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Enzyme Activity (enzyme + activity)
Kinds of Enzyme Activity Terms modified by Enzyme Activity Selected AbstractsEFFECTS OF RAW MATERIALS AND PROCESS VARIABLES ON THE HEAT PENETRATION TIMES, FIRMNESS, AND PECTIC ENZYME ACTIVITY OF DICED TOMATOES (HALLEY BOS 3155 CV)JOURNAL OF FOOD PROCESSING AND PRESERVATION, Issue 2 2001WENDY H. MA The effects of raw materials and process variables on the heat penetration times into diced tomatoes (Halley Bos 3155 cv) were evaluated. Variables included dice size (1.27 and 2.54 cm), maturity at harvest (red and red+2 weeks), and processing temperature (88 and 92C). Heat penetration times between dice sizes were significantly different, but not between maturities or processing temperatures. Tomatoes were also evaluated for firmness, pectin-methylesterase (PME) and polygalacturonase (PG) activities. Half-inch size diced tomatoes were processed at 88 and 92C, and evaluated for firmness using the shear-compression method. Firmness decreased to 60% of the initial raw firmness from 8.8 × 105 to 5.3 × 105 g-mm after 15 s at 88C, and to 50% from 8.8 × 105 to 4.4 × 105 g-mm after 15 s at 92C. Diced tomato firmness showed a slight firming trend after 150 s at both temperatures. PME was inactivated after 45 s, while 5% residual PG activity remained after 3 min. [source] RESPONSE OF GLUTAMINE SYNTHETASE GENE TRANSCRIPTION AND ENZYME ACTIVITY TO EXTERNAL NITROGEN SOURCES IN THE DIATOM SKELETONEMA COSTATUM (BACILLARIOPHYCEAE),JOURNAL OF PHYCOLOGY, Issue 1 2005Misaki Takabayashi To understand the enhanced ability of marine diatoms to assimilate nitrogen (N), we measured changes in the transcript abundance and enzyme activity of glutamine synthetase (GS), one of the key enzymes that link carbon (C) and N metabolism, in the common diatom Skeletonema costatum (Greville) Cleve. Transcript abundance of glnII (the gene that encodes the GSII isoenzyme), measured by quantitative reverse transcriptase-PCR, and total GS activity increased 2 to 3.5 times above background in the cells taking up nitrate (NO3,) but not the cells taking up ammonium (NH4+). A background level of glnII mRNA was maintained at a steady level up to 15 days of N starvation before decreasing to below detection after 21 days. These results confirm that transcription of glnII is induced to assimilate NH4+ derived from reduction of NO3,. Because of this role of GSII in diatoms assimilating NH4+ derived from NO3, reduction rather than from the environmental NH4+, quantification of glnII mRNA promises to be a useful indication of new production by phytoplankton. [source] Nitrogen Rates and Water Stress Effects on Production, Lipid Peroxidation and Antioxidative Enzyme Activities in Two Maize (Zea mays L.) GenotypesJOURNAL OF AGRONOMY AND CROP SCIENCE, Issue 6 2007L.-X. Zhang Abstract Effects of nitrogen rates and water stress (WS) on production, lipid peroxidation and antioxidative enzyme activities in two maize (Zea mays L.) genotypes were assessed at different stages under two levels of water supply conditions. WS caused a significant decline in dry matter, grain yield and activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) whereas a marked rise in malondialdehyde (MDA) concentration was observed in leaves for the two genotypes. However, the responses of the two varieties to WS were different: significantly higher dry matter, grain yield and antioxidative enzyme activities and lower MDA content were observed for Shaandan 9 than Shaandan 911, therefore the former could be treated as a drought tolerance variety comparatively. A better correlation was obtained amongst dry matter, grain yield and physiological traits. The addition of nitrogen increased dry matter and grain yield as well as activities of SOD, POD and CAT to different levels and significantly decreased MDA content under WS. These effects were higher for Shaandan 911 than for Shaandan 9. Furthermore, a significant effect was found for Shaandan 911 between N rates for all traits unlike Shaandan 9. Hence, we suggest that nitrogen should be applied to a water-sensitive variety to bring out its potential fully under drought. [source] Characterization of Cell Wall Enzyme Activities, Pectin Composition, and Technological Criteria of Strawberry Cultivars (Fragaria×ananassa Duch)JOURNAL OF FOOD SCIENCE, Issue 4 2004G. Lefever ABSTRACT: The effects of physical characteristics and cell wall enzymatic activities of several strawberry culti-vars were investigated for possible industrial use. The enzymes study showed that the softest varieties had the highest pectin methylesterase (PME) and polygalacturonase (PG) activities. Differences in alcohol-insoluble pectin, water-soluble pectin, and parietal residue compositions were observed between Darsanga ("firm fruit") and Senga sengana ("soft fruit"). Finally, the study of pectin composition of Darsanga and Senga sengana indicated that the softest fruit had the highest water-soluble pectin content. The measurement of fruit PME activity permitted a preliminary screening of fruit maturity characteristics. [source] Effect of Filtration Leukocytapheresis Therapy: Modulation of White Blood Cell Enzyme Activities in Patients with Rheumatoid ArthritisARTIFICIAL ORGANS, Issue 4 2002Satoshi Yamasaki Abstract: We treated 12 patients with rheumatoid arthritis by filtration leukocytapheresis (FLCP) and evaluated its effect on leukocyte enzyme activities. We calculated the number of leukocytes removed and assessed the clinical response. We also evaluated the cellular enzyme activities of elastase and dipeptidylpeptidase IV (DPP IV). Out of 12 patients, 7 patients achieved 20% improvement for 4 weeks following FLCP. The FLCP treatment resulted in removal of 96% of granulocytes, 98% of monocytes, and 61% of lymphocytes. Granulocytes and monocytes with high elastase activity