Characterization of cellulase-free xylanases from the newly isolated Bacillus sp. strain BP-23

1993 ◽  
Vol 39 (12) ◽  
pp. 1162-1166 ◽  
Author(s):  
A. Blanco ◽  
F. I. J. Pastor
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Xylanase Activity ◽  
Hydrolytic Activity ◽  
Bacillus Sp ◽  
Birchwood Xylan ◽  
Xylanase Production ◽  
Ph Values ◽  
Wide Range ◽  
Molecular Masses

A Bacillus strain with xylanase activity has been isolated. Maximum xylanase production was obtained when the strain was cultured in media supplemented with birchwood xylan or rice straw; production was repressed by glucose and xylose. The optimal temperature and pH for xylanase activity were 45–50 °C and 5.5–7.5, respectively. Crude xylanase was highly stable at a wide range of pH values, retaining 100% of the activity after 24 h of incubation at 37 °C in buffer at pH 10.0. Analysis by polyacrylamide gel electrophoresis and zymogram techniques showed four xylanase activity bands with apparent molecular masses of 32, 48, 61, and 66 kDa. The most active of them (molecular mass 32 kDa) apparently corresponded to a xylanase with an isoelectric point (pI) of 9.3 in isoelectrofocusing gels developed as zymograms. Four other bands with xylanase activity were detected at pIs of 7.7, 5.6, 5.0, and 4.5. Analysis for carboxymethylcellulase activity revealed that only the band of 48 kDa and the band with a pI of 7.7 showed hydrolytic activity against the cellulosic substrate.Key words: Bacillus sp., xylanase, isolation.

scholarly journals Optimization of production and partial characterization of xylanase from a newly isolated Bacillus amyloliquefaciens

2020 ◽  
Vol 42 ◽  
pp. e9
Author(s):  
Bruno Las-Casas Chaves ◽  
Ana Paula Martinazzo ◽  
Brisabella Coca ◽  
Adriane Nunes De Souza ◽  
Carlos Eduardo Teodoro
Keyword(s):  
Carbon Sources ◽  
Xylanase Activity ◽  
Residual Activity ◽  
Inoculum Size ◽  
Production Optimization ◽  
Partial Characterization ◽  
Birchwood Xylan ◽  
Xylanase Production ◽  
Local Soil

This paper reports the process of production optimization and partial characterization of xylanase from a newly isolated Bacillus amyloliquefacies VR002, isolated from local soil. The microorganism exhibited maximum xylanase production when 1.0% (v/v) of inoculum size was added to culture medium with initial pH 6, 1.0% (w/v) birchwood xylan, at 35 °C after 48h of incubation. Xylanase production in different carbon sources apart from birchwood xylan and xylose did not show high production levels. Optimum pH for xylanase activity was 6.0. The enzyme was alkali-stable and retained 100% of residual activity over the pH range from 6.0 to 10.0 for 24 h at 25°C. Optimum temperature for enzyme activity was 55°C. Xylanase was 100% stable at 4°C and 25°C even after 24h of incubation, a desirable characteristic for enzyme storage. Moreover, best crude extract volume and time reaction were found to be 10 µL and 5 min, respectively. After optimization of production and activity parameters, an increase of nearly 60-fold in xylanase activity (44.12 ± 4.36 U/mL) was achieved. Characteristics of B. amyloliquefaciens VR002 xylanase are particularly desirable for biotechnological applications

scholarly journals Characterization of the Different Dextransucrase Activities Excreted in Glucose, Fructose, or Sucrose Medium byLeuconostoc mesenteroides NRRL B-1299

1998 ◽  
Vol 64 (4) ◽  
pp. 1298-1302 ◽  
Author(s):  
Marguerite Dols ◽  
M. Remaud-Simeon ◽  
R. M. Willemot ◽  
M. Vignon ◽  
P. Monsan
Keyword(s):  
Gel Electrophoresis ◽  
Leuconostoc Mesenteroides ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Traditional Culture ◽  
Culture Conditions ◽  
Acceptor Reaction ◽  
Sucrose Medium ◽  
Molecular Masses

