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Transferase Activity (transferase + activity)
Selected AbstractsEFFECT OF HIGH-PRESSURE TREATMENT OF MILK ON LIPASE AND ,-GLUTAMYL TRANSFERASE ACTIVITYJOURNAL OF FOOD BIOCHEMISTRY, Issue 6 2004P. K. PANDEY ABSTRACT High-pressure (HP) treatment (0,180 min at 300,400 MPa) was applied to milk to evaluate the pressure effects on the activities of lipoprotein lipase and ,-glutamyl transferase. Short time pressure exposure resulted in some enhancement in the activity of both enzymes, and for lipase, there was no inactivation during the entire pressure hold time (up to 100 min). With ,-glutamyl transferase, the extent of enhancement in activity was pressure level dependent, with lower pressure resulting in a greater enhancement. Furthermore, longer pressure treatment times resulted in inactivation of ,-glutamyl transferase, following a first order kinetic model. The pressure sensitivity of the inactivation parameters (k and D -values) for ,-glutamyl transferase was adequately described by the pressure death time and Arrhenius models with a zpof 543 MPa and an associated volume of activation, ,V,, of ,3.28 × 10,8 m3/mole. [source] Cholinesterase and glutathione- S -transferase activities in freshwater invertebrates as biomarkers to assess pesticide contaminationENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 1 2010Inês Domingues Abstract Studies investigating the use of biomarkers in pesticide risk assessment have greatly increased in recent years; however, issues concerning the ecological meaning of enzymatic responses have proved controversial. Ideally a good biomarker response should be modulated by the environmental contaminants alone and demonstrate a predictable behavior towards certain types of toxins. As these premises are rarely observed, the present study aims to outline research that has contributed to an understanding of the behavior of two widely used biomarkers, cholinesterase and glutathione- S -transferase, describing environmental and biotic factors that affect their response in freshwater invertebrates. Studies were performed in the main classes of aquatic invertebrates with these biomarkers and conclusions were reached concerning their behavior towards the main classes of pesticides. Links between biomarker responses and conventional endpoints were evaluated so that ecological relevance could be attributed to enzymatic responses. Toxicity of mixtures was investigated, and cases of synergism and antagonism were pointed out as factors changing the expected toxicity of aquatic systems and leading to misinterpretations of biomarker responses. Finally, the use of biomarkers as a tool for biomonitoring and in situ assays was investigated, with discussion of advantages and disadvantages of their use. Environ. Toxicol. Chem. 2010;29:5,18. © 2009 SETAC [source] Attenuation of TCDD-induced oxidative stress by 670 nm photobiomodulation in developmental chicken kidneyJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 4 2008Jinhwan Lim 2,3,7,8-Tetrachlorodibenzo- p -dioxin (TCDD), a potent developmental teratogen inducing oxidative stress and sublethal changes in multiple organs, provokes developmental renal injuries. In this study, we investigated TCDD-induced biochemical changes and the therapeutic efficacy of photobiomodulation (670 nm; 4 J/cm2) on oxidative stress in chicken kidneys during development. Eggs were injected once prior to incubation with TCDD (2 pg/g or 200 pg/g) or sunflower oil vehicle control. Half of the eggs in each dose group were then treated with red light once per day through embryonic day 20 (E20). Upon hatching at E21, the kidneys were collected and assayed for glutathione peroxidase, glutathione reductase, catalase, superoxide dimutase, and glutathione- S -transferase activities, as well as reduced glutathione and ATP levels, and lipid peroxidation. TCDD exposure alone suppressed the activity of the antioxidant enzymes, increased lipid peroxidation, and depleted available ATP. The biochemical indicators of oxidative and energy stress in the kidney were reversed by daily phototherapy, restoring ATP and glutathione contents and increasing antioxidant enzyme activities to control levels. Photobiomodulation also normalized the level of lipid peroxidation increased by TCDD exposure. The results of this study suggest that 670 nm photobiomodulation may be useful as a noninvasive treatment for renal injury resulting from chemically induced cellular oxidative and energy stress. © 2008 Wiley Periodicals, Inc. J Biochem Mol Toxicol 22:230,239, 2008; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20233 [source] Phosphatidylinositol Synthase of Tetrahymena: Inositol Isomers as Substrates in Phosphatidylinositol Biosynthesis and Headgroup Exchange ReactionsTHE JOURNAL OF EUKARYOTIC MICROBIOLOGY, Issue 2 2007BRIDGET M. RIGGS ABSTRACT. Phosphatidylinositol (PtdIns) synthase in microsomal fractions derived from Tetrahymena vorax was studied to