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Showing papers on "Enzyme assay published in 2010"


Journal ArticleDOI
08 Oct 2010-Science
TL;DR: An enzyme is described that acts on the surface of crystalline chitin, where it introduces chain breaks and generates oxidized chain ends, thus promoting further degradation by chit inases, demonstrating the existence of a hitherto unknown enzyme activity.
Abstract: Efficient enzymatic conversion of crystalline polysaccharides is crucial for an economically and environmentally sustainable bioeconomy but remains unfavorably inefficient. We describe an enzyme that acts on the surface of crystalline chitin, where it introduces chain breaks and generates oxidized chain ends, thus promoting further degradation by chitinases. This enzymatic activity was discovered and further characterized by using mass spectrometry and chromatographic separation methods to detect oxidized products generated in the absence or presence of H 2 18 O or 18 O 2 . There are strong indications that similar enzymes exist that work on cellulose. Our findings not only demonstrate the existence of a hitherto unknown enzyme activity but also provide new avenues toward more efficient enzymatic conversion of biomass.

1,050 citations


Journal ArticleDOI
TL;DR: This review has been focused on the different types of techniques used for the immobilization of β-galactosidase and its potential applications in food industry.
Abstract: The enzyme β-galactosidase can be obtained from a wide variety of sources such as microorganisms, plants, and animals. The use of β-galactosidase for the hydrolysis of lactose in milk and whey is one of the promising enzymatic applications in food and dairy processing industries. The enzyme can be used in either soluble or immobilized forms but the soluble enzyme can be used only for batch processes and the immobilized form has the advantage of being used in batch wise as well as in continuous operation. Immobilization has been found to be convenient method to make enzyme thermostable and to prevent the loss of enzyme activity. This review has been focused on the different types of techniques used for the immobilization of β-galactosidase and its potential applications in food industry.

239 citations


Journal ArticleDOI
TL;DR: Hg-induced oxidative stress was responsible for the disturbance of the growth and antioxidant defensive systems in C. reinhardtii, a unicellular model organism, to Hg2+-induced toxicity.
Abstract: Investigation of mercury toxicology in green algae is of great importance from ecological point of view, because mercury has become a major contaminant in recent years. In higher plants, accumulation of mercury modifies many aspects of cellular functions. However, the process that mercury exerts detrimental effects on green algae is largely unknown. In this study, we performed an experiment focusing on the biological responses of Chlamydomonas reinhardtii, a unicellular model organism, to Hg(2+)-induced toxicity. C. reinhardtii was exposed to 0, 1, 2, 4, 6, and 8 μM Hg in media. Concentrations of Hg were negatively correlated with the cell growth. Treatment with Hg induced accumulation of reactive oxygen species and peroxidative products. Endogenous proline levels increased in Hg-exposed algae. Hg exposure activated superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX). To get insights into the molecular response, a RT-PCR-based assay was performed to analyze the transcript abundance of Mn-SOD, CAT and APX. Our analysis revealed that expression of the genes was up-regulated by Hg exposure, with a pattern similar to the enzyme activities. Additional investigation was undertaken on the effect of Hg on the transcript amount of ∆(1)-pyrroline-5-carboxylate synthetase, a key enzyme of proline biosynthesis and on that of heme oxygenase-1 (HO-1), an enzyme regulating heavy metal tolerance. Expressions of both P5CS and HO-1 were up-regulated by Hg. These data indicate that Hg-induced oxidative stress was responsible for the disturbance of the growth and antioxidant defensive systems in C. reinhardtii.

178 citations


Book ChapterDOI
01 Jan 2010
TL;DR: The controls on enzyme temperature sensitivity and their importance for understanding seasonal patterns in soil processes and their potential responses to global change are explored.
Abstract: Enzyme activities are commonly measured in lab assays at a single standard temperature, which does not provide any information on their temperature sensitivity. Temperature is one of the primary controls on enzyme activities, yet few studies have explored how temperature drives enzyme activities in the environment. The temperature sensitivity of enzyme activity is controlled by the structure and conformation of the isoenzymes that constitute an environmental enzyme pool as well as physical and chemical interactions with soil minerals, clays, and organic matter. Yet, these complex relationships are typically represented by a simple Q 10 of 2. There is sufficient evidence to suggest that even for the same enzyme class, temperature sensitivities vary between soils, and even seasonally in a single site. We will explore the controls on enzyme temperature sensitivity and their importance for understanding seasonal patterns in soil processes and their potential responses to global change.