ABSTRACT When grown in glucose or fructose medium in the absence of sucrose,Leuconostoc mesenteroides NRRL B-1299 produces two distinct extracellular dextransucrases named glucose glucosyltransferase (GGT) and fructose glucosyltransferase (FGT). The production level of GGT and FGT is 10 to 20 times lower than that of the extracellular dextransucrase sucrose glucosyltransferase (SGT) produced on sucrose medium (traditional culture conditions). GGT and FGT were concentrated by ultrafiltration before sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. Their molecular masses were 183 and 186 kDa, respectively, differing from the 195 kDa of SGT. The structural analysis of the dextran produced from sucrose and of the oligosaccharides synthesized by acceptor reaction in the presence of maltose showed that GGT and FGT are two different enzymes not previously described for this strain. The polymer synthesized by GGT contains 30% α(1→2) linkages, while FGT catalyzes the synthesis of a linear dextran only composed of α(1→6) linkages.

scholarly journals Production and Partial Characterization of an Alkaline Xylanase from a Novel FungusCladosporium oxysporum

2016 ◽  
Vol 2016 ◽  
pp. 1-7 ◽  
Author(s):  
Guo-Qiang Guan ◽  
Peng-Xiang Zhao ◽  
Jin Zhao ◽  
Mei-Juan Wang ◽  
Shu-Hao Huo ◽  
...  
Keyword(s):  
Nitrogen Source ◽  
Wheat Bran ◽  
Internal Transcribed Spacer ◽  
Gene Sequence ◽  
Agricultural Waste ◽  
Xylanase Activity ◽  
Maximum Activity ◽  
Alkaline Proteases ◽  
Xylanase Production

A new fungusCladosporium oxysporumGQ-3 producing extracellular xylanase was isolated from decaying agricultural waste and identified based on the morphology and comparison of internal transcribed spacer (ITS) rDNA gene sequence.C. oxysporumproduced maximum xylanase activity of 55.92 U/mL with wheat bran as a substrate and NH4Cl as a nitrogen source. Mg2+improvedC. oxysporumxylanase production.Partially purified xylanase exhibited maximum activity at 50°C and pH 8.0, respectively, and showed the stable activity after 2-h treatment in pH 7.0–8.5 or below 55°C. Mg2+enhanced the xylanase activity by 2% while Cu2+had the highest inhibition ratio of 57.9%. Furthermore,C. oxysporumxylanase was resistant to most of tested neutral and alkaline proteases. Our findings indicated thatCladosporium oxysporumGQ-3 was a novel xylanase producer, which could be used in the textile processes or paper/feed industries.

scholarly journals Optimization of alkali-thermostable and cellulase-free eylanase production from Bacillus Sp.

2012 ◽  
Vol 19 ◽  
pp. 7-14
Author(s):  
SCD Sharma ◽  
MS Shovon ◽  
AKM Asaduzzaman ◽  
MG Sarowar Jahan ◽  
T Yeasmin ◽  
...  
Keyword(s):  
Wheat Bran ◽  
Agar Medium ◽  
Xylanase Activity ◽  
Effect Of Temperature ◽  
Bacillus Sp ◽  
Dilution Technique ◽  
Isolation And Identification ◽  
Xylanase Production ◽  
Optimization Of Process Parameters ◽  
Bacterium Bacillus