determine its activity requirements. The suitability of inositol isomers as substrates for the synthase and in headgroup exchange reactions also was investigated. Tetrahymena PtdIn synthase activity was optimum in the presence of 2 mM MgCl2 plus 2 mM MnCl2, a pH of 7.8, and a temperature of 30°C. The enzyme retained approximately 80% of its activity after incubation at 70°C for 10 min. PtdIns headgroup exchange activity was maximal in the presence of cytidine monophosphate. By following either the accumulation of radiolabeled reaction products or the loss of radiolabel from precursors, each of the inositol isomers tested appeared to serve as substrates for both the PtdIns synthase and PtdIns:inositol phosphatidyl transferase activities. In each case, myo -inositol and scyllo -inositol were the preferred substrates. The data suggest two routes for the formation of phosphatidyl-non- myo -inositols in Tetrahymena and the potential for the production of novel, non- myo -inositol-containing second messengers. [source] Myo -inositol prevents oxidative damage, inhibits oxygen radical generation and increases antioxidant enzyme activities of juvenile Jian carp (Cyprinus carpio var. Jian)AQUACULTURE RESEARCH, Issue 15 2009Wei-Dan Jiang Abstract This study was conducted to evaluate the effects of dietary myo -inositol (MI) on the antioxidant status of juvenile Jian carp (Cyprinus carpio var. Jian). A total of 1050 Jian carp (22.28±0.07 g) were randomly distributed into seven groups of three replicates each, feeding diets containing graded levels of MI (163.5, 232.7, 384.2, 535.8, 687.3, 838.8 and 990.3 mg kg,1 diet) for 60 days. Results indicated that the malondialdehyde content was the lowest for fish fed diets containing ,384.2 mg MI kg,1, and the highest for fish fed the MI-unsupplemented basal diet (P<0.05). The protein carbonyl content was decreased with increasing dietary MI levels up to 535.8 mg kg,1 diet, and no differences were found with a further increase in the MI concentration. The anti-superoxide anion capacity (ASA) and anti-hydroxyl radical capacity (AHR) were increased with increasing MI levels up to 535.8 mg kg,1 diet, and plateaued thereafter. The superoxide dismutase and glutathione- S -transferase activities showed the same tendency with the ASA capacity. Catalase, glutathione peroxidase and glutathione reducase activities were improved with increasing MI levels up to 838.8, 384.2 and 687.3 mg kg,1 diet, respectively, and remained nearly constant thereafter. These results suggested that MI could inhibit oxygen radical generation, increase enzymatic antioxidant capacity and prevent oxidative damage of carp. Dietary MI requirements for ASA and AHR activities of juvenile Jian carp were 567.94 and 517.22 mg MI kg,1 diet respectively. [source] Sublethal responses of wolf spiders (Lycosidae) to organophosphorous insecticidesENVIRONMENTAL TOXICOLOGY, Issue 5 2002S. Van Erp Abstract The activities of cholinesterase (ChE) and glutathione S -transferase (GST) enzymes were assessed in the wolf spider (Lycosa hilaris) as biomarkers of organophosphate contamination in agricultural ecosystems. Spiders were exposed to simulated field rates of two commercially available organophosphorous insecticides [Basudin (diazinon) and Lorsban (chlorpyrifos)] under laboratory conditions. In terms of survival, chlorpyrifos and diazinon were more toxic to male than to female wolf spiders, but gender-specific differences in ChE activities were not evident. Cholinesterase activity in male spiders was inhibited to 14% and 61% of control activity by Basudin and Lorsban, respectively. Gluthathione S -transferase activity was not affected by either pesticide. Mortality and biomarker responses in the wolf spider were further investigated following the application of Basudin to pasture. Wolf spiders were deployed into field mesocosms; after 24 h mortality was 40%, and surviving spiders displayed significant inhibition of ChE activity (87%) compared with controls. Cholinesterase activity in spiders exposed for subsequent 24- or 48-h time periods was monitored until it returned to control levels 8 days post-application. Inhibition of ChE activity after a single application of Basudin indicate the potential use of this enzyme in wolf spiders as a biomarker for evaluating organophosphate contamination. © 2002 Wiley Periodicals, Inc. Environ Toxicol 17: 449,456, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/tox.10078 [source] Obesity as a cause of "false-positive" alcohol misuse laboratory investigationsADDICTION BIOLOGY, Issue 4 2002T. J. Peters Three patients are reported with a prior history of alcohol misuse accompanied by abnormal liver toxicity tests and other indices of alcohol misuse. A decreased but persistently raised serum , -glutamyl transferase activity during subsequent abstinence or controlled drinking was interpreted incorrectly as due to continued alcohol misuse whereas obesity-related fatty liver was the probable cause. The value of serum