178 citations


Journal ArticleDOI
TL;DR: It is demonstrated that the elevated levels of biochemicals, phenolic acids, and enzymes may play a major role in plant defense.
Abstract: A laboratory study was undertaken to ascertain the impact and the extent of feeding by different pests on biochemical constituents and various enzyme levels in rice plants. The difference in these parameters due to the pest damage by three different modes of feeding was also studied and compared. The borer pest—yellow stem borer (YSB), Scirpophaga incertulas (W); surface feeder—-leaf roller (LR), Cnaphalocrosis medinalis (G) and a sucking pest—brown plant hopper (BPH), Nilaparvata lugens (S) fed rice plants were analyzed for the quantitative and qualitative changes in biochemical profile and enzymatic changes that occur as plant’s defensive responses were analyzed spectrophotometrically. The phenolic acids were analyzed using HPLC and quantitated with the standard samples. The quantity of biochemicals such as proteins, phenols and carbohydrates has been enhanced along with the enzyme activities of peroxidase (POD), catalase (CAT), chitinase (CHI). A decrease in superoxide dismutase (SOD), phenyl alanine ammonia lyase (PAL), β-1, 3-glucanase (GLU) enzyme activities were evident in pest infested plants. Phenolic acids like vanillic acid, syringic acid, cinnamic acid, and p-coumaric acids were mostly found in the infested plants. We demonstrate that the elevated levels of biochemicals, phenolic acids, and enzymes may play a major role in plant defense.

169 citations



Book ChapterDOI
TL;DR: This chapter provides details of the assays used to determine the enzyme activities spectrophotometrically and Antioxidant enzyme responses to moderate water-deficit stress were studied.
Abstract: Reactive oxygen species (ROS) are formed in biological systems as part of normal metabolism. Adverse environmental factors like drought stress result in increased levels of ROS that are detrimental to the plant (1, 2). To avoid damage caused by these excess ROS, plants have developed elaborate mechanisms to manage them at sustainable levels. Enzymes play an important role in lowering the ROS levels and helping avoid oxidative stress. Superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase play a vital role in combating oxidative stress. Measuring these enzyme activities spectrophotometrically provides researchers an easy and precise way to study and understand an important part of the defense against oxidative stress. In this chapter we provide details of the assays we used to determine the enzyme activities spectrophotometrically. Antioxidant enzyme responses to moderate water-deficit stress were studied. All enzyme assays were conducted using wheat leaf tissue.

157 citations


Journal ArticleDOI
TL;DR: It is found that labile C and nutrient-acquiring enzymes, not those involved in the degradation of recalcitrant compounds, were the best predictors of respiration rates.
Abstract: Variation in organic matter inputs caused by differences in plant community composition has been shown to affect microbial activity, although the mechanisms controlling these effects are not entirely understood. In this study we determine the effects of variation in substrate composition, quantity, and diversity on soil extracellular enzyme activity and respiration in laboratory microcosms. Microbial respiration responded predictably to substrate composition and quantity and was maximized by the addition of labile substrates and greater substrate quantity. However, there was no effect of substrate diversity on respiration. Substrate composition significantly affected enzyme activity. Phosphatase activity was maximized with addition of C and N together, supporting the common notion that addition of limiting resources increases investment in enzymes to acquire other limiting nutrients. Chitinase activity was maximized with the addition of chitin, suggesting that some enzymes may be stimulated by the addition of the substrate they degrade. In contrast, activities of glucosidase and peptidase were maximized by the addition of the products of these enzymes, glucose and alanine, respectively, for reasons that are unclear. Substrate diversity and quantity also stimulated enzyme activity for three and four of the six enzymes assayed, respectively. We found evidence of complementary (i.e., non-additive) effects of additions of different substrates on activity for three of the six enzymes assayed; for the remaining enzymes, effects of adding a greater diversity of substrates appeared to arise from the substrate-specific effects of those substrates included in the high-diversity treatment. Finally, in a comparison of measures of microbial respiration and enzyme activity, we found that labile C and nutrient-acquiring enzymes, not those involved in the degradation of recalcitrant compounds, were the best predictors of respiration rates. These results suggest that while composition, quantity, and diversity of inputs to microbial communities all affect microbial enzyme activity, the mechanisms controlling these relationships are unique for each particular enzyme.