Context: To analyze the nutritional and physicochemical parameters for the production of alkali-thermostable and cellulase free xylanase from bacteria. Objectives: The aim of this study was to isolation and identification and of alkali-thermostable and cellulase free xylanase producing bacteria from soil as well as optimization of process parameters for xylanase production. Materials and Methods: The bacterium Bacillus sp. was isolated from soil by serial dilution technique on xylan agar medium and identified by morphological and biochemical studies. The production of xylanase was carried out on xylan broth medium and xylanase activity was assayed by dinitrosalicylic acid (DNS) method. The effect of cultural parameters on the production of xylanase was determined by measuring the activity of xylanase. The effect of temperature and pH on the activity of partially purified xylanase as well as substrate specificity of xylanase were examined. Results: The maximum xylanase production (4000 U/L) by a Bacillus sp. was attained when the medium containing 0.5% wheat bran xylan and peptone at pH 8.0 and 50-55°C within 48-60 h. The partially purified xylanase was optimally active at pH 9.0 and 55°C. The xylanase showed high substrate activity towards wheat bran xylan but no activity towards cellulose, carboxymethyl cellulose and starch. Thus the enzyme was alkali-thermostable and cellulase free xylanase. Conclusion: The results obtained in this study suggest that the Bacillus sp. used is highly potential and useful for the production of cellulase free xylanase. DOI: http://dx.doi.org/10.3329/jbs.v19i0.12994 J. bio-sci. 19: 7-14, 2011

scholarly journals Component analysis and characterization of a nuclear deoxyribonucleotidase

1994 ◽  
Vol 298 (3) ◽  
pp. 727-732 ◽  
Author(s):  
K Ford ◽  
J Waltho ◽  
D Hornby
Keyword(s):  
Mammalian Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Gel Filtration ◽  
Hydrolytic Activity ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Nuclear Fraction ◽  
Unidentified Species ◽  
Rf Values

We have previously reported the identification of a novel activity residing in the nuclear fraction of mammalian cells that selectively binds and hydrolyses deoxyribonucleoside triphosphates. Incubation of this protein with [alpha-32P]dATP leads to the appearance of a retarded band relative to free dATP when the reaction is analysed on non-denaturing polyacrylamide gels. We now show that the retarded species comprises the product of dATP hydrolysis (dADP or dAMP) bound to an as yet unidentified species. We have termed this complex the ‘product-nucleotide binding particle’ or PNBP*. Through a combination of continuous elution polyacrylamide-gel electrophoresis and gel-filtration chromatography, we demonstrate that the hydrolytic activity (dNTPase) is distinct from the radiolabelled species detected in gel-retardation experiments. T.l.c. confirms that the labelled product does not share RF values associated with a range of mono-, di- and tri-phosphate deoxyribonucleotide standards, and gel-filtration experiments suggest a molecular mass for PNBP* of between 2.5 and 3 kDa. The ability of purified PNBP* to retain its nucleotide ligand after a number of denaturing processes suggests that the ligand is covalently bound. The recovery of dNTPase activity from both gel-filtration and ion-exchange chromatography reveals that the as yet unliganded PNBP* (or a precursor form) is associated with the dNTPase enzyme as part of the active complex, prior to addition of dATP.

scholarly journals Characterization of Xylanolytic Enzymes in Clostridium cellulovorans: Expression of Xylanase Activity Dependent on Growth Substrates

2001 ◽  
Vol 183 (24) ◽  
pp. 7037-7043 ◽  
Author(s):  
Akihiko Kosugi ◽  
Koichiro Murashima ◽  
Roy H. Doi
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Xylanase Activity ◽  
Cellulolytic Bacterium ◽  
Clostridium Cellulovorans ◽  
Molecular Weights ◽  
Chromatography Analysis ◽  
Layer Chromatography ◽  
Activity Dependent