carbohydrate deficient transferrin assays in the differential diagnosis of abnormal liver toxicity tests is emphasized. [source] Escherichia coli thioredoxin inhibition by cadmiumFEBS JOURNAL, Issue 7 2004Asp2, Two mutually exclusive binding sites involving Cys3 Observations of thioredoxin inhibition by cadmium and of a positive role for thioredoxin in protection from Cd2+ led us to investigate the thioredoxin,cadmium interaction properties. We used calorimetric and spectroscopic methods at different pH values to explore the relative contribution of putative binding residues (Cys32, Cys35, Trp28, Trp31 and Asp26) within or near the active site. At pH 8 or 7.5 two binding sites were identified by isothermal titration calorimetry with affinity constants of 10 × 106 m,1 and 1 × 106 m,1. For both sites, a proton was released upon Cd2+ binding. One mole of Cd2+ per mole of reduced thioredoxin was measured by mass spectrometry at these pH values, demonstrating that the two binding sites were partially occupied and mutually exclusive. Cd2+ binding at either site totally inhibited the thiol,disulfide transferase activity of Trx. The absence of Cd2+ interaction detected for oxidized or alkylated Trx and the inhibition of the enzymatic activity of thioredoxin by Cd2+ supported the role of Cys32 at the first site. The fluorescence profile of Cd2+ -bound thioredoxin differed, however, from that of oxidized thioredoxin, indicating that Cd2+ was not coordinated with Cys32 and Cys35. From FTIR spectroscopy, we inferred that the second site might involve Asp26, a buried residue that deprotonates at a rather high and unusual pKa for a carboxylate (7.5/9.2). The pKa of the two residues Cys32 and Asp26 have been shown to be interdependent [Chivers, T. P. (1997) Biochemistry36, 14985,14991]. A mechanism is proposed in which Cd2+ binding at the solvent-accessible thiolate group of Cys32 induces a decrease of the pKa of Asp26 and its deprotonation. Conversely, interaction between the carboxylate group of Asp26 and Cd2+ at a second binding site induces Cys32 deprotonation and thioredoxin inhibition, so that Cd2+ inhibits thioredoxin activity not only by binding at the Cys32 but also by interacting with Asp26. [source] Molecular cloning, expression and characterization of cDNA encoding cis -prenyltransferases from Hevea brasiliensisFEBS JOURNAL, Issue 23 2003A key factor participating in natural rubber biosynthesis Natural rubber from Hevea brasiliensis is a high molecular mass polymer of isoprene units with cis -configuration. The enzyme responsible for the cis -1,4-polymerization of isoprene units has been idengified as a particle-bound rubber transferase, but no gene encoding this enzyme has been cloned from rubber-producing plants. By using sequence information from the conserved regions of cis -prenyl chain elongating enzymes that were cloned recently, we have isolated and characterized cDNAs from H. brasiliensis for a functional factor participating in natural rubber biosynthesis. Sequence analysis revealed that all of the five highly conserved regions among cis -prenyl chain elongating enzymes were found in the protein sequences of the Hevea cis -prenyltransferase. Northern blot analysis indicated that the transcript(s) of the Hevea cis -prenyltransferase were expressed predominantly in the latex as compared with other Hevea tissues examined. In vitro rubber transferase assays using the recombinant gene product overexpressed in Escherichia coli revealed that the enzyme catalyzed the formation of long chain polyprenyl products with approximate sizes of 2 × 103,1 × 104 Da. Moreover, in the presence of washed bottom fraction particles from latex, the rubber transferase activity producing rubber product of high molecular size was increased. These results suggest that the Hevea cis -prenyltransferase might require certain activation factors in the washed bottom fraction particles for the production of high molecular mass rubber. [source] Protein farnesyltransferase inhibitors interfere with farnesyl diphosphate binding by rubber transferaseFEBS JOURNAL, Issue 19 2003Christopher J. D. Mau Rubber transferase, a cis -prenyltransferase, catalyzes the addition of thousands of isopentenyl diphosphate (IPP) molecules to an allylic diphosphate initiator, such as farnesyl diphosphate (FPP, 1), in the presence of a divalent metal cofactor. In an effort to characterize the catalytic site of rubber transferase, the effects of two types of protein farnesyltransferase inhibitors, several chaetomellic acid A analogs (2, 4,7) and ,-hydroxyfarnesylphosphonic acid (3), on the ability of rubber transferase to add IPP to the allylic diphosphate initiator were determined. Both types of compounds inhibited the activity of rubber transferases from Hevea brasiliensis and Parthenium argentatum, but there were species,specific differences in the inhibition of rubber transferases by these compounds. Several shorter analogs of chaetomellic acid A did not inhibit rubber