136 citations


Journal ArticleDOI
TL;DR: Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO2 and may induce an alternative active protein state with altered enzyme activity, specificity and stability.
Abstract: Different types of enzymes such as lipases, several phosphatases, dehydrogenases, oxidases, amylases and others are well suited for the reactions in SC-CO2. The stability and the activity of enzymes exposed to carbon dioxide under high pressure depend on enzyme species, water content in the solution and on the pressure and temperature of the reaction system. The three-dimensional structure of enzymes may be significantly altered under extreme conditions, causing their denaturation and consequent loss of activity. If the conditions are less adverse, the protein structure may be largely retained. Minor structural changes may induce an alternative active protein state with altered enzyme activity, specificity and stability.

134 citations


Journal ArticleDOI
TL;DR: The supplementation of enzyme with essential oil in diets is likely more effective in view of performance, nutrient digestibility, enzyme activities and immune response of broilers fed on wheat–soybean meal based diets.
Abstract: 1. The study was conducted to determine the effects of dietary supplementation of enzyme and oregano essential oil at two levels, alone or together, on performance, digestive enzyme, nutrient digestibility, lipid metabolism and immune response of broilers fed on wheat-soybean meal based diets. 2. The following dietary treatments were used from d 0 to 21. Diet 1 (control, CONT): a commercial diet containing no enzyme or oregano essential oil, diet 2 (ENZY): supplemented with enzyme, diet 3 (EO250): supplemented with essential oil at 250 mg/kg feed, diet 4 (EO500): supplemented with essential oil at 500 mg/kg feed, diet 5 (ENZY + EO250): supplemented with enzyme and essential oil at 250 mg/kg, and diet 6 (ENZY + EO500): supplemented with enzyme and essential oil at 500 mg/kg. 3. Birds fed on diets containing ENZY, EO250 and ENZY + EO250 had significantly higher weight gain than those given CONT diet from d 0 to 7. No significant effects on feed intake, feed conversion ratio, mortality, organ weights except for jejunum weight and intestinal lengths was found with either enzyme or essential oil, alone or in combination, over the 21-d growth period. The supplementation of essential oil together with enzyme decreased jejunum weight compared with essential oil alone. 4. Supplementation with enzyme significantly decreased viscosity and increased dry matter of digesta, but did not alter pH of digesta. There was no effect of essential oil alone at either concentration on viscosity, dry matter or pH of digesta. A significant decrease in viscosity of digesta appeared when essential oil was used with together enzyme. 5. The supplementation of essential oil at both levels with or without enzyme significantly increased chymotrypsin activity in the digestive system, and improved crude protein digestibility. 6. The higher concentration of essential oil with and without enzyme significantly increased serum total cholesterol concentrations. No significant effect on immune response was found with either enzyme or essential oil, alone or together. 7. Enzymes and essential oil had different modes of actions. The supplementation of enzyme with essential oil in diets is likely more effective in view of performance, nutrient digestibility, enzyme activities and immune system.

133 citations


Journal ArticleDOI
TL;DR: This is the first report of using plant-derived enzyme cocktails for production of fermentable sugars from lignocellulosic biomass and pectate lyase transplastomic plants showed enhanced resistance to Erwina soft rot.
Abstract: It is widely recognized that biofuel production from lignocellulosic materials is limited by inadequate technology to efficiently and economically release fermentable sugars from the complex multi-polymeric raw materials. Therefore, endoglucanases, exoglucanase, pectate lyases, cutinase, swollenin, xylanase, acetyl xylan esterase, beta glucosidase and lipase genes from bacteria or fungi were expressed in Escherichia coli or tobacco chloroplasts. A PCR-based method was used to clone genes without introns from Trichoderma reesei genomic DNA. Homoplasmic transplastomic lines showed normal phenotype and were fertile. Based on observed expression levels, up to 49, 64 and 10, 751 million units of pectate lyases or endoglucanase can be produced annually, per acre of tobacco. Plant production cost of endoglucanase is 3100-fold, and pectate lyase is 1057 or 1480-fold lower than the same recombinant enzymes sold commercially, produced via fermentation. Chloroplast-derived enzymes had higher temperature stability and wider pH optima than enzymes expressed in E. coli. Plant crude-extracts showed higher enzyme activity than E. coli with increasing protein concentration, demonstrating their direct utility without purification. Addition of E. coli extracts to the chloroplast-derived enzymes significantly decreased their activity. Chloroplast-derived crude-extract enzyme cocktails yielded more (up to 3625%) glucose from filter paper, pine wood or citrus peel than commercial cocktails. Furthermore, pectate lyase transplastomic plants showed enhanced resistance to Erwina soft rot. This is the first report of using plant-derived enzyme cocktails for production of fermentable sugars from lignocellulosic biomass. Limitations of higher cost and lower production capacity of fermentation systems are addressed by chloroplast-derived enzyme cocktails.