ABSTRACT Xylanase activity of Clostridium cellulovorans, an anaerobic, mesophilic, cellulolytic bacterium, was characterized. Most of the activity was secreted into the growth medium when the bacterium was grown on xylan. Furthermore, when the extracellular material was separated into cellulosomal and noncellulosomal fractions, the activity was present in both fractions. Each of these fractions contained at least two major and three minor xylanase activities. In both fractions, the pattern of xylan hydrolysis products was almost identical based on thin-layer chromatography analysis. The major xylanase activities in both fractions were associated with proteins with molecular weights of about 57,000 and 47,000 according to zymogram analyses, and the minor xylanases had molecular weights ranging from 45,000 to 28,000. High α-arabinofuranosidase activity was detected exclusively in the noncellulosomal fraction. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that cellulosomes derived from xylan-, cellobiose-, and cellulose-grown cultures had different subunit compositions. Also, when xylanase activity in the cellulosomes from the xylan-grown cultures was compared with that of cellobiose- and cellulose-grown cultures, the two major xylanases were dramatically increased in the presence of xylan. These results strongly indicated that C. cellulovorans is able to regulate the expression of xylanase activity and to vary the cellulosome composition depending on the growth substrate.

scholarly journals Synthesis and Characterization of Pumice-Supported nZVI for Removal of Copper from Waters

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Bilgehan Ilker Harman ◽  
Mesut Genisoglu
Keyword(s):  
Surface Chemistry ◽  
Natural Waters ◽  
Water Source ◽  
Synthesis And Characterization ◽  
Characterization Methods ◽  
Ph Values ◽  
Surface Areas ◽  
Removal Capacity ◽  
Wide Range

The target of this work was to study the synthesis and characterization of pumice-supported nanoscale zero-valent iron (nZVI) and the effectiveness of nZVI coated pumice to remove copper from water. The impacts of pumice dose, pumice surface chemistry, pH, and water source on copper removal were studied. Natural pumice particles were used as granular support media and coated with nZVI. Results of nZVI coated pumice characterization showed nZVI coated successfully on pumice surface being proved with characterization methods such as SEM-EDS, XPS, and XRF. nZVI coating overwhelmed the surface chemistry properties of the underlying pumice particles. Higher surface areas and more iron content were obtained in nZVI coated pumice. nZVI coating significantly increased copper uptake compared to uncoated particles. High removal capacity has been observed for all tested pH values. Control experiments indicated that nZVI bound on pumice surfaces is stable at pH values of typical natural waters. The nZVI coated pumice was found to be effective in removing copper from waters having a wide range of specific UV absorbance (SUVA) values. Overall, the results indicated that nZVI coated pumice particles are maybe alternative adsorbents to remove copper.

scholarly journals Antifungal Activity Toward Fusarium culmorum in Soluble Wheat Extracts

2000 ◽  
Vol 90 (6) ◽  
pp. 666-671 ◽  
Author(s):  
F. M. Doohan ◽  
A. Mentewab ◽  
P. Nicholson
Keyword(s):  
Antifungal Activity ◽  
Ammonium Sulfate ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Fusarium Culmorum ◽  
Gel Filtration ◽  
Head Blight ◽  
Inhibitory Compounds ◽  
Molecular Masses

This study investigated antifungal activity in soluble extracts from seed of a range of wheat cultivars differing in susceptibility to Fusarium head blight. Antifungal activity was assessed in terms of β-D-glucuronidase (GUS) activity of a Fusarium culmorum GUS transformant using a sensitive laboratory assay. Significant antifungal activity was detected in seed extracts from WEK0609, CM 820036, and Arina. Initial characterization of the Arina seed extract indicated that it contained antifungal proteinaceous compounds. The Arina extract yielded two (60 and 80%) ammonium sulfate fractions containing inhibitory compounds. Gel filtration chromatography and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis of antifungal fractions showed that the antifungal activities detected in the Arina 60 and 80% ammonium sulfate fractions were associated with putative proteinaceous compounds with apparent molecular masses of approximately 60 and 28 kDa, respectively.

