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关键词： Gilthead sea bream   Feeding frequency   Fishmeal and fish oil level   Digestive enzymes   Feed digestibility  

摘要： In the context of Mediterranean aquaculture many efforts have been made in terms of reducing marine-derived ingredients in aquafeed formulation. On the other hand, little attention has been paid to the manipulation of feeding frequency at the on-growing phase, where the high costs related to feeding procedures and the optimisation of feed efficiency and fish health are key aspects for the economic and environmental sustainability of the production cycle. The effects of different feeding frequencies (F) (1F) one meal day(-1), (2F) two meals day(-1), (3F) three meals day(-1) on growth, digestive enzyme activity, feed digestibility and plasma biochemistry were studied in gilthead sea bream (Sparus aurata, L. 1758) fed with high (FM30/FO15, 30% fishmeal FM, 15% fish oil, FO) and low (FM10/FO3;10% FM and 3% FO) FM and FO levels. Isonitrogenous and isolipidic extruded diets were fed to triplicate fish groups (initial weight: 88.3 +/- 2.4 g) by a fixed ration over 109 days. No significant effects of feeding frequency on overall performance, feed efficiency and feed digestibility during the on-growing of gilthead sea bream fed high or low fishmeal and fish oil dietary level were observed. Pepsin activity showed an apparent decrease in fish receiving more than one meal a day which was not compensated by an increased production of alkaline proteases (either trypsin or chymotrypsin), particularly in fish fed on low FM. Although there were no effects on growth and feed utilisation at increasing feeding frequency, trypsin decreased significantly with an increasing number of meals only under low FMFO diet. Thus, it seemed that consecutive meals could have amplified the potential trypsin inhibitor effect of the vegetable meal-based diet adopted. Most of the plasma parameters related to nutritional and physiological conditions were not affected by feeding frequency, however an effect on electrolytes (Na+, Cl), cortisol and creatinine was observed. The higher level of plasma creatin

关键词： Chondroitin sulfate/dermatan sulfate   CSase ABC   Marine bacteria  

摘要： Chondroitin sulfate (CS)/dermatan sulfate (DS) lyases play important roles in structural and functional studies of CS/DS. In this study, a novel CS/DS lyase (enCSase) was identified from the genome of the marine bacterium Photobacterium sp. QA16. This enzyme is easily heterologously expressed and purified as highly active form against various CS, DS and hyaluronic acid (HA). Under the optimal conditions, the specific activities of this enzyme towards CSA, CSC, CSD, CSE, DS and HA were 373, 474, 171, 172, 141 and 97 U/mg of proteins, respectively. As an endolytic enzyme, enCSase degrades HA to unsaturated hexa- and tetrasaccharides but CS/DS to unsaturated tetra- and disaccharides as the final products. Sequencing analysis showed that the structures of tetrasaccharides in the final products of CS variantswere not unique butwere highly variable, indicating the randomness of substrate degradation by this enzyme. Further studies showed that the smallest substrate of enCSase was octasaccharide for HA but hexasaccharide for CS/DS, which could explain why this enzyme cannot degrade HAhexa- and tetrasaccharides and CS/DS tetrasaccharides further. It is believed that enCSasemay be a very useful tool for structural and functional studies and related applications of CS/DS and HA. (C) 2020 Elsevier B.V. All rights reserved.

关键词： Pullulan-C-60   Levan-C-60   Synchrotron radiation FTIR spectro-microscopy  

摘要： Objects of the present study are improved fullerene C-60 drug carrier properties trough encapsulation by microbial polysaccharides, levan (LEV), pullulan (PUL), and their hydrophobized cholesterol-derivatives (CHL and CHP), that show better interaction with cancer cells. The zeta potential, polydispersity index, and the diameter of particles were determined, and their cytotoxicity against three cancer cell lines were tested. Biochemical changes in HeLa cells are analyzed by synchrotron radiation (SR) FTIR spectro-microscopy combined with the principal component analysis (PCA). The most significant changes occur in HeLa cells treated with LEV-C-60 and correspond to the changes in the protein region, i.e. Amide I band, and the changes in the structure of lipid bodies and membrane fluidity are evident. The highest cytotoxicity was also induced by LEV-C-60. In HeLa cells, cytotoxicity could not be strictly associatedwith biochemical changes in lipids, proteins and nucleic acids, but these findings are significant contribution to the study of the mechanism of interaction of C-60-based nanoparticles with cellular biomolecules. In conclusion, LEV, PUL, CHL, and CHP enhanced fullerene C-60 potential to be used as target drug delivery system with the ability to induce specific intracellular changes in HeLa cancer cells. (C) 2020 Elsevier B.V. All rights reserved.