were effectively removed by FLCP. The elastase activity of granulocytes was increased 4 weeks after the last FLCP only in responders. On the other hand, the DPP IV activity of lymphocytes was low at 4 weeks after the last FLCP in responders. Modulation of leukocyte enzyme activities is one of the main effects of FLCP therapy and alteration of granulocytes, monocytes, and lymphocytes. [source] Extracellular Enzyme Activities and Carbon Chemistry as Drivers of Tropical Plant Litter DecompositionBIOTROPICA, Issue 3 2004Steven D. Allison ABSTRACT Litter quality parameters such as nitrogen and lignin content correlate with decomposition rates at coarse scales, but fine-scale mechanisms driving litter decomposition have proven more difficult to generalize. One potentially important driver of decomposition is the activity of extracellular enzymes that catalyze the degradation of complex compounds present in litter. To address the importance of this mechanism, we collected 15 Hawaiian plant litter types and decomposed them in fertilized and control plots for up to two years. We measured litter nutrient content and carbon chemistry prior to decomposition, as well as extracellular enzyme activities, mass loss, and litter nutrient content over time. We found that water-soluble carbon content, cellobiohydrolase activities, and polyphenol oxidase activities were significantly correlated with mass loss. Enzyme activities and decomposition rate constants both varied significantly by litter type, and fertilization increased mass loss rates in five litter types. Some litter types that decayed faster under fertilization also showed time-dependent increases in carbon-degrading enzyme activities, but others decayed faster independent of enzyme changes. These results suggest that extracellular enzyme activities partially determine litter decomposition rates, but high soluble carbon content may circumvent the requirement for enzyme-catalyzed decomposition. [source] Inhibition of Biochemical Reactions by Silicon Nanowires through Modulating Enzyme ActivitiesCHEMBIOCHEM, Issue 11 2007Changqing Yi Through the wire. We have investigated the potential effects of silicon nanowires (SiNW-SiO2) and SiNWs functionalized with carboxylic groups (SiNW-COOH) on restriction endonucleases and Taq DNA polymerase. The results show that these SiNWs can inhibit enzyme activity (lower band in gel). Our findings suggest that this could be due to chemical interactions between the functional groups on SiNWs and the enzymes. [source] Regulating Enzyme Activities in a Multiple-Enzyme ComplexCHEMBIOCHEM, Issue 6 2005Yi Chen Off and on. This paper reports a general strategy (strand displacement) to isothermally, individually, and reversibly regulate each DNA enzyme in a multiple-enzyme complex. The graph shows the time course of the codigestion of two substrate strands (Sa and Sb) with the sequential addition of inhibitor (I) and remover (R) strands. [source] Selective Long-Term Electrical Stimulation of Fast Glycolytic Fibres Increases Capillary Supply but not Oxidative Enzyme Activity in Rat Skeletal MusclesEXPERIMENTAL PHYSIOLOGY, Issue 5 2000S. Egginton Glycolytic fibres in rat extensor digitorum longus (EDL) and tibialis anterior (TA) were selectively activated, as demonstrated by glycogen depletion, by indirect electrical stimulation via electrodes implanted in the vicinity of the peroneal nerve using high frequency (40 Hz) trains (250 ms at 1 Hz) and low voltage (threshold of palpable contractions). This regime was applied 10 times per day, each bout being of 15 min duration with 60 min recovery, for 2 weeks. Cryostat sections of muscles were stained for alkaline phosphatase to depict capillaries, succinate dehydrogenase (SDH) to demonstrate oxidative fibres, and periodic acid-Schiff reagent (PAS) to verify glycogen depletion. Specific activity of hexokinase (HK), 6-phosphofructokinase, pyruvate kinase, glycogen phosphorylase and cytochrome c oxidase (COX) were estimated separately in homogenates of the EDL and the predominantly glycolytic cortex and oxidative core of the TA. Stimulation increased the activity of HK but not that of oxidative enzymes in fast muscles. Comparison of changes in oxidative capacity and capillary supply showed a dissociation in the predominantly glycolytic TA cortex. Here, COX was 3.9 ± 0.68 ,M min-1 (g wet wt)-1 in stimulated muscles compared with 3.7 ± 0.52 ,M min-1 (g wet wt)-1 in contralateral muscles (difference not significant), while the percentage of oxidative fibres (those positively stained for SDH) was also similar in stimulated (14.0 ± 2.8%) and contralateral (12.2 ± 1.9%) muscles. In contrast, the capillary to fibre ratio was significantly increased (2.01 ± 0.12 vs. 1.55 ± 0.04, P < 0.01). We conclude that capillary supply can be increased independently of oxidative capacity, possibly due to haemodynamic factors, and serves metabolite removal to a greater extent than substrate delivery. [source] Selected Line Difference in the Effects of Ethanol Dependence and Withdrawal on Allopregnanolone Levels and 5,-Reductase Enzyme Activity and ExpressionALCOHOLISM, Issue 12 2009Michelle A. Tanchuck Background:, Allopregnanolone (ALLO) is a progesterone derivative that rapidly potentiates ,-aminobutyric acidA (GABAA) receptor-mediated inhibition and modulates symptoms of ethanol withdrawal. Because clinical and preclinical data indicate that ALLO levels are inversely related to symptoms of withdrawal, the present studies determined whether ethanol dependence and withdrawal differentially altered plasma and cortical ALLO levels in mice selectively bred for differences in ethanol withdrawal severity and determined whether the alterations in ALLO levels corresponded to a concomitant change in activity and expression of the biosynthetic enzyme 5,-reductase. Methods:, Male Withdrawal Seizure-Prone (WSP) and -Resistant (WSR) mice were exposed to 72 hours ethanol vapor or air and euthanized at select times following removal from the inhalation chambers. Blood was collected for analysis of ALLO and corticosterone levels by radioimmunoassay. Dissected amygdala, hippocampus, midbrain, and cortex as well as adrenals were examined for 5,-reductase enzyme activity and expression levels. Results:, Plasma ALLO was decreased significantly only in WSP mice, and this corresponded to a decrease in adrenal 5,-reductase expression. Cortical ALLO was decreased up to 54% in WSP mice and up to 46% in WSR mice, with a similar decrease in cortical 5,-reductase activity during withdrawal in the lines. While cortical gene expression was significantly decreased during withdrawal in WSP mice, there was a 4-fold increase in expression in the WSR line during withdrawal. Hippocampal 5,-reductase activity and gene expression was decreased only in dependent WSP mice. Conclusions:, These results suggest that there are line and brain regional differences in the regulation of the neurosteroid biosynthetic enzyme 5,-reductase during ethanol dependence and withdrawal. In conjunction with the finding that WSP mice exhibit reduced sensitivity to ALLO during withdrawal, the present results are consistent with the hypothesis that genetic differences in ethanol withdrawal severity are due, in part, to modulatory effects of GABAergic neurosteroids such as ALLO. [source] Nano-Encapsulation of Lipase by Self-Assembled Nanogels: Induction of High Enzyme Activity and Thermal StabilizationMACROMOLECULAR BIOSCIENCE, Issue 4 2010Shin-ichi Sawada Abstract The effects of self-assembled polysaccharide nanogels on colloidal and thermal stability of lipase from Pseudomonas cepacia were investigated. The enzyme activity, especially kcat, drastically increased in the presence of nanogels of cholesterol-bearing pullulan (CHP). The thermostability of lipase complex increased because the denaturation temperature of lipase increased by more than 20,°C by complexation with CHP nanogels. Lipase denaturation and aggregation upon heating was effectively prevented by complexation with CHP nanogels. Moreover, complexation with CHP nanogels protected lipase from lyophilization-induced aggregation. Nano-encapsulation with CHP nanogel is a useful method for colloidal and thermal stabilization of unstable enzyme. [source] Effects of Cryopreservation and Hypothermic Storage on Cell Viability and Enzyme Activity in Recombinant Encapsulated Cells Overexpressing Alpha-L-IduronidaseARTIFICIAL ORGANS, Issue 5 2010Fabiana Quoos Mayer Abstract Here, we show the effects of cryopreservation and hypothermic storage upon cell viability and enzyme release in alginate beads containing baby hamster kidney cells overexpressing alpha-L-iduronidase (IDUA), the enzyme deficient in mucopolysaccharidosis type I. In addition, we compared two different concentrations of alginate gel (1% and 1.5%) in respect to enzyme release from the beads and their shape and integrity. Our results indicate that in both alginate concentrations, the enzyme is released in lower amounts compared with nonencapsulated cells. Alginate 1% beads presented increased levels of IDUA release, although this group presented more deformities when compared with alginate 1.5% beads. Importantly, both encapsulated groups presented higher cell viability after long cryopreservation period and hypothermic storage. In addition, alginate 1.5% beads presented higher enzyme release after freezing protocols. Taken together, our findings suggest a benefic effect of alginate upon cell viability and functionality. These results may have important application for treatment of both genetic and nongenetic diseases using microencapsulation-based artificial organs. [source] A Mass Spectrometry Plate Reader: Monitoring Enzyme Activity and Inhibition with a Desorption/Ionization on Silicon (DIOS) PlatformCHEMBIOCHEM, Issue 7 2004Zhouxin Shen Dr. Abstract A surface-based laser desorption/ionization mass spectrometry assay that makes use of Desorption/Ionization on Silicon Mass Spectrometry (DIOS-MS) has been developed to monitor enzyme activity and enzyme inhibition. DIOS-MS has been used to characterize inhibitors from a library and then to monitor their activity against selected enzyme targets, including proteases, glycotransferase, and acetylcholinesterase. An automated DIOS-MS system was also used as a high-throughput screen for the activity of novel enzymes and enzyme inhibitors. On two different commercially available instruments, a sampling rate of up to 38 inhibitors per minute was accomplished, with thousands of inhibitors being monitored. The ease of applying mass spectrometry toward developing enzyme assays and the speed of surface-based assays such as DIOS for monitoring inhibitor effectiveness and enzyme activity makes it attractive for a broad range of screening applications. [source] Brain aromatase, 5,-reductase, and 5,-reductase change seasonally in wild male song sparrows: Relationship to aggressive and sexual behaviorDEVELOPMENTAL NEUROBIOLOGY, Issue 3 2003Kiran K. Soma Abstract In many species, territoriality is expressed only during the breeding season, when plasma testosterone (T) is elevated. In contrast, in song sparrows (Melospiza melodia morphna), males are highly territorial during the breeding (spring) and nonbreeding (autumn) seasons, but not during molt (late summer). In autumn, plasma sex steroids are basal, and castration has no effect on aggression. However, inhibition of aromatase reduces nonbreeding aggression, suggesting that neural steroid metabolism may regulate aggressive behavior. In wild male song sparrows, we examined the neural distribution of aromatase mRNA and seasonal changes in the activities of aromatase, 5,-, and 5,-reductase, enzymes that convert T to 17,-estradiol, 5,-dihydrotestosterone (5,-DHT, a potent androgen), or 5,-DHT (an inactive metabolite), respectively. Enzyme activities were measured in the diencephalon, ventromedial telencephalon (vmTEL, which includes avian amygdala), caudomedial neostriatum (NCM), and the hippocampus of birds captured during spring, molt, or autumn. Aromatase and 5,-reductase changed seasonally in a region-specific manner. Aromatase in the diencephalon was higher in spring than in molt and autumn, similar to seasonal changes in male sexual behavior. Aromatase activity in the vmTEL was high in both spring and autumn but significantly reduced at molt, similar to seasonal changes in aggression. 