Journal ArticleDOI
TL;DR: It is suggested that while excess copper slightly alters the activity of the antioxidative enzyme system in young expanding leaves of bean plants, it exerts its toxicity primarily through causing a disturbance in the nutrient balance.

Journal ArticleDOI
S. Amena1, N. Vishalakshi1, M. Prabhakar1, A Dayanand1, K. Lingappa1 
TL;DR: A novel strain of Streptomyces gulbargensis was explored for the production of extra-cellular L-asparaginase using groundnut cake extract and it was found to be more stable at the alkaline pH than at the acidic one.


Journal ArticleDOI
TL;DR: In this article, the authors identify potential angiotensin I-converting enzyme (ACE) inhibitory peptide from egg white protein by HPLC assay and the 50% inhibitory concentrations of Arg-Val-Pro-Ser-Leu (RVPSL) was 20μM.

Journal ArticleDOI
TL;DR: Horseeradish peroxidase (HRP) from horseradish roots was purified using a simple, rapid, three-step procedure: ultrasonication, ammonium sulfate salt precipitation, and hydrophobic interaction chromatography on phenyl Sepharose CL-4B with high specific activity.
Abstract: Peroxidase (EC 1.11.1.7) from horseradish (Armoracia rusticana) roots was purified using a simple, rapid, three-step procedure: ultrasonication, ammonium sulfate salt precipitation, and hydrophobic interaction chromatography on phenyl Sepharose CL-4B. The preparation gave an overall yield of 71%, 291-fold purification, and a high specific activity of 772 U mg−1 protein. Sodium dodecyl sulfate−polyacrylamide gel electrophoresis revealed that the purified enzyme was homogeneous and had a molecular weight of approximately 40 kDa. The isolated enzyme had an isoelectric point of 8.8 and a Reinheitszahl value of 3.39 and was stable when stored in the presence of glycerol at −20 °C, with >95% retention of original enzyme activity for at least 6 months. Maximal activity of purified horseradish peroxidase (HRP) was obtained under different optimized conditions: substrate (guaiacol and H2O2) concentrations (0.5 and 0.3 mM, respectively), type of buffer (50 mM phosphate buffer), pH (7.0), time (1.0 min), and tempera...

Journal ArticleDOI
Yang Gao1, Pei Zhou1, Liang Mao1, Yue-er Zhi1, Wanjun Shi1 
TL;DR: In this article, a laboratory study was conducted to evaluate the response of soil enzyme activities (namely dehydrogenase, phosphatase and urease) to different levels of trace element pollution in soil representative area.
Abstract: A laboratory study was conducted to evaluate the response of soil enzyme activities (namely dehydrogenase, phosphatase and urease) to different levels of trace element pollution in soil representative area. The improved ecological dose model and random-amplified polymorphic DNA (RAPD) were used to assess soil health. The 50% ecological dose (ED50) values modified by toxicant coefficient were calculated from the best-fit model, and determination values from the regression analysis for the three enzyme activities were studied after the incubation periods. The results showed that the elevated heavy metal concentration negatively affects the total population size of bacteria and actinomycetes and enzymatic activity; dehydrogenase (ED50 = 777) was the most sensitive soil enzyme, whereas urease activity (ED50 = 2,857) showed the lowest inhibition; combined pollution or elevated toxicant level would increase disappearing RAPD bands, and the number of denoting polymorphic bands was greater in combined polluted soils. All three mathematical modified models satisfactorily described the inhibition of soil enzyme activities caused by Cd and Pb, by giving the best fit.

Journal ArticleDOI
TL;DR: Gene expression and enzyme activity were compared during agar interactions between Trametes versicolor and five other wood decay fungi resulting in a range of interaction outcomes from deadlock to replacement of one fungus by another.