Characterization of Laccases and Peroxidases from Wood-Rotting Fungi (Family Coprinaceae)

1998 ◽  
Vol 64 (5) ◽  
pp. 1601-1606 ◽  
Author(s):  
Marion Heinzkill ◽  
Lisbeth Bech ◽  
Torben Halkier ◽  
Palle Schneider ◽  
Timm Anke
Keyword(s):  
Sodium Azide ◽  
Polyacrylamide Gel Electrophoresis ◽  
Ligninolytic Enzymes ◽  
Coprinus Cinereus ◽  
Tetraacetic Acid ◽  
Alkaline Ph ◽  
Ph Values ◽  
Native Polyacrylamide Gel Electrophoresis ◽  
Diethyldithiocarbamic Acid

Panaeolus sphinctrinus, Panaeolus papilionaceus, and Coprinus friesii are described as producers of ligninolytic enzymes. P. papilionaceus andP. sphinctrinus both produced a laccase. In addition,P. sphinctrinus produced a manganese peroxidase. C. friesii secreted a laccase and two peroxidases similar to the peroxidase of Coprinus cinereus. The purified laccases and peroxidases were characterized by broad substrate specificities, significant enzyme activities at alkaline pH values, and remarkably high pH optima. The two peroxidases of C. friesii remained active at pH 7.0 and 60°C for up to 60 min of incubation. The peroxidases were inhibited by sodium azide and ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), whereas the laccases were inhibited by sodium azide andN,N-diethyldithiocarbamic acid. As determined by native polyacrylamide gel electrophoresis and isoelectric focusing, all three fungi produced laccase isoenzymes.

scholarly journals A multifunctional GH39 glycoside hydrolase from the anaerobic gut fungus Orpinomyces sp. strain C1A

2016 ◽  
Author(s):  
Jessica M Morrison ◽  
Mostafa S Elshahed ◽  
Noha Youssef
Keyword(s):  
Active Site ◽  
Glycoside Hydrolase ◽  
Biochemical Characterization ◽  
Xylanase Activity ◽  
Comparative Modeling ◽  
Glycoside Hydrolase Family ◽  
Wide Range ◽  
Promising Source ◽  
Substrate Competition

Background. The anaerobic gut fungi (phylum Neocallimastigomycota) represent a promising source of novel lignocellulolytic enzymes. Here, we report on the cloning, expression, and characterization of a glycoside hydrolase family 39 (GH39) enzyme (Bgxg1) that is highly transcribed by the anaerobic fungus Orpinomyces sp. strain C1A under different growth conditions. This represents the first study of a GH39-family enzyme from the anaerobic fungi. Methods. Using enzyme activity assays, we performed a biochemical characterization of Bgxg1 on a variety of substrates over a wide range of pH and temperature values to identify the optimal enzyme conditions and the specificity of the enzyme. In addition, substrate competition studies and comparative modeling efforts were completed. Results. Contrary to the narrow range of activities (β-xylosidase or α-L-iduronidase) observed in previously characterized GH39 enzymes, Bgxg1 is unique in that it is multifunctional, exhibiting strong β-xylosidase, β-glucosidase, β-galactosidase activities (11.5 ± 1.2, 73.4 ± 7.15, and 54.6 ± 2.26 U/mg, respectively) and a weak xylanase activity (10.8 ± 1.25 U/mg), strength determined as compared to previously characterized enzymes. Physiological characterization revealed that Bgxg1 is active over a wide range of pH (3-8, optimum 6) and temperatures (25-60°C, optimum 39°C), and possesses excellent temperature and thermal stability. Substrate competition assays suggest that all observed activities occur at a single active site. Using comparative modeling and bioinformatics approaches, we putatively identified ten amino acid differences between Bgxg1 and previously biochemically characterized GH39 β-xylosidases that we speculate could impact active site architecture, size, charge, and/or polarity. The putative contributions of these changes to the observed relaxed specificities in Bgxg1 are discussed. Discussion. Collectively, the unique capabilities and multi-functionality of Bgxg1 render it an excellent candidate for inclusion in enzyme cocktails mediating cellulose and hemicellulose saccharification from lignocellulosic biomass.

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