5,-Reductase was not elevated during molt, suggesting that low aggression during molt is not a result of increased inactivation of androgens. These data highlight the relevance of neural steroid metabolism to the expression of natural behaviors by free-living animals. © 2003 Wiley Periodicals, Inc. J Neurobiol 56: 209,221, 2003 [source] Prediction of polymorphic N -acetylation of new drug candidates by correlation with human NAT1 and NAT2DRUG DEVELOPMENT RESEARCH, Issue 1 2002Katalin Jemnitz Abstract Due to interindividual variation in N -acetyltransferase 2 (NAT2) activity, pharmaceutical companies face the problem of polymorphic metabolism in drugs that are metabolized mainly or exclusively by this enzyme. An in vitro method has been developed to predict in vivo polymorphic N -acetylation at an early stage of drug development. Two new type 5H-2,3-benzodiazepine derivatives, Nerisopam (NER) with anxiolytic activity and GYKI47261 with antiepileptic activity, are metabolized mainly by N -acetylation in the rat and human. The selectivity of human N -acetyltransferases (NAT1,2) to form the acetylated metabolites has been investigated by correlation analysis. Twelve human liver samples were characterized for NAT1 and NAT2 phenotype based on their enzyme activity toward two selective NAT1 (p -aminobenzoic acid, PABA; p -aminosalicylic acid, PAS) and two selective NAT2 (sulfamethazine, SMZ; procainamide, PROC) substrates. Significant correlation was found between enzyme activities NAT1PABA/NAT1PAS and NAT2SMZ/NAT2PROC, respectively, and no correlation was observed comparing enzyme activities toward NAT1PABA/NAT2PROC. Enzyme activities using NER and GYKI 47261 as substrates were compared to activities obtained with NAT1 and NAT2 selective substrates, and the correlation coefficients were calculated. Good correlation was established between the rates of acetylation of the two drugs and that of the NAT2 selective substrate (NER/NAT2SMZ, r2=0.91, GYKI 47261/NAT2SMZ, r2=0.91). In contrast, no correlation was found between the rate of conjugation of the drugs and that of NAT1 selective substrate (NER/NAT1PABA, r2=0.022, GYKI 47261/NAT1PABA, r2=0.0004), suggesting polymorphic in vivo metabolism, since both drugs are acetylated preferably by NAT2. According to our results, correlation analysis based on in vitro acetylation activity may be used to predict in vivo polymorphic metabolism. Drug Dev. Res. 56:17,22, 2002. © 2002 Wiley-Liss, Inc. [source] Hydrocarbon-induced changes to metabolic and detoxification enzymes of the Australian crimson-spotted rainbowfish (Melanotaenia fluviatilis)ENVIRONMENTAL TOXICOLOGY, Issue 1 2003Carmel A. Pollino Abstract The toxicity of petroleum hydrocarbons to marine aquatic organisms has been widely investigated; however, the effects on freshwater environments have largely been ignored. Selected biomarkers were measured in a freshwater species, the crimson-spotted rainbowfish (Melanotaenia fluviatilis). Fish were exposed to either a water-accommodated fraction (WAF) of crude oil or a dispersed crude oil water-accommodated fraction (DCWAF) for 3 days and were depurated for 14 days. Generally, biomarkers were altered following the short-term exposures but recovered after 14 days of depuration. Metabolic enzymes measured in gill tissue were citrate synthase and lactate dehydrogenase (LDH). As a result of WAF and DCWAF exposures, citrate synthase and LDH activities increased. Enzyme activities returned to control levels following depuration. Subsequent to the WAF exposure, hepatic ethoxyresorufin- O -deethylase (EROD) activity levels were higher than controls and they returned to control levels during depuration. For the DCWAF exposure, EROD was induced by a TPH (total petroleum hydrocarbons) concentration of 14.5 mg/L; however, after depuration the 14.5 mg/L TPH group had lower EROD activity than did controls. There were no changes in liver- to body-weight ratios or the histopathological organization of gill or liver tissues. As the majority of biomarkers returned to control levels after 14 days of depuration, rainbowfish were able to recover from short-term exposures to crude oil and dispersed crude oil. © 2003 Wiley Periodicals, Inc. Environ Toxicol 18: 21,28, 2003. [source] Glycolysis in Ustilago maydisFEMS YEAST RESEARCH, Issue 8 2008Emma Saavedra Abstract The kinetic parameters of the 10 glycolytic enzymes and glycolytic fluxes were determined for the first time in Ustilago maydis. Enzyme activities in yeast grown in minimal medium and harvested in the stationary stage were twofold higher than those from yeast grown in rich medium. In contrast, in yeast harvested in the exponential stage, the enzyme activities were higher in cells grown in rich medium. Phosphofructokinase activity was the lowest in the four culture conditions analyzed, suggesting that this enzyme is a flux-controlling step in U. maydis glycolysis. The Vmax and Km values of hexokinase and pyruvate kinase were similar under all conditions. The results revealed that U. maydis aldolase belongs to the class II type of metalo-aldolases. 