TemperatureABSTRACT
Phytase is one of the most effective feed additives to increase the availability of phosphorus and minerals by catalyzing the hydrolysis of phytic acid. A modified appA gene (mappA) was transformed into soybean (Glycine max) under the control of a seed-specific promoter from common bean (Phaselous vulgaris). The soybean recombinant phytase showed optimal activity at pH 4.5 and 70 °C. A slight increase in enzyme activity occurred when the recombinant enzyme was pre-incubated with n-hexane. In addition, the phytase activity from our transgenic soybean does not reduce even after 2 hours of extraction with n-hexane at 55~65 °C. In conclusion, the oil extraction process using n-hexane does not inactivate the phytase expressed in the mAppA transgenic soybean, and the meal derived from the transgenic soybean processing can be used as feed supplement to livestock.
Subject(s)
6-Phytase/genetics , 6-Phytase/metabolism , Animal Feed/analysis , Glycine max/genetics , Temperature , Enzyme Stability , Gene Expression , Kinetics , Metals/pharmacology , Plants, Genetically Modified , Solvents/pharmacologyABSTRACT
We aimed to determine whether the use of injectable vitamins and minerals improves growth performance and immune and antioxidant responses in dairy calves during pre- and post-weaning period in summer. Twenty dairy calves (45 days of age) were randomized to two groups (10 each): control group (CON) and treated group [TREAT; injection providing 0.20, 0.80, 0.20, 0.10, 35 and 1â¯mg/kg of copper, zinc, manganese selenium, and vitamins A and E, during two periods (15 days pre- and 15 days post-weaning)]. The animals were weighed and blood samples were collected on days 1, 15, 30 and 45 of the study. Levels of serum copper, selenium, zinc, and manganese were measured on day 1; and the results showed that calves were not deficient in these minerals. The TREAT group had greater BW gain during the final third of the experiment. There was an increase in total leukocyte numbers as a result of elevation in neutrophil counts (day 45) and monocytes (days 30 and 45) in the TREAT group. This group also had lower reactive oxygen species (ROS) content (days 15, 30 and 45) and lipid peroxidation (LPO; days 15 and 45). Furthermore, the TREAT group had greater antioxidant capacity against peroxyl radicals (ACAP; days 15 and 30), activities of the enzymes glutathione peroxidase (GPx; days 15, 30 and 45) and superoxide dismutase (SOD; day 15), concentrations of total serum proteins (day 30), serum globulin (days 15 and 30), ceruloplasmin (day 15), tumor necrosis factor-alpha (TNF-α), interleukin-1, (IL-1; days 30 and 45) and interferon gamma (IFNγ; day 45), compared to CON group. High respiratory rates during hot times of the day in all study calves was suggestive of heat stress. Taken together, the data suggest that mineral and vitamins injections increased the growth performance and boosted the antioxidant and immunological systems of dairy calves during the diet transition period in summer.
Subject(s)
Cattle , Dietary Supplements , Metals/pharmacology , Selenium/pharmacology , Vitamin A/pharmacology , Vitamin E/pharmacology , Vitamins/pharmacology , Animals , Cattle/blood , Cattle/growth & development , Cattle/immunology , Cattle/metabolism , Cytokines/blood , Diet/veterinary , Female , Glutathione Peroxidase/blood , Heat-Shock Response/physiology , Metals/blood , Reactive Oxygen Species/blood , Respiratory Rate , Seasons , Selenium/blood , Superoxide Dismutase/bloodABSTRACT
The current study was undertaken to investigate the immunomodulatory and protective effects of Withania somnifera (WS) extract and Withaferin A (WA) supplementation on zinc oxide nanoparticles mediated toxicity in Balb/c mice. The animals were exposed to ZnO NPs along with WS and WA for 28 days and various parameters like body weight, organ coefficient, cytotoxicity, nitric oxide (NO), total serum protein, phagocytosis, and the gene expression levels of TLR6 and ARG genes were determined. In vivo study showed that, dose-dependent reduction in phagocytosis, an increase in the levels of NO production along with up-regulation of TLR6, arginase gene was significant (P < 0.05) when ZnO NPs were given. However toxicity of ZnO NP was reduced in presence of WS and WA with decreased TLR6 over expression and restoration of phagocytic activities. Our results provided a valuable insight into the underlying mechanism for the protective effects of WS. Mechanism of toxicity induced by Zinc oxide nanoparticles ZnO NPs and immunomodulatory protective effects of Withania somnifera extract (WS) and Withaferin A (WA), in Balb/c mice modal of peritoneal macrophages. Red arrows: effect of ZnO NPs independently leads to ROS production which attenuated the phagocytosis of yeast by macrophages through, up-regulation of TLR6 and down-regulation of arginase gene expression. Green arrows: co-treatment, Impact of Withania somnifera extract with zinc oxide nanoparticles (WS + ZnO NPs), Withaferin A along with zinc oxide nanoparticles (WA + ZnO NPs)-enhance phagocytic activity by counteracting mechanism of ZnO NPs toxicity. Black arrows: increasing or decreasing effects. Per oral (P.O).