transferase activity, even though the analogs contained chemical features that are present in an elongating rubber molecule. These results indicate that the initiator-binding site in rubber transferase shares similar features to FPP binding sites in other enzymes. [source] RMF inactivates ribosomes by covering the peptidyl transferase centre and entrance of peptide exit tunnelGENES TO CELLS, Issue 4 2004Hideji Yoshida In gram-negative bacteria such as Escherichia coli, protein synthesis is suppressed by the formation of 100S ribosomes under stress conditions. The 100S ribosome, a dimer of 70S ribosomes, is formed by ribosome modulation factor (RMF) binding to the 70S ribosomes. During the stationary phase, most of the 70S ribosomes turn to 100S ribosomes, which have lost translational activity. This 100S formation is called the hibernation process in the ribosome cycle of the stationary phase. If stationary phase cells are transferred to fresh medium, the 100S ribosomes immediately go back to active 70S ribosomes, showing that inactive 100S , active 70S interconversion is a major system regulating translation activity in stationary phase cells. To elucidate the mechanisms of translational inactivation, the binding sites of RMF on 23S rRNA in 100S ribosome of E. coli were examined by a chemical probing method using dimethyl sulphate (DMS). As the results, the nine bases in 23S rRNA were protected from DMS modifications and the modification of one base was enhanced. Interestingly A2451 is included among the protected bases, which is thought to be directly involved in peptidyl transferase activity. We conclude that RMF inactivates ribosomes by covering the peptidyl transferase (PTase) centre and the entrance of peptide exit tunnel. It is surprising that the cell itself produces a protein that seems to inhibit protein synthesis in a similar manner to antibiotics and that it can reversibly bind to and release from the ribosome in response to environmental conditions. [source] Effects of rumen-protected methionine in a low protein ration on metabolic traits and performance of early lactating cows as opposed to rations with elevated crude protein contentJOURNAL OF ANIMAL PHYSIOLOGY AND NUTRITION, Issue 5 2000T. F. Kröber Summary A 5-week experiment with 24 multiparous early lactating Brown Swiss cows was conducted to investigate the effects of supplementary rumen-protected methionine in conjunction with dietary protein reduction on metabolism and performance after 1 week of control measurement. Three rations containing 175, 150 and 125 g of crude protein/kg feed dry matter were supplemented with methionine. The fourth ration, also only containing 125 g of crude protein/kg dry matter, remained unsupplemented. The four treatment groups had a similar metabolic supply of other essential amino acids, protein and energy, as calculated by various approaches. The two low protein rations were, however, slightly deficient in ruminally degraded protein. Treatment effects remained low on feed intake, forage meal pattern, milk yield and fat as well as lactose content. In contrast, the content and yield of milk protein significantly declined only in the unsupplemented low protein ration relative to the initial value. Compared with this ration, the decline in milk protein yield was clearly delayed in the supplemented low protein ration. Blood plasma methionine tended to be reduced without supplementation and to be increased with additional methionine. Supplementation of methionine reduced other plasma amino acids. Plasma insulin, glucose, lactate, ketone bodies and aspartate amino transferase activity indicated a certain liver stress and a somewhat elevated energy requirement with high and particularly with low protein content (when unsupplemented). Methionine improved metabolic protein utilization, followed by the lowest plasma, urine and milk urea levels in the supplemented low protein diet. In conclusion, no major adverse effects were assessed under the conditions tested. Supplementation of methionine may nevertheless be useful in rations with particularly low protein content fed to early lactating cows in order to prevent negative long-term effects which were only visible here as trends. Zusammenfassung Auswirkungen von pansengeschütztem Methionin in einer Niedrigproteinration im Vergleich zu Rationen mit erhöhtem Rohproteingehalt auf Stoffwechselmerkmale und Leistung von frischlaktierenden Milchkühen In einem fünfwöchigen Experiment mit 24 frischlaktierenden Braunviehkühen wurden die Auswirkungen einer Ergänzung mit pansengeschütztem Methionin bei gleichzeitiger Reduktion der Proteinzufuhr nach einer einwöchigen Kontrollphase geprüft. Drei Rationen mit 175, 150 und 125 g Rohprotein/kg T wurden mit Methionin ergänzt. Eine weitere Variante, ebenfalls nur mit 125 g Rohprotein/kg T, wurde nicht supplementiert. Die vier Varianten stellten gemäß verschiedener Futterbewertungsysteme eine vergleichbare metabolische Versorgung mit den übrigen essentiellen