3-Phosphoglycerate mutase (PGAM) activity was 2,3-bisphosphoglycerate cofactor independent, which contrasted with the cofactor dependency predicted by the amino acid sequence alignment analysis. Pyruvate was secreted by U. maydis yeast in the presence and absence of external glucose. The glycolytic enzyme activities in the U. maydis mycelial form were similar to those found in yeast, except for one order of magnitude higher phosphofructokinase and PGAM activities, thus suggesting differences in the glycolysis regulatory mechanisms between the two cellular forms. [source] Specificities of proteases for use in leather manufactureJOURNAL OF CHEMICAL TECHNOLOGY & BIOTECHNOLOGY, Issue 3 2006Farhad Foroughi Abstract Proteases are used in leather manufacture in the processes of soaking, unhairing and bating of hides and skins. However proteases can be relatively non-specific in their usage, and for improved efficacy of enzyme biocatalysis within the industry, an analysis of specific activities of enzymes towards skin proteins was undertaken. Most commercial proteases for soaking showed substantial activity against the substrates elastin,Congo Red and Azocoll but little or no activity against keratin,azure and hide powder black. Enzymes used for unhairing in conjunction with 30% of the usual concentration of sulfide to effect chemical unhairing showed moderate activity against all substrates tested (selected as representative of skin proteins), while proteases used in bating showed activity against Azocoll and elastin,Congo Red but had no keratinase activity and little activity against hide powder black. Bating proteases and soaking proteases displayed similar activities at pH 8. Microbes isolated in the screening of organisms from putrefied skins included one fungal and two bacterial isolates whose extracellular enzymes had efficient unhairing activity without the addition of sulfide. Enzyme activities for these proteases included high activity measured against Azocoll with little or no activity against elastin,Congo Red, keratin,azure and hide powder black. Neither elastase nor keratinase activities were determined as being essential for unhairing. Copyright © 2005 Society of Chemical Industry [source] Mitochondrial function and apoptotic susceptibility in aging skeletal muscleAGING CELL, Issue 1 2008Béatrice Chabi Summary During aging, skeletal muscle undergoes sarcopenia, a condition characterized by a loss of muscle cell mass and alterations in contractile function. The origin of these decrements is unknown, but evidence suggests that they can be partly attributed to mitochondrial dysfunction. To characterize the nature of this dysfunction, we investigated skeletal muscle contractile properties, subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondrial biogenesis and function, as well as apoptotic susceptibility in young (6 months old) and senescent (36 months old) Fischer 344 Brown Norway rats. Muscle mass and maximal force production were significantly lower in the 36-month group, which is indicative of a sarcopenic phenotype. Furthermore, contractile activity in situ revealed greater fatigability in the 36-month compared to the 6-month animals. This decrement could be partially accounted for by a 30% lower mitochondrial content in fast-twitch muscle from 36-month animals, as well as lower protein levels of the transcriptional coactivator peroxisome proliferator-activated receptor , coactivator-1,. Enzyme activities and glutamate-induced oxygen consumption rates in isolated SS and IMF mitochondria were similar between age groups. However, mitochondrial reactive oxygen species (ROS) production during state 3 respiration was ~1.7-fold greater in mitochondria isolated from 36-month compared to 6-month animals, and was accompanied by a 1.8-fold increase in the DNA repair enzyme 8-oxoguanine glycosylase 1 in fast-twitch muscle. Basal rates of release of cytochrome c and endonuclease G in SS mitochondria were 3.5- to 7-fold higher from senescent animals. These data suggest that the age-related sarcopenia and muscle fatigability are associated with enhanced ROS production, increased mitochondrial apoptotic susceptibility and reduced transcriptional drive for mitochondrial biogenesis. [source] A Novel Process for the Recovery of Polyphenols from Grape (Vitis vinifera L.) PomaceJOURNAL OF FOOD SCIENCE, Issue 2 2005Dietmar Kammerer ABSTRACT: A novel process for enzyme-assisted extraction of polyphenols from winery by-products was established on a pilot-plant scale. Optimization of enzymatic hydrolysis of grape skins, that is, selection of pectinolytic and cellulolytic enzymes, enzyme-substrate ratio, and time-temperature regime of enzymatic treatment, was conducted on a laboratory scale. Enzyme activities were monitored by viscosity measurement of resuspended grape pomace and by quantification of oligomeric pectin and cellulose degradation products released from cell wall material. Optimal conditions were obtained with 5000 ppm (based on dry matter) of a pectinolytic and 2500 ppm of a cellulolytic enzyme preparation, respectively, at 50°C, which were also applied in pilot-plant scale experiments. Concomitant determination of individual polyphenolics demonstrated a significantly improved yield for most compounds when compared with experiments without enzyme addition. Recovery rates were comparable to those obtained when grape pomace was extracted using sulfite. Pre-extraction of the pomace with hot water followed by treatment with cell wall degrading enzymes even increased yields of phenolic compounds. Only some quercetin glycosides and malvidin coumaroylglucoside were partly hydrolyzed due to enzyme side activities. This new process may provide a valuable alternative to the application of sulfite, which is considered crucial in food processing. [source] Induction of resistance in cocoa against Crinipellis perniciosa and Verticillium dahliae by acibenzolar- S -methyl (ASM)PLANT PATHOLOGY, Issue 5 2002M. L. V. Resende The benzothiadiazole compound