关键词： Pyrobacullum calidifontis   4-alpha-glucanotransferases   Family GH77  

摘要： Pcal_0768 gene encoding an amylomaltase, a 4-alpha-glucanatransferase belonging to family 77 of glycosyl hydrolases, from Pyrobaculum calidifontis was cloned and expressed in Escherichia coli. The recombinant protein was produced in E. coli in soluble and active form. However, the expression level was not very high. Analysis of the mRNA of initial seven codons at the 5'-end of the gene revealed the presence of a hair pin like secondary structure. This secondary structure was removed by site directed mutagenesis, without altering the amino acids, which resulted in enhanced expression of the cloned gene. Recombinant Pcal_0768 exhibited optimal amylomaltase activity at 80 degrees C and pH 6.9. Under these conditions, the specific activity was 690 U/mg. Recombinant Pcal_0768 was highly thermostable with a half-life of 6 h at 100 degrees C. It exhibited the highest k(cat) value among the characterized glucanotransferases. No metal ions were required for activity or stability of the enzyme. Recombinant Pcal_0768 was successfully employed in the synthesis of modified starch for producing thermoreversible gel. To the best of our knowledge, till now this is the most thermostable enzyme among the characterized amylomaltases. High thermostability and starch modification potential make it a novel and distinct amylomaltase. (C) 2020 Elsevier B.V. All rights reserved.

关键词： Handling   Welfare   Carnivorous fish   Eugenol  

摘要： This study tested the use of essential oil of Ocimum gratissimum (EOOG) as an anesthetic for juvenile Lophiosilurus alexandri and its efficiency in mitigating the effects of handling stress. Two experiments were performed. Experiment 1 determined the time for anesthesia induction and recovery and the ventilatory frequency of juveniles exposed to different concentrations of EOOG (0, 10, 30, 90, 150 and 300 mg L-1). Based on the results of Experiment 1, Experiment 2 evaluated the effects of 0, 10 and 90 mg L-1 of EOOG on blood parameters and oxidative stress in liver and brain immediately after induction and one hour after recovery. No mortality was observed in any experiment. In Experiment 1, times for anesthesia induction were shorter, and recovery times longer, with increased EOOG concentrations. Compared to the animals anesthetized with 0, 10 and 30 mg L-1 of EOOG, higher ventilatory frequencies were observed during induction for animals anesthetized with 90, 150 and 300 mg L-1. In experiment 2, variations on hematological variables derived from EOOG exposition were determined, but parameters returned to baseline levels one hour after recovery. The use of 90 mg L-1 EOOG prevented increased plasma glucose and cortisol levels one hour after handling. However, animals anesthetized with 90 mg L-1 EOOG had higher concentrations of ROS (hepatic and brain) after induction and one hour after recovery. The increase in ROS for animals anesthetized with 90 mg L-1 was not matched by the antioxidant defense system, which showed a reduction in brain GST activity one hour after recovery. Juveniles anesthetized with 10 mg L-1 EOOG only presented an increase in hepatic ROS one hour after recovery from anesthesia, however, there was a response from the antioxidant defense system, with an increase in brain and hepatic GST. The increase in ROS in treatments submitted to handling under the effect of anesthesia did not cause tissue damage. It is concluded that the use of 90 and 150 mg

关键词： Guanidine   Peptides and proteins   Ligands   Genetics   Monomers  

摘要： An intriguing consequence of ongoing riboswitch discovery efforts is the occasional identification of metabolic or toxicity response pathways for unusual ligands. Recently, we reported the experimental validation of three distinct bacterial riboswitch classes that regulate gene expression in response to the selective binding of a guanidinium ion. These riboswitch classes, called guanidine-I, -II, and -III, regulate numerous genes whose protein products include previously misannotated guanidine exporters and enzymes that degrade guanidine via an initial carboxylation reaction. Guanidine is now recognized as the primal substrate of many multidrug efflux pumps that are important for bacterial resistance to certain antibiotics. Guanidine carboxylase enzymes had long been annotated as urea carboxylase enzymes but are now understood to participate in guanidine degradation. Herein, we report the existence of a fourth riboswitch class for this ligand, called guanidine-IV. Members of this class use a novel aptamer to selectively bind guanidine and use an unusual expression platform arrangement that is predicted to activate gene expression when ligand is present. The wide distribution of this abundant riboswitch class, coupled with the striking diversity of other guanidine-sensing RNAs, demonstrates that many bacterial species maintain sophisticated sensory and genetic mechanisms to avoid guanidine toxicity. This finding further highlights the mystery regarding the natural source of this nitrogen-rich chemical moiety.

关键词： Thioredoxin reductase   Selenocysteine   Cysteine   Antioxidant   Cancer   Parasite  

摘要： The thiol-based glutathione reductase (GR) and thioredoxin reductase (TrxR) are the major antioxidant enzymes present in various organisms that maintain the internal redox homeostasis. The thioredoxin system has attracted the attention of researchers from diverse investigation fields of biological sciences. Apart from redox regulation, this system is thought to be the major regulator of various biological processes including transcription, apoptosis, etc. Identification and physicobiochemical characterization of the reductase enzyme i.e. Thioredoxin reductase (TrxR) revealed the potency of it to become a promising target. Novel therapeutic interventions by selective targeting of TrxR in parasitic organisms as well as in the cancer cells have now become a usual treatment approach. However, different isoforms and their variation in the penultimate amino acid (Selenocysteine or cysteine) present in the catalytic site of the enzyme have made this enzyme to respond differently towards various drugs and synthetic and/or natural compounds. Therefore, the present article seeks to highlight the importance and the detailed molecular mechanism, functional perspective underlying the TrxR inhibition in various parasitic protozoans, helminthes as well as in cancer cells for devising suitable anti-TrxR candidates. (C) 2020 Elsevier B.V. All rights reserved.