Aminosäuren, Protein und Energie sicher. Die Niedrigproteinvarianten enthielten allerdings etwas zu wenig pansenabbaubares Protein. Futteraufnahme, Muster des Grundfutterverzehrs, Milchleistung sowie Fett-und Laktosegehalt der Milch zeigten nur geringe Reaktion auf die Behandlungen. Milchproteingehalt und -menge waren nur in der nicht ergänzten Niedrigproteinvariante relativ zum Ausgangswert signifikant verringert. Im Vergleich zur unsupplementierten Niedrigproteinration war dagegen der Abfall mit Ergänzung deutlich verzögert. Gegenüber dem Ausgangswert war die Methioninkonzentration im Blutplasma ohne Ergänzung tendenziell erniedrigt, mit Ergänzung erhöht. Es erfolgte eine Verringerung der Plasmakonzentration anderer Aminosäuren durch die Methioninergänzung der Niedrigproteinration. Die Plasmaniveaus von Insulin, Glucose, Laktat, Ketonkörpern und Aspartataminotransferase-Aktivität lassen auf eine gewisse Leberbelastung und einen etwas höheren Energiebedarf mit hohem und besonders mit niedrigem Proteingehalt (unsupplementiert) schließen. Die Zulage an Methionin verbesserte die metabolische Proteinverwertung, so dass die Harnstoffgehalte in Blut, Milch und Harn in dieser Niedrigproteinvariante am niedrigsten waren. Insgesamt ergaben sich keine grösseren ungünstigen Effekte unter den getesteten Bedingungen. Dennoch könnte die Ergänzung von Rationen mit besonders niedrigem Rohproteingehalt mit Methionin beim Einsatz an frischlaktierende Kühe hilfreich sein, um negative Langzeitwirkungen zu verhindern, die sich hier lediglich andeuteten. [source] FDB2 encodes a member of the arylamine N -acetyltransferase family and is necessary for biotransformation of benzoxazolinones by Fusarium verticillioidesJOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2009A.E. Glenn Abstract Aims:, To clone and characterize genes from the mycotoxigenic fungus, Fusarium verticillioides, which are associated with its ability to biotransform allelopathic benzoxazolinones produced by maize, wheat, and rye. Methods and Results:, Suppression subtractive hybridization identified F. verticillioides genes up-regulated in response to 2-benzoxazolinone (BOA), including a cluster of genes along chromosome 3. One of these genes, putatively encoding an arylamine N -acetyltransferase (NAT), was highly represented in the subtracted library and was of particular interest since previous analyses identified the FDB2 locus as possibly encoding transferase activity. The gene was subcloned and complemented a natural fdb2 mutant. Conversely, disruption of the gene eliminated the ability of F. verticillioides to metabolize BOA. Other genes in the cluster also were assessed using a complementation assay. Metabolic profiles of fdb2 mutants suggest that minor acylation activity occurred independently of the NAT activity encoded by FDB2. Conclusions:, The previously defined FDB2 locus was functionally associated with the gene encoding putative NAT activity, and the FDB2 gene was essential for biotransformation of BOA. The flanking gene FDB3 encodes a putative Zn(II)2Cys6 transcription factor and contributes to efficient BOA biotransformation but was not essential. Significance and Impact of the Study:, Biotransformation of benzoxazolinones by F. verticillioides may enhance its ecological fitness in maize field environments and our results provide greater understanding of the genes that modulate the biotransformation process. Additionally, this is the first homologue of the NAT gene family to be characterized in a filamentous fungus. [source] Gene expression of AGS cells stimulated with released proteins by Helicobacter pyloriJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 4 2008Nayoung Kim Abstract Background and Aim:, Interactions between released proteins by Helicobacter pylori (H. pylori) and the cells of gastric epithelium to which it adheres may contribute to gastric inflammation and epithelial damage. The present study was performed to evaluate the gene expression of AGS gastric cancer cells stimulated with released proteins by H. pylori. Methods:, Gene expression of AGS cells to the stimulation by H. pylori -released proteins (G27 strain) were monitored using oligonucleotide microarrays. Results:, Eighty-eight genes (0.88%) and eight genes (0.08%) were up- or downregulated, respectively, by treating AGS cells with H. pylori -released proteins but not by H. pylori adhesion after 12 h of coculture. Out of the selected 40 up- and five downregulated genes, 29 upregulated genes classified as general RNA polymerase II transcription factor activity (GTF2B, PPARGC1A), SH3/SH2 adaptor activity (CRKL), transferase activity (ACLY, CRKL, PIGC, PLK4), and oxidoreductase activity (IDH1) were confirmed to be upregulated by released proteins and not by H. pylori adhesion by real-time reverse transcription,polymerase chain reaction. When the concentrated H. pylori -cultured supernatant prepared by our protocol was treated by boiling, the upregulations of 26 of these 29 genes (89.7%) except for CD160, ZNF268, and PSAT1 disappeared. This confirmed that most of these upregulations were caused by released proteins. Conclusion:, Host genes involving transcription, signaling and stress are significantly modulated by the proteins released by H. pylori. This might strengthen the gastroduodenal pathogenesis induced by H. pylori. [source] Functional analysis of aromatic biosynthetic pathways in Pseudomonas putida KT2440MICROBIAL BIOTECHNOLOGY, Issue 1 2009M. Antonia Molina-Henares Summary Pseudomonas putida KT2440 is a non-pathogenic prototrophic bacterium with high potential for biotechnological applications. Despite all that is known about this strain, the biosynthesis of essential chemicals has not been fully analysed and auxotroph mutants are scarce. We carried out massive mini-Tn5 random mutagenesis and screened for auxotrophs that require aromatic amino acids. The biosynthesis of aromatic amino acids was analysed in detail including physical and transcriptional organization of genes, complementation assays and feeding experiments to establish pathway intermediates. There is a single pathway from chorismate leading to the biosynthesis of tryptophan, whereas the biosynthesis of phenylalanine and tyrosine is achieved through multiple convergent pathways. Genes for tryptophan biosynthesis are grouped in unlinked regions with the trpBA and trpGDE genes organized as operons and the trpI, trpE and trpF genes organized as single transcriptional units. The pheA and tyrA gene-encoding multifunctional enzymes for phenylalanine and tyrosine biosynthesis are linked in the chromosome and form an operon with the serC gene involved in serine biosynthesis. The last step in the biosynthesis of these two amino acids requires an amino transferase activity for which multiple tyrB -like genes are present in the host chromosome. [source] Thermoregulation of the Escherichia coli O157:H7 pO157 ecf operon and lipid A myristoyl transferase activity involves intrinsically curved DNAMOLECULAR MICROBIOLOGY, Issue 2 2004Jang W. Yoon Summary Escherichia coli O157:H7 survives in diverse environments from the ruminant gastrointestinal tract to cool nutrient-dilute water. We hypothesized that the gene regulation required for this flexibility includes intrinsically curved DNA that responds to environmental changes. Three intrinsically curved DNAs were cloned from the E. coli O157:H7 virulence plasmid (pO157), sequenced and designated Bent 1 through Bent 3 (BNT1, BNT2 and BNT3). Compared to BNT1 and BNT3, BNT2 had characteristics typical of intrinsically curved DNA including electrophoretic gel retardation at 4°C, six partially phased adenine:thymine tracts and transcriptional activation. BNT2::lacZ operon fusions showed that BNT2 activated transcription at 24°C compared to 37°C and was partially repressed by a bacterial nucleoid-associated protein H-NS. BNT2 regulated the E. coli attaching and effacing gene-positive conserved fragments 1,4 (ecf1,4) that are conserved in Shiga toxin-producing E. coli associated with human disease. Experimental analyses showed that ecf1,4 formed an operon. ecf1, 2 and 3 encoded putative proteins associated with bacterial surface polysaccharide biosynthesis and invasion and ecf4 complemented a chromosomal deletion of lpxM encoding lipid A myristoyl transferase. Mass spectrometric analysis of lipid A from ecf and lpxM single and double mutants showed that myristoylation was altered at lower temperature. [source] Developmental changes in glutathione S -transferase activity in herbicide-resistant populations of Alopecurus myosuroides Huds (black-grass) in the fieldPEST MANAGEMENT SCIENCE (FORMERLY: PESTICIDE SCIENCE), Issue 12 2001Lucy J Milner Abstract Herbicide-resistant populations of Alopecurus myosuroides Huds (black-grass) have become widespread throughout the UK since the early 1980s. Clear evidence suggests that more than one resistance mechanism exists, and glutathione S -transferases (GSTs) have been implicated in resistance due to enhanced metabolism. This study reports the determination of GST activity in four UK black-grass populations from field sites situated in the East Midlands. Data demonstrate that, as untreated plants in the field mature, there is an accompanying natural elevation of GST activity with natural environmental changes from winter to spring. We speculate that this endogenous change in enzyme activity with plant development in the field contributes to reduced efficacy of some graminicides applied in the spring. These observations are discussed in relation to predicting herbicide efficacy to achieve maximum control of this important grass weed. © 2001 Society of Chemical Industry [source] Resistance to ACCase-inhibiting herbicides and isoproturon in UK populations of Lolium multiflorum: mechanisms of resistance and implications for controlPEST MANAGEMENT SCIENCE (FORMERLY: PESTICIDE SCIENCE), Issue 7 2001Kay M Cocker Abstract Herbicide-resistant Lolium multiflorum (Italian rye-grass) was