acibenzolar- S -methyl (ASM) was assessed as an inducer of resistance against Crinipellis perniciosa, agent of witches' broom, and Verticillium dahliae, agent of vascular wilt, both on cocoa. ASM induced a reduction in incidence of witches' broom of up to 84·5% when sprayed 30 days before inoculation on cocoa seedlings of cv. Catongo. ASM also induced a reduction in severity of Verticillium wilt to 55·4% on cv. Theobahia. For both pathosystems, effects of dose on disease were not clearly observed. The efficacy of the inducer increased with the interval between sprayings and the respective inoculations with the pathogens. In another experiment, the effect of ASM on the control of witches' broom on cocoa seedlings was compared with that of cuprous oxide and tebuconazole, all sprayed 15 days before inoculation. ASM reduced disease incidence by 60·1% compared with the inoculated control. ASM was superior to tebuconazole, and there was also a tendency for ASM to be better than cuprous oxide. To understand the mechanism of action of ASM as an inducer of resistance, alterations in the levels of total phenolics, polyphenol oxidases and peroxidases were evaluated 3, 15 and 30 days after spraying of seedlings of cv. Catongo. Enzyme activities from seedlings of cv. Theobahia were evaluated 30 days after spraying. On cv. Catongo, no significant differences in total phenolic content and polyphenol oxidase activity were detected after spraying. However, an increase in peroxidase activity was detected at all times of evaluation. On cv. Theobahia, significant increases in activities of peroxidase and polyphenol oxidase were detected, indicating that defence responses due to ASM were dependent on host genotype. [source] Structure and function of GlmU from Mycobacterium tuberculosisACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2009Zhening Zhang Antibiotic resistance is a major issue in the treatment of infectious diseases such as tuberculosis. Existing antibiotics target only a few cellular pathways and there is an urgent need for antibiotics that have novel molecular mechanisms. The glmU gene is essential in Mycobacterium tuberculosis, being required for optimal bacterial growth, and has been selected as a possible drug target for structural and functional investigation. GlmU is a bifunctional acetyltransferase/uridyltransferase that catalyses the formation of UDP-GlcNAc from GlcN-1-P. UDP-GlcNAc is a substrate for two important biosynthetic pathways: lipopolysaccharide and peptidoglycan synthesis. The crystal structure of M. tuberculosis GlmU has been determined in an unliganded form and in complex with GlcNAc-1-P or UDP-GlcNAc. The structures reveal the residues that are responsible for substrate binding. Enzyme activities were characterized by 1H NMR and suggest that the presence of acetyl-coenzyme A has an inhibitory effect on uridyltransferase activity. [source] Extracellular Enzyme Activities and Carbon Chemistry as Drivers of Tropical Plant Litter DecompositionBIOTROPICA, Issue 3 2004Steven D. Allison ABSTRACT Litter quality parameters such as nitrogen and lignin content correlate with decomposition rates at coarse scales, but fine-scale mechanisms driving litter decomposition have proven more difficult to generalize. One potentially important driver of decomposition is the activity of extracellular enzymes that catalyze the degradation of complex compounds present in litter. To address the importance of this mechanism, we collected 15 Hawaiian plant litter types and decomposed them in fertilized and control plots for up to two years. We measured litter nutrient content and carbon chemistry prior to decomposition, as well as extracellular enzyme activities, mass loss, and litter nutrient content over time. We found that water-soluble carbon content, cellobiohydrolase activities, and polyphenol oxidase activities were significantly correlated with mass loss. Enzyme activities and decomposition rate constants both varied significantly by litter type, and fertilization increased mass loss rates in five litter types. Some litter types that decayed faster under fertilization also showed time-dependent increases in carbon-degrading enzyme activities, but others decayed faster independent of enzyme changes. These results suggest that extracellular enzyme activities partially determine litter decomposition rates, but high soluble carbon content may circumvent the requirement for enzyme-catalyzed decomposition. [source] Diabetes-induced decrease in rat brain microsomal Ca2+ -ATPase activityCELL BIOCHEMISTRY AND FUNCTION, Issue 4 2005Bilgehan Do, ru Pekiner Abstract The Ca2+ -ATPase activity of rat brain microsomes was studied in streptozotocin (STZ)-induced diabetes. Male rats, 200,250,g, were rendered diabetic by injection of STZ (45,mg,kg,1 body weight) via the teil vein. Brain tissues were collected at 1, 4 and 10 weeks after diabetes was induced for determination of Ca2+ -ATPase activity, lipid peroxidation and tissue calcium levels. Diabetic rats had significantly elevated blood glucose levels compared to controls. Blood glucose levels were 92.92,±,1.22,mg,dl,1 (mean,±,SEM) for the control group, 362.50,±,9.61,mg,dl,1 at 1 week and >500,mg,dl,1 at 4, 8 and 10 weeks for the diabetics. Enzyme activities were significantly decreased at 1, 4, 8 and 10 weeks of diabetes relative to the control group (p,<,0.001). Ca2+ -ATPase activity was 0.084,±,0.008,U,l,1, 0.029,±,0.005,U,l,1, 0.029,±,0.006,U,l,1, 0.033,±,0.003,U,l,1 and 0.058,±,0.006,U,l,1 (mean,±,SEM) at control, 1, 4, 8 and 10 week of diabetes respectively. The change in calcium levels in diabetic rat brain at 8 and 10 weeks of diabetes was significantly higher than that of the control group (p,<,0.05). On the other hand lipid peroxidation measured as TBARS (thiobarbituric acid reactive substances) was significantly higher at 8 and 10 weeks of