Subject(s)
Oxidative Stress/drug effects , Withania/chemistry , Zinc Oxide/toxicity , Animals , Cell Line , Immunologic Factors/pharmacology , Macrophages/drug effects , Male , Metal Nanoparticles , Metals/pharmacology , Mice , Mice, Inbred BALB C , Nanoparticles/toxicity , Nitric Oxide , Plant Extracts/pharmacology , Plant Roots/drug effects , Withania/immunology , Withania/metabolism , Withanolides/pharmacology , Zinc Oxide/adverse effectsABSTRACT
US Food and Drug Administration (FDA) permitted for direct addition of castor oil into food for human consumption and recently FDA approved castor oil as over-the-counter (OTC) for laxative drug. Castor oil (COL) is a vegetable oil and naturally polyol which is inexpensive, environmental friendly and a very valuable renewable resource. Metals are believed to influence antioxidant property of ligands. Metals copper, zinc, cobalt and barium were incorporated with castor oil and subsequently structures were established by FT-IR, UV-Visible, and 1H NMR spectroscopic techniques. In vitro antioxidant activities of metal containing COL were determined by DPPH and superoxide scavenging methods and the results were compared with vitamin C. Enhanced antioxidant nature of metal containing castor oil was noticed and compared to virgin castor oil. This study reveals that synthesized metal containing-COL is a potential antioxidant material.
Subject(s)
Antioxidants/pharmacology , Castor Oil/chemistry , Castor Oil/pharmacology , Metals/pharmacology , Antioxidants/chemistry , Ascorbic Acid/pharmacology , Cations, Divalent , Free Radical Scavengers/pharmacology , Ligands , Magnetic Resonance Spectroscopy , Metals/chemistry , Spectroscopy, Fourier Transform Infrared , Superoxides/metabolismABSTRACT
This study reports the photocatalytic degradation of Methylene Blue (MB) dye (a class of dyestuffs that are resistant to biodegradation) under the influence of UV-light irradiation. Antibacterial and antibiofilm activities of ferrite nanoparticles (FO NPs) were examined against some pathogenic bacteria isolated from the medical operating room surfaces. In the same context, metals-substituted spinel cobalt ferrite nanoparticles with nominal composition [MxCo1-xFe2O4 NPs; (Mâ¯=â¯Zn, Cu, Mn; xâ¯=â¯0.0, 0.25, 0.5 and 0.75)] were synthesized by citrate sol-gel method. Also, the structures of the synthesized FO NPs were characterized by X-ray diffraction, and Williamson-Hall (WH) method was used to determine the crystallite size. The estimated specific surface area is found in the range from 37.99 to 107.05â¯m2/g, between the synthesized ferrites, Zn0.5Co0.5Fe2O4 NPs have average pore radius 1.84â¯nm and the pore volume was 0.136â¯ml/g. SEM images revealed that, the synthesized FO NPs have an unique pores and uniformly distribution, while EDX spectra shows the elemental composition for the synthesized FO NPs. The elastic properties of FO NPs have been estimated using FTIR data, whereas (Mâ¯-â¯H) hysteresis loops revealed that, by replacing cobalt ions with Zn, Cu, and Mn ions the magnetic behaviour changed from ferromagnetic to paramagnetic. Results obtained from the photocatalysis indicated that Mn0.75Co0.25Fe2O4 NPs (30.0â¯mg) were a promising photocatalyst achieving 96.0% removal of MB after 100â¯min of UV-light exposure in the alkaline solution. Antibacterial results showed that the most effective combination was Zn0.75Co0.25Fe2O4 NPs (20.0â¯ppm) displaying activity against Staphylococcus aureus, Enterococcus columbae, and Aerococcus viridians by 15.0, 13.0, and 12.0â¯mm ZOI, respectively. Additionally, Zn0.75Co0.25Fe2O4 NPs were active as antibiofilm factors producing activity by 63.7, 57.9, and 45.5% towards S. aureus, A. viridians, and E. columbae, respectively. Accordingly, Zn0.75Co0.25Fe2O4 and Mn0.75Co0.25Fe2O4 NPs can be utilized in industrial, biological and medical applications.