Journal ArticleDOI
TL;DR: The present review will highlight the literature of CE-based enzyme assays from 2006 to November 2009 and one section will be devoted to applications of microfluidic devices.

Journal ArticleDOI
TL;DR: High ratio of milk clotting to proteolytic activity coupled with low thermal stability strengthens the potential usefulness of milk-clotting enzyme of A. oryzae MTCC 5341 as a substitute for calf rennet in cheese manufacturing.
Abstract: Microbial milk-clotting enzymes are valued as calf rennet substitutes in the cheese industry. Aspergillus oryzae MTCC 5341 was identified to produce the highest milk-clotting activity during screening of 16 fungal strains. Solid state fermentation using wheat bran along with 4% defatted soy flour and 2% skim milk powder as substrate was optimal for growth of A. oryzae and production of the enzyme. Nearly 40,000 U/g bran of milk-clotting activity was present at the end of 120 h. The enzyme could be recovered by percolating the bran with 0.1 M sodium chloride for 60 min at 4°C. The decolorized enzyme preparation had high ratio of milk clotting to proteolytic activity. Affinity precipitation with alginate and subsequent elution with 0.5 M sodium chloride containing 0.2 M CaCl2 resulted in an enzyme preparation with specific activity of 3,500 U/mg and 72% yield. Optimum pH and temperature for activity of the enzyme were characterized as 6.3 and 55°C, respectively. Milk-clotting enzyme showed differential degree of hydrolysis on casein components. High ratio of milk clotting to proteolytic activity coupled with low thermal stability strengthens the potential usefulness of milk-clotting enzyme of A. oryzae MTCC 5341 as a substitute for calf rennet in cheese manufacturing.

Journal ArticleDOI
TL;DR: Though immobilized enzyme system was able to reduce Cr(VI), the performance was not very encouraging in continuous mode of operation.
Abstract: In the present study, hexavalent chromium (Cr(VI)) reduction potential of chromium reductase associated with the cell-free extracts (CFE) of Arthrobacter rhombi-RE species was evaluated. Arthrobacter rhombi-RE, an efficient Cr(VI) reducing bacterium, was enriched and isolated from a chromium-contaminated site. Chromium reductase activity of Arthrobacter rhombi-RE strain was associated with the cell-free extract and the contribution of extracellular enzymes to Cr(VI) reduction was negligible. NADH enhanced the chromium reductase activity. The enzyme activity was optimal at a pH of 5.5 and a temperature of 30 °C. Among the ten electron donors screened, sodium pyruvate was the most effective one followed by NADH and propionic acid. Michaelis–Menten constant, Km, and maximum reaction rate, Vmax, obtained from the Lineweaver–Burk plot were 48 μM and 4.09 nM/mg protein/min, respectively, in presence of NADH as electron donor and 170.5 μM and 4.29 nM/mg protein/min, respectively, in presence of sodium pyruvate as electron donor. Ca2+ enhanced the enzyme activity while Hg2+, Cd2+, Ba2+, and Zn2+ inhibited the enzyme activity. Among the various immobilization matrices screened, calcium alginate beads seemed to be the most effective one. Though immobilized enzyme system was able to reduce Cr(VI), the performance was not very encouraging in continuous mode of operation.

Journal ArticleDOI
TL;DR: The feasibility of using tagged enzyme to enhance delivery and pathological effectiveness in Morquio A mice is indicated and substantial clearance of the storage materials in bone, bone marrow, and heart valves is shown, especially after 24 weekly infusions.

Journal ArticleDOI
TL;DR: A mycelial β-glucosidase from the thermophilic mold Humicola insolens was purified and biochemically characterized in this paper, and the enzyme showed carbohydrate content of 21% and apparent molecular mass of 94 kDa, as estimated by gel filtration.