diabetes (p,<,0.05). The increase in lipid peroxidation observed in diabetic rat brain may be partly responsible for the decrease in calcium ATPase activity. Copyright © 2004 John Wiley & Sons, Ltd. [source] Use of monoclonal antibodies to quantify the dynamics of ,-galactosidase and endo-1,4-,-glucanase production by Trichoderma hamatum during saprotrophic growth and sporulation in peatENVIRONMENTAL MICROBIOLOGY, Issue 5 2005Christopher R. Thornton Summary Trichoderma species are ubiquitous soil and peat-borne saprotrophs that have received enormous scientific interest as biocontrol agents of plant diseases caused by destructive root pathogens. Mechanisms of biocontrol such as antibiosis and hyperparasitism are well documented and the biochemistry and molecular genetics of these processes defined. An aspect of biocontrol that has received little attention is the ability of Trichoderma species to compete for nutrients in their natural environments. Trichoderma species are efficient producers of polysaccharide-degrading enzymes that enable them to colonize organic matter thereby preventing the saprotrophic spread of plant pathogens. This study details the use of monoclonal antibodies (mAbs) to quantify the production of two enzymes implicated in the saprotrophic growth of Trichoderma species in peat. Using mAbs specific to the hemicellulase enzyme ,-galactosidase (AGL) and the cellulase enzyme endo-1,4-,-glucanase (EG), the relationship between the saprotrophic growth dynamics of a biocontrol strain of Trichoderma hamatum and the concomitant production of these enzymes in peat-based microcosms was studied. Enzyme activity assays and enzyme protein concentrations derived by enzyme-linked immunosorbent assay (ELISA) established the precision and sensitivity of mAb-based assays in quantifying enzyme production during active growth of the fungus. Trends in enzyme activities and protein concentrations were similar for both enzymes, during a 21-day sampling period in which active growth and sporulation of the fungus in peat was quantified using an independent mAb-based assay. There was a sharp increase in active biomass of T. hamatum 3 days after inoculation of microcosms with phialoconidia. After 3 days there was a rapid decline in active biomass which coincided with sporulation of the fungus. A similar trend was witnessed with EG activities and concentrations. This showed that EG production related directly to active growth of the fungus. The trend was not found, however, with AGL. There was a rapid increase in enzyme activities and protein concentrations on day 3, after which they remained static. The reason for the maintenance of elevated AGL probably resulted from secretion of the enzyme from conidia and chlamydospores. ELISA, immunofluoresence and immunogold electron microscopy studies of these cells showed that the enzyme is localized within the cytoplasm and is secreted extracellularly into the surrounding environment. It is postulated that release of oligosaccharides from polymeric hemicellulose by the constitutive spore-bound enzyme leads to AGL induction and could act as an environmental cue for spore germination. [source] Defining the caspase-containing apoptotic machinery contributing to cornification in human epidermal equivalentsEXPERIMENTAL DERMATOLOGY, Issue 1 2006Vijaya Chaturvedi Abstract:, Whether terminal differentiation/stratum corneum formation of keratinocytes (KCs) represents a form of programmed cell death, utilizing mediators of classical apoptosis, is unclear. Apoptosis, an evolutionarily conserved death process, is comprised of extrinsic and intrinsic pathways, which converge using caspase 3. To define upstream and downstream caspases involved in terminal differentiation, we utilized human epidermal equivalents (EEs). Using submerged cultures comprised of human KCs, EEs were sequentially analyzed before and after being raised to an air/liquid (A/L) interface at 3,24 h intervals. At each time point, EEs were analyzed morphologically and for specific enzyme activity to distinguish different initiator (caspases 1, 2, 8, 9) and effector caspases (3, 6, 7). Terminal differentiation began at 6,8 h, as defined by stratum corneum with loricirin expression and completed at 18,24 h producing an epidermis resembling normal skin. Enzyme activity for caspases 1, 2, 3, 6, 7, 8, and 9 (but not 4, 5) was enhanced (>two-fold nmol/mg/h) at 3,6 h compared with submerged cultures. Processing of caspase 14 occurred at 18 h, and cleaved caspase 14 was increased at 24 h. Activated caspase 3-positive and terminal deoxynucleotidyl transferase-mediated nick end labeling-positive KCs were identified in EEs at 3,6 h corresponding to initiation sites of terminal differentiation. Addition of caspase inhibitors reduced levels of involucrin and loricrin in EEs raised to an A/L interface. We conclude caspases function as important death effectors strategically positioned at intersection of intrinsic and extrinsic pathways in KCs undergoing stratum corneum formation. [source] The identification of a phospholipase B precursor in human neutrophilsFEBS JOURNAL, Issue 1 2009Shengyuan Xu A phospholipase B (PLB) precursor was purified from normal human granulocytes using Sephadex G-75, Mono-S cation-exchange and hydroxyapatite columns. The molecular mass of the protein was estimated to be , 130 kDa by gel filtration and 22 and 42 kDa by SDS/PAGE. Tryptic peptide and sequence analyses by MALDI-TOF and tandem mass spectrometry (MS/MS) identified the protein as a FLJ22662 (Homo sapiens) gene product, a homologue of the amoeba Dictyostelium discoideum PLB. The native protein needed modifications to acquire deacylation activity against phospholipids including phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine and lysophospholipids. Enzyme activity was associated with fragments derived from the 42 kDa fragment. The