first reported in the UK in 1993 and had been confirmed on 25 farms by 1999. In this study, resistance to five herbicides belonging to the aryloxyphenoxypropionate, cyclohexanedione and phenyl-urea classes was determined in six populations of L multiflorum from the UK under glasshouse and simulated field conditions. Glasshouse conditions tended to exaggerate the degree of resistance, but experiments performed in both environments detected resistance in four populations of L multiflorum. Four populations (Essex A1, Lincs A1, Wilts B1, Yorks A2) were resistant to diclofop-methyl, fluazifop-P-butyl, tralkoxydim and partially resistant to isoproturon, but only the population from Yorkshire (Yorks A2) showed resistance to cycloxydim. Biochemical analyses of acetyl coenzyme A carboxylase (ACCase) activity, oxygen consumption by thylakoids, diclofop metabolism and glutathione S -transferase activity showed that, in three of the resistant populations, an enhanced rate of herbicide metabolism conferred resistance. This is the first report world-wide of an enhanced metabolism mechanism of diclofop resistance in L multiflorum. In the Yorks A2 population, an insensitive ACCase was detected (target-site resistance) which also conferred cross-resistance to all of the other ACCase inhibitors investigated. © 2001 Society of Chemical Industry [source] Antioxidant Defenses and DNA Damage Induced by UV-A and UV-B Radiation in the Crab Chasmagnathus granulata (Decapoda, Brachyura),PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 2 2005Glauce R. Gouveia ABSTRACT The photoprotector role of pigment dispersion in the melanophores of the crab, Chasmagnathus granulata, against DNA and oxidative damages caused by UV-A and UV-B was investigated. Intact and eyestalkaless crabs were used. In eyestalkless crabs, the dorsal epidermis of the cephalothorax (dispersed melanophores) and the epidermis of pereiopods (aggregated melanophores) were analyzed. Intact crabs showed only dispersed melanophores in the two epidermis. Antioxidant enzymes activity and lipoperoxidation content were analyzed after UV-A (2.5 J/cm2) or UV-B (8.6 J/sm2) irradiation. DNA damage was analyzed by single cell electrophoresis (comet) assay, after exposure to UV-B (8.6 J/cm2). UV-A radiation increased the glutatione- S -transferase activity in the pereiopods epidermis of eyestalkless crabs (P < 0.05). UV-B radiation induced DNA damage in the dorsal epidermis of eyestalkless crabs (P < 0.005). In pereiopod epidermis of eyestalkless crabs, there was no significant difference between control and UV-B-exposed crabs. In the pereiopods epidermis of eyestalkless, the control group showed higher scores of DNA damage and ,50% of cellular viability. Because in eyestalkless and irradiated crabs the cellular viability was ,5%, it was not possible to observe nuclei for determination of DNA damage. The findings show that melanophores can play a role in the defense against harmful effects of a momentary exposure to UV radiation. [source] Safety of meloxicam to critically endangered Gyps vultures and other scavenging birds in IndiaANIMAL CONSERVATION, Issue 2 2007D. Swarup Abstract Widespread veterinary use of the non-steroidal anti-inflammatory drug diclofenac is responsible for the population collapse of three species of Gyps vulture in south Asia; these species are now critically endangered. Vultures die when they consume carcasses of livestock that contain lethal residues of diclofenac. National and international conservation organizations have urgently recommended that diclofenac be banned and replaced with alternative drugs that are relatively safe to Gyps vultures and other scavenging birds. We tested the safety of the NSAID meloxicam on the oriental white-backed vulture, long-billed vulture and a range of other scavenging birds in India (Egyptian vulture Neophron percnopterus, cattle egret Bubulcus ibis, house crow Corvus splendens, large-billed crow Corvus machrorhynchos and common mynah Acridotheres tristis). Meloxicam was administered by oral intubation [at 0.5 and 2.0 mg kg,1 vulture body weight (bw)], or through feeding with muscle or liver tissue (at 0.3 to 2.1 mg kg,1 vulture bw) from meloxicam-treated buffalo Bubalus bubalis. We estimate that 2.0 mg kg,1 bw is the maximum likely exposure in the wild. All 31 Gyps vultures and the 20 other scavenging birds given meloxicam survived. Feeding behaviour remained normal and there were no significant differences between the treated and control groups in body mass, or the blood haematology and biochemistry parameters monitored, including those known to be affected by diclofenac (uric acid levels and alanine transferase activity). Meloxicam is used to treat a wide range of livestock ailments and is licensed and manufactured in India. We recommend that meloxicam be introduced as rapidly as possible across the Indian sub-continent as an alternative to diclofenac. [source] Isolation of a porcine UDP-GalNAc transferase cDNA mapping to the