Journal ArticleDOI
TL;DR: Results indicated that AtNUDX6 is a modulator of NADH rather than ADP-Rib metabolism and that, through induction of TRX-h5 expression, AtNudX6 significantly impacts the plant immune response as a positive regulator of NPR1-dependent SA signaling pathways.
Abstract: Here, we investigated the physiological role of Arabidopsis (Arabidopsis thaliana) AtNUDX6, the gene encoding ADP-ribose (Rib)/NADH pyrophosphohydrolase, using its overexpressor (Pro35S:AtNUDX6) or disruptant (KO-nudx6) The level of NADH in Pro35S:AtNUDX6 and KO-nudx6 plants was decreased and increased, respectively, compared with that of the control plants, while the level of ADP-Rib was not changed in either plant The activity of pyrophosphohydrolase toward NADH was enhanced and reduced in the Pro35S:AtNUDX6 and KO-nudx6 plants, respectively The decrease in the activity of NADH pyrophosphohydrolase and the increase in the level of NADH were observed in the rosette and cauline leaves, but not in the roots, of the KO-nudx6 plants Notably, the expression level of AtNUDX6 and the activity of NADH pyrophosphohydrolase in the control plants, but not in the KO-nudx6 plants, were increased by the treatment with salicylic acid (SA) The expression of SA-induced genes (PR1, WRKY70, NIMIN1, and NIMIN2) depending on NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1), a key component required for pathogen resistance, was significantly suppressed and enhanced in the KO-nudx6 and Pro35S:AtNUDX6 plants, respectively, under the treatment with SA Induction of thioredoxin h5 (TRX-h5) expression, which catalyzes a SA-induced NPR1 activation, was suppressed and accelerated in the KO-nudx6 and Pro35S:AtNUDX6 plants, respectively The expression of isochorismate synthase1, required for the regulation of SA synthesis through the NPR1-mediated feedback loop, was decreased and increased in the KO-nudx6 and Pro35S:AtNUDX6 plants, respectively Judging from seed germination rates, the KO-nudx6 plants had enhanced sensitivity to the toxicity of high-level SA These results indicated that AtNUDX6 is a modulator of NADH rather than ADP-Rib metabolism and that, through induction of TRX-h5 expression, AtNUDX6 significantly impacts the plant immune response as a positive regulator of NPR1-dependent SA signaling pathways

Journal ArticleDOI
TL;DR: Two different carbon sources, glucose and maltose were used for the enzyme production and glucose was found to be the better carbon source and the enzyme was partially purified by ammonium sulphate precipitation.
Abstract: 2Azyme Biosciences Private Limited, 1188/20, 3 rd Floor, 26 th Main, 9 th Block, Jaya Nagar, Bengaluru 560 069, India. Accepted 22 July, 2010 L-asparaginase is an anti-neoplastic agent used in lymphoblastic leukaemia chemotherapy. Soil microbial isolates were screened for potential producers of L-asparaginase using a phenol red indicator growth medium and the microbe producing the largest hydrolysis zone was selected. The isolate was characterised by biochemical tests and was found to belong to Bacillus sp. The enzyme production was carried out by submerged fermentation. Two different carbon sources, glucose and maltose were used for the enzyme production and glucose was found to be the better carbon source. The enzyme was partially purified by ammonium sulphate precipitation. Dialysis was carried out to remove the excess salt and complete purification was achieved by ion-exchange chromatography. The characterised enzyme exhibited maximal enzyme activity at pH 7 and temperature 37°C. The enzyme was activated by MgCl2 and inhibited by EDTA. Protein profiling by SDS-PAGE revealed the molecular weight of the protein to be 45 kDa.

Journal ArticleDOI
TL;DR: O2- produced by NADPH oxidases after imbibition regulates seed germination and seedling growth in barley, indicating the importance of NAD PH oxidases in germination of barley seeds.
Abstract: Reactive oxygen species (ROS) play an important role in seed germination. Although hydrogen peroxide (H2O2), a type of ROS, enhances the germination rate of various plant seeds, little is known about the mechanism. NADPH oxidases catalyze the production of superoxide anion (O2-) that is one of the ROS and the enzymes regulate plant development. We, therefore, investigated the role of NADPH oxidases in seed germination and seedling growth in barley (Hordeum vulgare L.). The production of O2- was observed both in embryo and aleurone layers in barley seeds treated with distilled water (DW). However, it was suppressed in seeds treated with diphenylene iodonium (DPI) chloride, NADPH oxidase inhibitor. Moreover, DPI markedly delayed germination and remarkably suppressed α-amylase activity in barley seeds, indicating the importance of NADPH oxidases in germination of barley seeds. The gene expression and the enzyme activity of NADPH oxidases gradually increased after imbibition, and the enzyme activities were cl...

Journal ArticleDOI
TL;DR: Results confirmed that the studied enzyme was an α-amylase ((1→4)-α-glucan glucanohydrolase), which was activated mainly by calcium and cobalt, and it presented as a glycoprotein with 23% carbohydrate content.