关键词： Phytochemicals   Guava leaves   Nile tilapia   Cypermethrin toxicity   Hemato-biochemical variables   Immunity  

摘要： Cypermethrin (CYP) is a synthetic pyrethroid insecticide which is used for insects control in agricultural activities near the aquatic ecosystem. It could reach the aquatic environment leading to severe degradation in the fish welfare. Guava leaves extract (GLE) was used as a natural feed supplement for fish because of its stimulating effects on their growth and welfare status. The present research was aimed at investigating the protective effects of dietary GLE on growth, hemato-biochemical, and immunity response of CYP-intoxicated Nile tilapia, Oreochromis niloticus (L.). The half lethal dose of CYP for Nile tilapia fingerlings was determined and it was found to be 5.88 mu g/L and a sublethal concentration of 0.294 mu g/L of CYP was used in the present study. Fish were classified in five replicates into four groups;the first group was the control group (T1), the second group was fed a diet enriched with GLE (T2), the third group was exposed to 0.294 mu g CYP/L (T3), and the fourth group was exposed to 0.294 mu g CYP/L and co-supplemented with GLE (T4). After 6 weeks, growth, hemato-biochemical, and immunological variables were determined. Compared to the control group, the toxicity of CYP (T3) caused significant reductions in counts of white blood cells and red blood cells, concentrations of hemoglobin, hematocrit levels, and activity of acetyl cholinesterase accompanied with significant reductions in glucose and cortisol levels. In addition, CYP exposure (T3) exhibited significant immune suppression with marked decreases in total serum protein, albumin, globulin, lysozyme activity, respiratory burst activity, and total immunoglobulin than the control group (T1). On the other hand, the dietary GLE (T2) increased substantially fish growth, hematobiochemical, and immune variables. The feeding fish on a GLE-enriched diet minimized the negative impacts of CYP toxicity and normalized the above-mentioned variables to be similar to the control group. The present study ha

关键词： Hyssopus officinalis L.   Antioxidant activity   Compatible solutes   Isopinocamphone   Foliar treatment  

摘要： This study examined the effects of exogenous citrulline (control, 1 and 2 mM) and water availability (100%, 50% and 25% WA) on antioxidant attributes and essential oil constituents of Hyssopus officinalis L. in two successive harvests. Hyssop tolerantly responded to water deficiency by well-promoted antioxidant enzymes (i.e., superoxide dismutase, and catalase), strong DPPH-scavenging activity, and increasing polyphenols;however, the essential oil content was negatively reduced by water stress. External citrulline further increased the activity of antioxidant enzymes. Citrulline application at 2 mM under severe water stress could also improve essential oil (EO) content in the first and second harvests by about 15 and 30%, respectively. Furthermore, under severe drought, citrulline at 2 mM could obtain the highest yield of isopinocamphone (47%) as the main component of EO. The results showed the high potential of this novel applied metabolite agent to be used in a well-fulfilled production of this medicinal plant.

关键词： M1 class aminopeptidases   Human puromycin sensitive aminopeptidase   PSA   Selective inhibitor   Aminopeptidase N   Puromycin   O-methyl-L-tyrosine  

摘要： Puromycin sensitive aminopeptidase (PSA or NPEPPS) is a M1 class aminopeptidase is selectively inhibited by the natural product puromycin, an aminonucleoside antibiotic produced by the bacterium Streptomyces alboniger. The molecular basis for this selective inhibition has not been understood well. Here, we report the basis for selectivity of puromycin using biochemical, structural and molecular modeling tools on four different M1 family enzymes including human PSA. Except for PSA, the other three enzymes were not inhibited. Instead, the peptide bond in the puromycin is hydrolyzed to O-methyl-L-tyrosine (OMT) and puromycin aminonucleoside (PAN). Neither of the hydrolyzed products, individually or together inhibit any of the four enzymes. Crystal structure of ePepN using crystals that are incubated with puromycin contained the hydrolyzed products instead of intact puromycin. On the other hand, intact puromycin molecule was observed in the crystal structure of the inactivemutant ePepN (E298A)-puromycin complex. Surprisingly, puromycin does not enter the active site of the mutant enzyme but binds near the entrance. Comparison of puromycin binding region in ePepN mutant enzyme and molecular modeling studies suggest that PSA might be inhibited by similar mode of binding there by blocking the entrance of the active site. (C) 2020 Elsevier B.V. All rights reserved.