enzyme revealed a PLB nature by removing fatty acids from both the sn -1 and sn -2 positions of phospholipids. The enzyme is active at a broad pH range with an optimum of 7.4. Immunoblotting of neutrophil postnuclear supernatant using antibodies against the 42 kDa fragment detected a band at a molecular mass of 42 kDa, indicating a neutrophil origin of the novel PLB precursor. The existence of the PLB precursor in neutrophils and its enzymatic activity against phospholipids suggest a role in the defence against invading microorganisms and in the generation of lipid mediators of inflammation. [source] Methotrexate induction of human sulfotransferases in Hep G2 and Caco-2 cellsJOURNAL OF APPLIED TOXICOLOGY, Issue 5 2005Xinrong Chen Abstract Methotrexate (MTX) was the first antifolate drug developed for the treatment of cancer. It is also effective in treating inflammatory and autoimmune diseases. Sulfotransferases are phase II drug-metabolizing enzymes and their induction by hormones and endogenous molecules is relatively well known, although xenobiotic drug induction of sulfotransferases has not been well studied. In the present investigation, MTX is shown to be a xenobiotic inducer of human sulfotransferases in transformed human liver (Hep G2) and intestinal (Caco-2) cells. Following MTX treatment, various sulfotransferases were induced in both cell lines. Enzyme assay, Western blot and reverse-transcription polymerase chain reaction (RT-PCR) results demonstrated that protein and mRNA expressions of human simple phenol sulfotransferase (P-PST), human monoamine sulfotransferase (M-PST), human dehydroepiandrosterone sulfotransferase (DHEA-ST) and human estrogen sulfotransferase (EST) were induced in Hep G2 cells; M-PST and DHEA-ST were induced in Caco-2 cells. Inductions in both cell lines were dose dependent. Enzyme activity and Western blot results were in good agreement with RT-PCR results, suggesting that the induction is at the gene transcription level. Folic acid had a significantly lesser effect on sulfotransferases compared with MTX. Interestingly, the induction of different sulfotransferases by MTX was inhibited by high doses of folic acid at both protein and mRNA levels in Hep G2 cells. Methotrexate is the first antifolate and apoptosis-inducing drug to show induction of sulfotransferases in Hep G2 cells and Caco-2 cells. The inhibition by folic acid suggests a possible mechanism for MTX induction. Copyright © 2005 John Wiley & Sons, Ltd. [source] CHARACTERIZATION OF POLYPHENOL OXIDASE FROM ROOSTER POTATO (SOLANUM TUBEROSUM CV ROOSTER)JOURNAL OF FOOD BIOCHEMISTRY, Issue 1 2010D. NI EIDHIN ABSTRACT The isolation and purification of polyphenol oxidase from potatoes (Solanum tuberosum cv. Rooster) is described. A 64-fold purified preparation has been obtained with 10% yield by a procedure involving (NH4)2SO4 precipitation, phenyl sepharose chromatography, ion exchange chromatography and hydroxyapatite chromatography. The partially purified enzyme has both cresolase and catecholase activity. Activity was lower toward monophenols than diphenols. Enzyme activity was optimal at pH 6.0,6.5 and at 30C. Greater than 50% activity was retained during storage for 72 h at pH 6.0,7.5. Residual activity was greater than 50% after incubation at 20C for 72 h, 30C for 48 h, 40C for 24 h, 50C for 2 h and 60C for 15 min. The most effective inhibitors tested were sodium metabisulfite and ascorbic acid. Sodium dodecyl sulphate appeared to activate the enzyme. The enzyme was capable of cross-linking casein but did not increase gel-strengths in acidified milk gels. PRACTICAL APPLICATIONS Rooster is the most important potato cultivar grown in Ireland and data on its isolation and characterization has not been reported previously. This work describes a method to isolate polyphenol oxidase and characterization of the enzyme. Information on characterization of the enzyme could be valuable in relation to control of enzymatic browning during current processing and in minimum processing. There is potential for use of the enzyme in the emerging cross-linking area, as the results show some success and there may be potential of more cross-linking as the field develops and as interest in natural methods of cross-linking for food texture grows. This could lead to an important use for potato waste. Food product applications are given. [source] KINETIC BEHAVIOR OF SOYBEAN LIPOXYGENASE: A COMPARATIVE STUDY OF THE FREE ENZYME AND THE ENZYME IMMOBILIZED IN AN ALGINATE SILICA SOL-GEL MATRIX,JOURNAL OF FOOD BIOCHEMISTRY, Issue 1 2000AN-FEI HSU Lipoxygenase (LOX) is an enzyme that regioselectively introduces a hydroperoxide into polyunsaturated fatty acids (PUFA). We recently reported a procedure that immobilizes soybean LOX within an alginate sol-gel matrix. In this study, the kinetic profile of free LOX was compared with that of the sol-gel immobilized LOX. The temperature dependent activity profile of free LOX was optimal at 25C whereas immobilized LOX had optimal activity over the temperature range of 25,35C. Enzyme activity, measured in aqueous buffer, for both the free and immobilized LOX preparations had Km values of 2.5 and 1.40 mmoles/L, respectively, and Vmax values of 0.056 and 0.02 ,mol/min, respectively. The relative rates of oxidation of linoleic acid and acylgfycerols containing linoleoyl residues catalyzed by free and immobilized LOX also were determined The results showed that both free and immobilized LOX favor linoleic acid as a substrate. Relative substrate preference for free LOX was linoleic acid >1-monolinolein > 1,3-dilinolein >trilinolein, and for immobilized LOX was linoleic acid >l, 3-dilinolein >1-monolinolein >trilinolein. In general, LOX immobilized in alginate silica sol-gel matrix retained the physical and chemical characteristics of free LOX. [source] | ||||||||||||||||||||||||||||||||||||||||