Journal ArticleDOI
TL;DR: A newly isolated Geobacillus sp.
Abstract: A newly isolated Geobacillus sp. IIPTN (MTCC 5319) from the hot spring of Uttarakhand's Himalayan region produced a hyperthermostable α-amylase. The microorganism was characterized by biochemical tests and 16S rRNA gene sequencing. The optimal temperature and pH were 60°C and 6.5, respectively, for growth and enzyme production. Although it was able to grow in temperature ranges from 50 to 80°C and pH 5.5–8.5. Maximum enzyme production was in exponential phase with activity 135 U ml−1 at 60°C. Assayed with cassava as substrate, the enzyme displayed optimal activity 192 U ml−1 at pH 5.0 and 80°C. The enzyme was purified to homogeneity with purification fold 82 and specific activity 1,200 U mg−1 protein. The molecular mass of the purified enzyme was 97 KDa. The values of K m and V max were 36 mg ml−1 and 222 µmol mg−1 protein min−1, respectively. The amylase was stable over a broad range of temperature from 40°C to 120°C and pH ranges from 5 to 10. The enzyme was stimulated with Mn2+, whereas it was inhibited by Hg2+, Cu2+, Zn2+, Mg2+, and EDTA, suggesting that it is a metalloenzyme. Besides hyperthermostability, the novelty of this enzyme is resistance against protease.

Journal ArticleDOI
TL;DR: The characteristics of novel A. baylyi lipase, as high temperature stability, organic solvent tolerance, and transesterification capacity from palm oil to FAMEs, indicate that it could be a vigorous biocatalyzer in the prospective fields as bioenergy industry or even in organic synthesis and pharmaceutical industry.
Abstract: The benzene tolerant Acinetobacter baylyi isolated from marine sludge in Angsila, Thailand could constitutively secrete lipolytic enzymes. The enzyme was successfully purified 21.89-fold to homogeneity by ammonium sulfate precipitation and gel-permeable column chromatography with a relative molecular mass as 30 kDa. The enzyme expressed maximum activity at 60°C and pH 8.0 with p-nitrophenyl palmitate as a substrate and found to be stable in pH and temperature ranging from 6.0-9.0 to 60-80°C, respectively. A study on solvent stability revealed that the enzyme was highly resisted to many organic solvents especially benzene and isoamyl alcohol, but 40% inhibited by decane, hexane, acetonitrile, and short-chain alcohols. Lipase activity was completely inhibited in the presence of Fe2+, Mn2+, EDTA, SDS, and Triton X-100 while it was suffered detrimentally by Tween 80. The activity was enhanced by phenylmethylsulfonyl fluoride (PMSF), Na+, and Mg2+ and no significant effect was found in the presence of Ca2+ and Li+. Half of an activity was retained by Ba2+, Ag+, Hg+, Ni2+, Zn2+, and DTT. The enzyme could hydrolyze a wide range of p-nitrophenyl esters, but preferentially medium length acyl chains (C8-C12). Among natural oils and fats, the enzyme 11-folds favorably catalyzed the hydrolysis of rice bran oil, corn oil, sesame oil, and coconut oil in comparison to palm oil. Moreover, the transesterification activity of palm oil to fatty acid methyl esters (FAMEs) revealed 31.64 ± 1.58% after 48 h. The characteristics of novel A. baylyi lipase, as high temperature stability, organic solvent tolerance, and transesterification capacity from palm oil to FAMEs, indicate that it could be a vigorous biocatalyzer in the prospective fields as bioenergy industry or even in organic synthesis and pharmaceutical industry.

Journal ArticleDOI
TL;DR: Sardans et al. as mentioned in this paper investigated the role of soil enzyme activity in a Mediterranean forest after six years of Drought in Soil Biology & Biochemistry and found that the enzyme activity was positively correlated with soil fertility.
Abstract: Soil Sci. Soc. Am. J. 74:838–851 Published online 17 Mar. 2010 doi:10.2136/sssaj2009.0225 Received 16 June 2009. *Corresponding author (j.sardans@creaf.uab.cat). © Soil Science Society of America, 5585 Guilford Rd., Madison WI 53711 USA All rights reserved. No part of this periodical may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher. Permission for printing and for reprinting the material contained herein has been obtained by the publisher. Soil Enzyme Activity in a Mediterranean Forest after Six Years of Drought Soil Biology & Biochemistry