region of the blood group EAA locus on pig chromosome 1ANIMAL GENETICS, Issue 3 2001E. Meijerink In our studies of the genes constituting the porcine A0 blood group system, we have characterized a cDNA, encoding an ,(1,3)N-acetylgalactosaminyltransferase, that putatively represents the blood group A transferase gene. The cDNA has a 1095-bp open reading frame and shares 76.9% nucleotide and 66.7% amino acid identity with the human ABO gene. Using a somatic cell hybrid panel, the cDNA was assigned to the q arm of pig chromosome 1, in the region of the erythrocyte antigen A locus (EAA), which represents the porcine blood group A transferase gene. The RNA corresponding to our cDNA was expressed in the small intestinal mucosae of pigs possessing EAA activity, whereas expression was absent in animals lacking this blood group antigen. The UDP-N-acetylgalactosamine (UDP-GalNAc) transferase activity of the gene product, expressed in Chinese hamster ovary (CHO) cells, was specific for the acceptor fucosyl- ,(1,2)galactopyranoside; the enzyme did not use phenyl- , - D -galactopyranoside (phenyl- , -D-Gal) as an acceptor. Because the ,(1,3)GalNAc transferase gene product requires an ,(1,2)fucosylated acceptor for UDP-GalNAc transferase activity, the ,(1,2)fucosyltransferase gene product is necessary for the functioning of the ,(1,3)GalNAc transferase gene product. This mechanism underlies the epistatic effect of the porcine S locus on expression of the blood group A antigen. Abbreviations: CDS: coding sequence; CHO: Chinese Hamster Ovary; EAA: erythrocyte antigen A; FCS: foetal calf serum; Fuc,(1,2)Gal: fucosyl- ,(1,2)galactopyranoside; Gal: galactopyranoside; GGTA1: Gal,(1,3)Gal transferase; PCR: polymerase chain reaction; phenyl- , -D-Gal: phenyl- , - D -galactopyranoside; R: Gal,1-4Glc,1-1Cer; UDP-GalNAc: uridine diphosphate N-acetylgalactosamine [source] Simultaneous analysis of catechol- O -methyl transferase activity, S -adenosylhomocysteine and adenosineBIOMEDICAL CHROMATOGRAPHY, Issue 3 2010Ilkka Reenilä Abstract Novel HPLC method utilizing UV-detection was developed to analyse catechol- O -methyltransferase (COMT) products, vanillic acid and isovanillic acid, S -adenosylhomocysteine (SAH) and adenosine formed from dihydroxybenzoic acid and S -adenosyl-L-methionine (SAM) by incubation of the rat tissues. Entacapone, a COMT inhibitor, prevented the formation of SAH only partially in the striatal homogenate whereas in the kidney homogenate the increase of SAH was prevented by entacapone. In conclusion, this method was reliable, rapid and simple. COMT seemed to be partially responsible on the SAM utilizing methylations in the striatal homogenates while in the high COMT activity tissue, COMT was the main SAH producing methyltransferase. Copyright © 2009 John Wiley & Sons, Ltd. [source] Tyrosine aminotransferase and gamma-glutamyl transferase activity in human fetal hepatocyte primary cultures under proliferative conditionsCELL BIOCHEMISTRY AND FUNCTION, Issue 2 2004Khaja K. Rehman Abstract The ontogeny of gamma-glutamyl transferase (GGTase; E.C.2.3.2.2) and tyrosine aminotransferase (TAT; E.C.2.6.1.5) activities in 14 to 36 weeks gestational and neonatal hepatocytes during development of human fetal liver was studied. Subsequently, 20,24 weeks gestational hepatocytes were cultured in media supplemented with epidermal growth factor (EGF) and insulin with or without glucagon and dexamethasone to investigate the proliferation and differentiation of fetal hepatocyte in vitro using GGTase and TAT as biochemical markers. During the development of the liver, the activity of GGTase increased continuously from the first trimester through the third trimester and decreased (p,<,0.001) in neonates. A low basal level of TAT activity was seen only during the third trimester, which then increased significantly (p,<,0.001) in neonates. Fetal hepatocytes, in the presence of EGF and insulin, undergo proliferation from the fourth to 10th day with an increase in cell number (p,<,0.001) and concomitant increase (p,<,0.001) in GGTase activity. As the cells attain confluence, enzyme activity decreased significantly (p,<,0.001) from the 10th to 16th day. Maximal TAT activity (p,<,0.001) was observed at 48,h of culture, which decreased, but not significantly, during cell proliferation and the enzyme activity was regained as the cultures attained confluence. Furthermore, TAT activity was induced synergistically (p<0.001) in the presence of glucagon and dexamethasone, while GGTase was inhibited (p<0.001). These results indicate that GGTase increases with proliferation, whereas TAT, once it has been expressed, is not suppressed during cell proliferation. In conclusion, human fetal hepatocytes undergo enzymic differentiation by 48,h of culture, and proliferate with an increase in GGTase in the presence of growth factors with maintenance of differentiated status up to the studied 16 days of culture. Copyright © 2003 John Wiley & Sons, Ltd. [source] | ||||||||||||||||